diff --git "a/deduped/dedup_0454.jsonl" "b/deduped/dedup_0454.jsonl"
new file mode 100644--- /dev/null
+++ "b/deduped/dedup_0454.jsonl"
@@ -0,0 +1,39 @@
+{"text": "Although there are numerous ways children with asthma and allergies can reduce attacks and live a more normal life, researchers at the 2004 annual meeting of the American Academy of Allergy, Asthma, and Immunology (AAAAI) in San Francisco said cigarette smoking in the home virtually negates those interventions. \u201cThe data are clearly there,\u201d said Robert Holzhauer, a clinical assistant professor of pediatrics at the University of Rochester School of Medicine and Dentistry. \u201cWe have unequivocal data to show that sidestream smoke is dangerous to people with asthma.\u201dIn one study presented at the March meeting, Holzhauer and colleagues identified Rochester schoolchildren aged 3\u20137 who had mild persistent to severe persistent asthma. Children were assigned randomly to school-based care groups. One group received daily inhaled corticosteroids\u2014a proven, effective treatment to prevent asthma attacks\u2014at school, while the other did not.Archives of Pediatrics and Adolescent Medicine.Children in the treated group had fewer attacks and school absences; their parents reported fewer worries about their children\u2019s health, work absences, and unexpected changes in plans. But if there was smoking in the home, those advantages were almost completely nullified. These findings have since been published in the May 2004 issue of In another presentation, Dennis Ownby, chief of allergy and immunology at the Medical College of Georgia, described his examination of the relationship between exposure to cats and dogs during the first year of life and the risk of allergy at age 6\u20137 years. He selected a birth cohort of 474 children, who were classified as having no exposure to cats or dogs during the first year of life, exposure to 1 cat or dog, or exposure to 2 or more cats or dogs.Children of nonsmoking parents were significantly less likely to have allergies if they were exposed to 2 or more cats or dogs; about 14% of these children were allergic, compared to 37.5% of children with 1 pet and 36.8% of children with no pets. But this benefit was not seen in children of smoking parents. \u201cThis research shows that cigarette smoking is not innocuous to young children,\u201d Ownby said. \u201cWe see evidence that it\u2019s affecting their immune system.\u201dHolzhauer said parents must be convinced of how important it is to stop smoking in the home if they have children with asthma. However the doctors stopped short of advocating persuasion through legislation. \u201cI think that if we were to report these parents to the authorities for child abuse, we would lose the children as patients,\u201d Holzhauer said.Rather, Kathleen Sheerin, a private-practice allergy specialist and chair of the AAAAI Public Education Committee, suggested that pediatricians and other health care professionals more strongly emphasize to parents the link between asthma, allergies, and smoking. She said, \u201cWe counsel parents to go to another room to smoke or to go outside if there is a child in the house.\u201d"}
+{"text": "Biagini JM, LeMasters GK, Ryan PH, Levin L, Reponen T, Bernstein DI, et al. 2006. Environmental risk factors of rhinitis in early infancy. Pediatr Allergy Immunol 17(4):278\u2013284.Many environmental exposures have been confirmed to affect children\u2019s respiratory health, but few have been studied in very young children. Now NIEHS grantees Grace K. LeMasters, Jocelyn M. Biagini, and their colleagues at the University of Cincinnati and the Cincinnati Children\u2019s Hospital Medical Center demonstrate for the first time the relationship between exposure to environmental tobacco smoke (ETS) and allergy in infants.About one-fifth of all American adults smoke cigarettes, resulting in about 43% of children being exposed to ETS at home. ETS exposure, along with mold exposure, has been documented as a risk factor for health problems such as wheezing, asthma, and otitis media in both children and adults.In the current study, the researchers observed the effects of ETS and indoor mold exposure on the development of rhinitis and symptoms such as nasal blockage, sneezing, and nasal itching in a cohort of 633 infants under the age of 1 enrolled in the Cincinnati Childhood Allergen and Air Pollution Study. They used interviewer-administered questionnaires to collect demographics and information on smoking habits, family health history, and other covariates. They also analyzed any upper respiratory symptoms of the infants recorded by the parents in a monthly diary. In addition, they performed a skin-prick test on the parents and the infants (at approximately 12 months of age) to test for sensitivity to at least 1 of 15 airborne allergens.The investigators found that exposure to ETS increased an infant\u2019s risk of developing allergic rhinitis by almost threefold. They also found that exposure to mold in the home was associated with increased risk of upper respiratory infections but not allergy, which differed from previously reported research in older children and adults.Other findings included a protective effect of having older siblings in the home. Infants with at least one older sibling were less likely to have allergic rhinitis by their first birthday. This finding supports the hygiene hypothesis, a theory that exposure to infectious agents early in life may decrease the risk for allergic diseases such as asthma later in life. Presumably, by having older siblings these infants were exposed to a wider variety of viruses and bacteria, causing their immune systems to develop in a way that decreased the risk of allergy.The authors conclude that further research is necessary to confirm their results. Continued research is also needed to determine the components of cigarette smoke that cause these health effects, and to ascertain the role of possible gene\u2013environment interactions."}
+{"text": "To determine whether a pass/fail system is more appropriate for medical education instead of a grade-based system, a survey of medical students and faculty members of Hallym University, Korea, was taken. A questionnaire was delivered to 54 junior students and 36 faculty members from a medical school in Korea and analyzed. Of these participants, 37.7% of students and 36.1% of faculty agreed to the pass/fail system, while 28.3% of students and 52.8% of faculty objected to it. The most frequent reason for objection was the potential decrease in learning achievement. A pass/fail system should be considered after persuasion of the students and faculty to think positively of this system. In some instances, some classes are evaluated by the pass/fail system (P/F system). However, some medical schools outside of Korea have adopted the numeric grading system (a continuous numeric system), letter grading system  and 36 faculty members  in a medical school. The average duration of education of the faculty was 9.77\u00b15.6 years. Initially, the agreement or objection to the system was noted, and potential effects of the P/F system were then presented. Open opinions were also gathered. The content of the questionnaire is described in Of all survey participants, 37.7% of students and 36.1% of faculty agreed with the P/F system, while 28.3% of students and 52.8% of faculty objected to this system. Males (44.2%) agreed more than females (18.8%) . StudentHalf of all faculty, however, worried about the potential decrease in learning achievement. Furthermore, 79.6% of students and 66.7% of faculty worried about the difficulty in setting the cutoff score for a \"pass\" .In the open-ended questions, the reasons for agreement by students were the possibility of in-depth study and a cooperative environment among students without the worry over grades. Many faculty agreed, since the ultimate goal of medical education is the sufficient achievement of minimum requirements to be a doctor.Reasons for objection by students and faculty were the lack of motivation to study and the likely decrease in learning achievement. Students had qualms over setting too high a cutoff score and the greater potential for failure in courses. The faculty were worried about the resistance students might generate if the cutoff score was so high that massive failure occurred. Also, faculty were afraid that the decision to award scholarships may be difficult if it was based solely on a binary pass/fail system.Although the P/F system has some merits, there are still many obstacles to this system, particularly with regard to a potential decrease in learning achievement and a lack of motivation for study. Although reports on use of the P/F system in the United States have been positive, it is uncertain that these results can be recapitulated in Korea , 2.In our survey, positive responses to P/F system were based on anticipated cooperative work. However, negative responses developed over worries that learning achievement will decline. These results suggest that while the study environment may be improved, the total time spent learning may be decreased. Also, students said that they could study harder if there was no stress related to high grade achievement, a rationale that the faculty did not believe.Concerns about cutoff score determination by both students and faculty were substantial. A reasonable selection of the cutoff score should be considered, and should be reviewed to adapt the modified Angoff or Bookmark method for setting such scores, if necessary.Meanwhile, compensation or motivation for high performance in students is another difficult task. The faculty mentioned new subject criteria for scholarship awards. A previous report suggested that if numeric grades were used for evaluation, learning achievement can be anticipated and much information can be deduced, a result that was not possible in the P/F system .Program directors for recruitment of medical residents prefer the numeric grade system , 6. TherStill, there are many problems that need solved in implementing the P/F system. Team-learning or group study and self-directed study should be introduced in the premed period in order to provide a familiar environment for the P/F system. The setting of the cutoff score and a valid evaluation method should be prepared. The motivation of high-performance students should also be considered. Grading systems, such as the recruitment system of internship by a hospital, are sometimes necessary, and it will be challenging to report learning achievement in the P/F system in such a recruitment process.Although there are some difficulties at hand, the P/F system evidently has some merit, such as the mitigation of stress, as well as the motivation for cooperative work. This system should be considered in medical schools in Korea after considerable support by students and faculty to think positively of this system and after providing a suitable environment in which to establish it ."}
+{"text": "L. casei DN-114 001 (Lc) could decrease the severity of intestinal inflammation in a murine model of IBD.Probiotic bacteria can be used for the prevention and treatment of human inflammatory diseases including inflammatory bowel diseases (IBD). However, the nature of active components and exact mechanisms of this beneficial effects have not been fully elucidated. Our aim was to investigate if lysate of probiotic bacterium in vivo and in vitro, we evaluated intestinal permeability using the FITC-labeled dextran method and analysed tight junction proteins expression by immunofluorescence and PCR. We also measured CD4+FoxP3+ regulatory T cells proportion by FACS analysis, microbiota composition by pyrosequencing, and local cytokine production by ELISA. Lc leads to a significant protection against increased intestinal permeability and barrier dysfunction shown by preserved ZO-1 expression. We found that the Lc treatment increases the numbers of CD4+FoxP3+ regulatory T cells in mesenteric lymph nodes (MLN), decreases production of pro-inflammatory cytokines TNF-\u03b1 and IFN-\u03b3, and anti-inflammatory IL-10 in Peyer's patches and large intestine, and changes the gut microbiota composition. Moreover, Lc treatment prevents lipopolysaccharide-induced TNF-\u03b1 expression in RAW 264.7 cell line by down-regulating the NF-\u03baB signaling pathway.The preventive effect of oral administration of Lc significantly reduces the severity of acute dextran sulfate sodium (DSS) colitis in BALB/c but not in SCID mice. In order to analyze how this beneficial effect interferes with well-known phases of intestinal inflammation pathogenesis Our study provided evidence that even non-living probiotic bacteria can prevent the development of severe forms of intestinal inflammation by strengthening the integrity of intestinal barrier and modulation of gut microenvironment. Inflammatory bowel diseases (IBD), such as Crohn's disease and ulcerative colitis, are severe chronic inflammatory illnesses of the gastrointestinal tract. Although their etiology and pathogenesis are not fully understood, it is generally accepted, that the inflammation is a result of an aberrant immune response to antigens of resident gut microbiota in genetically susceptible individuals When administered in adequate amounts, probiotics, defined as live microorganisms, confer a health benefit to the host Colitis induced by dextran sulfate sodium (DSS) is a well established and reliable model of IBD because its clinical features resemble the ulcerative colitis L. casei DN-114 001 has been found to reduce the duration and severity of diarrhea and common infectious diseases in children E. coli strain The most intensively studied and used probiotic bacteria are lactobacilli L. casei DN-114001 protects mice from subsequent acute DSS-induced colitis in BALB/c mice In our previous study, we have shown that the preventive treatment with live probiotic bacterium Lactobacillus casei DN-114 001 , Lactobacillus plantarum CCDM 185 , were grown in an anaerobic chamber in De Man, Rogosa, and Sharpe broth  at 37\u00b0C until the cultures were in the late log phase of growth. Both lactobacilli were harvested by centrifugation  and washed twice with sterile phosphate-buffered saline (PBS). After the treatment with the French press, lactobacilli were freeze-dried and diluted to a working concentration of 30 g/l. In order to kill all remaining viable bacteria, the lysate was heated to 60\u00b0C for 30 min and the sterility of all components was verified by both aerobic and anaerobic 48 hours cultivation before administration.Ethics statement: All animal experiments were approved by the Animal Care and Use Committee of the Institute of Microbiology, Academy of Sciences of the Czech Republic, approval ID: 10/2005, 94/2008 and 211/2009. Female BALB/c mice (8\u201312 weeks old) or severe combined immunodeficient mice BALB/cJHanHsd-SCID (SCID) were obtained from a breeding colony at the Institute of Physiology  or at the Institute of Microbiology , respectively, and reared under conventional conditions.8 CFU of heat killed bacteria, by gavage. To reduce proteolytic activity in the gut, the Lc components were co-administered with 1 mg of soybean trypsin inhibitor  dissolved in 50 \u00b5l of 0.15 M sodium bicarbonate buffer (pH 8.0). Control mice were given only sterile PBS with soybean trypsin inhibitor in bicarbonate buffer. The administration of lysates was repeated every 7 days for a total number of 4 doses . Acute colitis was induced 7 days later by 3% (wt/v) DSS  dissolved in tap water for 7 days, and on the last day of the experiment the colitis was evaluated by using a clinical activity score, colon length, and the histological scoring system as described previously We administered 1.5 mg of Lc in 50 \u00b5l of sterile PBS, i.e. 6\u00d7102, Tecan Group Ltd., M\u00e4nnedorf, Switzerland) with an excitation wavelength of 483 nm and an emission wavelength of 525 nm using serially diluted samples of the marker as standard.The intestinal permeability was measured by determining the amount of FITC-dextran in blood after it was orally administered as described previously Segments of colon and terminal ileum were frozen in liquid nitrogen immediately after removal and stored at \u221280\u00b0C until used. Frozen sections (6 \u00b5m) were mounted on the poly-L-lysine-coated slides. Then the slides were dried and fixed in 4% buffered paraformaldehyde (pH 7.4) for 10 min at room temperature. Fixed sections were washed in PBS and blocked with 2% donkey serum (Sigma-Aldrich) in PBS for 20 min at room temperature. The slides were incubated with the rabbit polyclonal anti-mouse ZO-1 or occludin antibodies  overnight at 4\u00b0C. The negative controls were performed similarly using 1% bovine serum albumin (BSA) in PBS instead of primary antibody. After washing, the sections were incubated with donkey anti-rabbit antibody conjugated either with Texas Red or with DyLight 488 fluorochrome . Nuclei were counterstained using DAPI  stain. Finally, the sections were mounted in Vectashield mounting medium for fluorescence  and viewed with a fluorescence microscope Olympus AX-70 .Intestinal mucosa from terminal ileum and colon was placed in RNAlater stabilization reagent . Total messenger RNA (mRNA) was extracted by using the RNeasy Mini isolation kit (QIAGEN GmbH) following the manufacturer's instructions. RNA integrity was determined by gel electrophoresis in 1.5% agarose gel stained with ethidium bromide. The purity of the RNA was assessed by the ratio of absorbance at 260 and 280 nm. RNA purity was within a range of 2.0\u20132.1. The total RNA concentration was estimated by spectrophotometric measurements at 260 nm assuming that 40 \u00b5g of RNA per mililitre equal one absorbance unit. Real time PCR was performed as described previously 2 in complete RPMI medium with 10% fetal bovine serum  and 100,000 U/l penicillin, 100 mg/l streptomycin (Sigma-Aldrich), as described previously Sections of Peyer's patches (PP), ileum, cecum, and colon were taken from every mouse. The intestines were then opened longitudinally, washed in ice cold PBS containing antibiotics and cultivated for 48 hours at 37\u00b0C and 5% COThe samples were processed as described above . Gene expression assays for IL-10, IL-6, TNF-\u03b1 and \u03b2-actin were all purchased by Applied Biosystems, Foster City, CA, USA.Sera and small intestine washings were collected for specific antibody evaluation. Gut washings were obtained by flushing the content of isolated small intestine with 2 ml of sterile PBS containing a mixture of proteinase inhibitors (Sigma-Aldrich). The samples were then vortexed and centrifuged at 4\u00b0C, and the supernatant was collected and stored at \u221280\u00b0C until analysis. Indirect ELISA, optimized in our laboratory as previously described regs using FoxP3 Staining Set  with fluorochrome-labeled anti-mouse mAbs: CD4-Qdot\u00ae 605 , CD8-BD Horizon\u2122 V500 , CD3-FITC and FoxP3-Phycoerythrin (both from eBioscience) according to the manufacturer's recommendation.Single-cell suspensions of spleens, MLNs and PPs were prepared and stained for TRAW 264.7 cells were cultivated and stained for IL-7R-Alexa647 , CD206-PE , CD-11c-NC625 and F4/80-APC780 (both from eBioscience). Hoechst 33342 (Sigma-Aldrich) was used to determine cell viability. Flow cytometric analysis was performed on LSRII (BD Biosciences), and the data was analyzed using FlowJo software .2 in Dulbecco's modified Eagle's medium  containing 10% heat-inactivated fetal bovine serum (Biochrom AG), 4.5 g/l glucose, 1.5 g/l sodium bicarbonate, 4 mM glutamine (Institute of Molecular Genetics AS CR), 100,000 U/l penicillin and 100 mg/l streptomycin (both Sigma-Aldrich). The cells were cultured together with Lc, lysate of L. plantarum (Lp) or sterile PBS in the presence or absence of LPS . After cultivation, the concentration of TNF-\u03b1 in the supernatant was measured with ELISA (Invitrogen). The nuclear proteins were extracted from stimulated RAW264.7 cells by a nuclear extract kit  and used to quantify the DNA binding activity of p65 subunit using the TransAM NF-\u03baB family transcription factor assay kit (Active Motif). In NF-\u03baB assay, only the concentration with the strongest immunomodulatory properties of Lc was used, i.e. 10 pg/l. All assays were performed according to the manufacturer's recommendation.The LPS-activated macrophage cell line  was cultivated in the presence of different concentrations of bacterial lysate, as previously described Stool samples from PBS or Lc-treated mice, on day 0, 28 (just before DSS administration) and 35 (the last day of experiment) were collected. Total DNA from these samples was then isolated with ZR Fecal DNA Kit\u2122  according to the manufacturer's recommendation.2, 0.2 mmol/l of dNTPs, 1\u00d7 PCR buffer and 1 U platinum TAQ DNA polymerase (Invitrogen) and 0.40 \u00b5mol/l of forward modified primer consisting of 454 adaptor A , unique 10-base tag sequence  and universal broad-range bacterial primer 5\u2032-AYTGGGYDTAAAGNG and 0.40 \u00b5mol/l of reverse primer consisting of adaptor B (5\u2032-CCATCTCATCCCTGCGTGTCTCCGACTCAG-3\u2032) and universal primer TACNVGGGTATCTAATCC. PCR conditions were as follows: 1\u00d7: 95\u00b0C, 3 min; 35\u00d7: 94\u00b0C, 50 sec; 40\u00b0C, 30 sec; 72\u00b0C, 60 sec; 1\u00d7: 72\u00b0C, 5 min and final hold at 4\u00b0C. The length of PCR product was checked on the agarose gel electrophoresis. PCR product was subsequently purified using magnetic beads . Concentration was measured on Qubit fluorometer . Equimolar amounts of PCR product from each sample were used for unidirectional 454 FLX amplicon pyrosequencing using LIB-L emPCR kits following the manufacturer's protocols .DNA was subsequently gel-purified and PCR was performed in triplicate for each primer pair, and pooled to minimize random PCR bias. The reaction mixture contained 1 \u00b5l of DNA (10 ng/\u00b5l), 1.5 mmol/l MgClFlowgrams were processed using amplicon analysis option in data processing software from Roche. The sequencing resulted in 161551 overall number of reads. Quality trimmed sequences obtained from the FLX sequencing run were processed using RDP pyrosequencing pipeline. Beforehand, data files were depleted of chimeras by Black Box chimera Checker One-way analysis of variance (ANOVA) with Dennett's multiple comparison test was used to compare multiple experimental groups with the control group. Differences between two groups were evaluated using an unpaired two-tailed Student's t-test and deviation of values from hypothetical mean were calculated by one sample t-test. The data is presented as the mean \u00b1 standard deviation (SD) unless stated otherwise and differences were considered statistically significant at P\u22640.05. GraphPad Prism statistical software  was used for analyses.L. casei DN-114 001 attenuates the severity of acute experimental colitis In our previous study we showed that oral treatment with Increased intestinal permeability caused by impairment of the gut barrier function drives the pathogenesis of intestinal inflammation in both DSS-induced colitis and human IBD Barnesiella, from the Bacteroidetes phylum, one of the most abundant phylum in intestinal microbiota. The next most abundant genus with very well described capability to ameliorate intestinal inflammation Lactobacillus increased in abundance after exposure to DSS and the Lc treatment. This increase in abundance was not observed in the control PBS group. The Bacteroides, known to be increased during DSS-induced colitis, proliferated after intestinal inflammation, was induced in Lc and PBS treated groups. Moreover, there is an increase in the biggest group of genera from Clostridium cluster: butyrate producing Butyricicoccus, Coprococcus and Anaerostipes. Butyrate is crucial for energy homeostasis of mammalian colonocytes, capable to prevent their autophagy Changes in the intestinal gut microbial ecology are expected to be associated with the state of disease and could be influenced by probiotic treatment Changes in cytokine microenvironment in the gut mucosa can influence the mucosal immune response to luminal antigens leading to the decrease of intestinal inflammation. Therefore, we investigated if the protective effect of Lc is associated with modifications in inflammatory response in the key compartments of the gut. We cultivated tissues from four distinct parts of the gut of either DSS/PBS or DSS/Lc-treated mice for 48 h and then measured the cytokines in supernatants by ELISA. We found that pretreatment with DSS/Lc decreased the production of pro-inflammatory cytokines  and anti-inflammatory cytokine IL-10 in PPs, cecum and colon as compared to DSS/PBS-treated mice . These rregs , we analyzed the changes in CD4+FoxP3+ Tregs in the spleen, MLNs and PPs of DSS/PBS-, DSS/Lc-treated mice. We found a statistically significant increase in Tregs in MLN of DSS/Lc-treated mice as compared to DSS/PBS-treated mice. There were no statistically significant differences in the numbers of Tregs in spleen and PPs between these groups  can enter and after the interaction with gut-committed cells (\u03b14\u03b27-integrin expressing cells) from intestine became TregsThe lack of protective effect in SCID mice suggests that mechanisms of adaptive immunity are essential for the beneficial effect of Lc. We did not find any changes neither in Lc-specific serum IgA, IgG and IgM, nor in gut SIgA during our experiments (data not shown), so we analyzed another mechanism executed by adaptive immune response, oral tolerance. Major role in this mechanism is played by TL. casei and other probiotics can strengthen the gut barrier function E. coli Nissle 1917 provided protection against DSS-mediated leakiness and was capable to produce specific up-regulation of ZO-1 expression in the intestinal epithelial barrier L. casei, prevents changes in expression and distribution of tight junction proteins ZO-1 and occludin The intestinal barrier prevents viable enteric bacteria and the microbiota derived components from excessive interaction with the immune system. Here, we demonstrated that increase in intestinal permeability and the decrease in local ZO-1 expression, typical for DSS-treated mice, are both significantly improved by oral application of Lc. These results are in agreement with several studies showing that in vivo, which can shape the gut microbiota composition long time after the probiotic therapy has ended Several studies showed that there is a marked difference in the gut microbiota composition in IBD patients (\u201cdysbiosis\u201d) as compared to healthy individuals. These changes in microbiota composition, or presence of certain microbial species with increased virulence, cause or perpetuate the intestinal inflammation in IBD Bacteroides genus after induction of intestinal inflammation as shown previously Lactobacillus genus which suggests that treatment with Lc promotes this genus among others. This effect could be caused by formation of niche ideal to lactobacilli. These and other differences in microbiota could be also explained by decreased inflammation in Lc-treated mice mediated by differing immunological mechanisms.By using the pyrosequencing technique we observed an increase in L. casei can downregulate the pro-inflammatory mediators in the lamina propria of inflamed mucosa from Crohn's disease patients during ex vivo cultivation The cytokines produced in the gut mucosa greatly influence the resulting immunological outcome. The production of anti-inflammatory cytokines induces the mucosal unresponsiveness and tolerance and high levels of pro-inflammatory cytokines induce protective immune response and inflammation in vitro. We found that Lc, but not Lp, decreases the production of TNF-\u03b1, and the activation of NF-\u03baB cascade and polarizes macrophages to M2 phenotype, suggesting a possible direct effect of Lc on the cells of the innate immunity. It is not excluded that negative regulators are involved in beneficial anti-inflammatory effects of probiotics Acute DSS colitis is believed to be driven initially by innate immunity mechanisms and, in particular, the role of macrophages has been proposed L. casei DN-114 001 can protect from induction of intestinal inflammation, thus confer a health benefit for the host. This is achieved by mechanisms that comprise: a) improvement in the gut barrier function, b) correction of the dysbiosis, and c) modulation of the mucosal immune response. These complex immunomodulatory properties of bacterial lysates may lead to the development of new therapeutic approaches for treatment of chronic intestinal inflammation. Moreover, oral administration of sterile bacteria, in contrast to live bacteria, may be safer in severely ill or immunocompromised patients.In conclusion, our data provide evidence that even lysate of"}
+{"text": "Data concerning the effects of phthalate exposure on preterm delivery and fetal growth are limited in humans. In this paper, we assessed the relationship between 15 phthalate levels in cord blood and preterm delivery and fetal growth parameters in 207 Chinese women going into labor. Exposure to phthalates except DCHP was associated with gestational age reduction and preterm delivery (p<0.05). There were associations between phthalates and fetal growth parameters, many of which disappeared when analyses were adjusted for gestational age, especially in male infants . This study indicates that prenatal exposure to phthalates is associated with younger gestational age and preterm delivery. Also, phthalate exposure may adversely affect fetal growth parameters via gestational age reduction and preterm delivery with a significant gender effect. Phthalate Acid Esters (PAEs) are a class of synthetic chemicals which are produced in large volumes and used in a wide variety of industrial and common household products. High molecular weight phthalates (HWMP) act as additives that impart flexibility to vinyl resins. Low molecular weight phthalates (LWMP) have different uses, including in personal care products, cosmetics, food conveyor belts, artificial leather, automotive trim, traffic cones, latex adhesives, cellulose plastics and solvent for dyes In our previous studies In this study we investigated the association between phthalate levels in cord blood and fetal growth, pregnancy complications, preterm delivery and other fetal growth parameters.This study was approved by the Ethics Committee of the Third Military Medical University. All participants, the children participant were presented by their parents and signed at the same consent, provide their written informed consent to participate in this study. Some consents were signed by the husbands on behalf of their wife. This consent procedure was approved by the ethics committees.Study subjects included Chinese women residing in Chongqing (Southwest China) for at least two years, who delivered at Southwest Hospital in Chongqing between October 2011 and September 2012. Volunteers had no history of alcohol or tobacco use, were 18\u201335 years old, had undergone prenatal examination at the department of gynecology and obstetrics, Southwest Hospital, and had no family or personal history of occupational exposure to phthalates. Gestational age was determined by the last menstrual period. We recruited 207 consecutive volunteers meeting the inclusion/exclusion criteria mentioned above, 33 of which had preterm delivery . A questionnaire was administered to participants after labor to obtain information on socio-demographic characteristics, medical history and lifestyle factors. Delivery characteristics and fetal growth parameters were obtained from the perinatal database of Southwest Hospital in Chongqing where the patients delivered. This included presence of premature rupture of membranes (PROM), chorioamnionitis, intrahepatic cholestasis of pregnancy (ICP), placenta previa, pregnancy-induced hypertension syndrome , gestational diabetes mellitus (GDM), abruption placentae, intravenous infusions therapy history (within the last week before labor), gestational age, birth weight, birth length, head circumference (HC), biparietal diameter (BPD), abdominal circumference (AC) and femur length (FL). Five ml of cord blood were obtained from each subject within 10 min of delivery and stored in a heparinized glass container at \u221280\u00b0C until it was analyzed.A certified standard mixture of 15 phthalates  including Dimethyl phthalate (DMP), Diethyl phthalate (DEP), Diisobutyl phthalate (DIBP), Dibutyl phthalate (DBP), bis (2-methoxyethyl) phthalate (DMEP), bis (4-methyl-2-pentyl) phthalate (BMPP), bis (2-ethoxyethyl) phthalate (DEEP), Diamyl phthalate (DPP), Dihexyl phthalate (DNHP), Benzyl butyl phthalate (BBP), bis (2-n- butoxyethyl) phthalate (DBEP), Dicyclohexyl phthalate (DCHP), bis (2-ethyl hexyl) phthalate (DEHP), Di-n-octyl phthalate (DNOP) and Dinonyl phthalate (DNP) was used as a calibration standard.Phthalates in cord blood were extracted according to the method described by Hogberg et al All statistical analyses were performed with SPSS for windows version 18.0. Independent sample T-tests and Chi-square tests were used to analyze the characteristics of pregnant women in the preterm and term delivery groups and their offspring. Linear regression and independent sample T-test were used to estimate the association between phthalates and gestational age and fetal growth parameters, and binary logistic regression and Chi-square test were used to estimate the association between phthalates and preterm delivery. Normal distribution was confirmed in all tests performed by one sample Kolmogorov-Smirnov Test.For linear regression and binary logistic regression analyses, we replaced the concentration below the limits of detection (LOD) by LOD/\u221a2 when the detection rate of the phthalate was more than 60% and transformed them into Ln-transformation for analyses. When the detection rate was less than 60%, the concentrations were converted into categorical variables by replacing the values for 1 (detected) or 0 (undetected).For independent sample T-test and Chi-square test analyses, pregnant women were graded from low cord blood PAEs concentration to high cord blood PAEs concentration by the phthalates level (dividing at the median when the detection rate was more than 50% and at the LOD when the detection rate was less than 50%).p<0.05, p<0.05, p<0.01, p<0.05, Cord blood levels of all 15 phthalates predicted gestational age reduction p<0.05,  and pretp<0.05, p<0.05, p<0.05, p<0.05, p<0.05, p<0.05, p<0.05, p<0.05, p<0.05, p<0.05, p>0.05, In female infants, 14 phthalates except DCHP were significantly associated with decreased birth weight p<0.05, ; 13 phthp<0.05, p<0.05, p<0.05, p<0.05, p<0.05, p<0.05, p<0.05, p>0.05, In male infants, 13 phthalates  were significantly associated with decreased birth weight p<0.05, ; 13 phthp>0.05, data not show).Other adjusting factors we tested including maternal age, BMI, frequency of prenatal examination and pregnancy history did not affect the associations between PAEs exposure and fetal growth parameters in female or male infants .In our previous studies, DBP levels in cord blood were more significantly associated with birth weight than its levels in venous blood, breast milk and urine. Based on this we hypothesized that phthalate levels in cord blood, which avoid the effect of the placental barrier, would be associated with fetal development.All 15 phthalates were detected in cord blood samples, indicating that exposure to phthalates was very prevalent among pregnant women in Chongqing . AlthougIn our previous study we reported that DBP was generally present in pregnant women in Chongqing In our study population, all phthalates except DCHP were associated with decreased gestational age and preterm delivery.Certain medical equipment is known to contain phthalates, and phthalate exposure is associated with intensive hospital care that occurs during preterm parturition. In china, nearly all of the preterm women underwent intravenous infusion therapy to prolong pregnancy Prenatal phthalate exposure may reduce gestational age and cause preterm delivery through other mechanisms such as activation of peroxisome proliferator activated receptor (PPARs). PPARs, a ligand-activated transcription factor that is associated with preterm labor In our study population, higher phthalate concentrations in cord blood were associated with the reduction of birth weight, birth length, AC, FL, BPD and HC, both in male and female infants. The majority of these associations lost significance after adjusting for gestational age. Moreover, gestational age were also associated with fetal growth parameters. This suggests that the adverse effects on fetal growth parameters caused by phthalates may be partially dependent on the gestational age reduction and are not a direct effect of phthalates. Also, after adjusting for gestational age, the effects phthalates were different between genders. While DNP, DCHP, DBP, DEHP, DEEP, DNHP, DEP and BBP appear to have an effect on female infants, only DNEP and DPP had an effect on male infants. These differences were also present when the term group was analyzed alone with DNP, DCHP, DBP, DEEP, DNHP, DEP and BBP affecting female infants, and DIBP and DPP affecting male infants. This data suggests that gender differences may exist in the associations between phthalates exposure and fetal growth parameters.In our study, we found an association between phthalate levels in cord blood and gestational duration and preterm delivery. Also, exposure to some phthalates was associated with adverse effects on fetal growth parameters which may partially be due to a shortened gestational duration and appears to be modulated by gender.Figure S1Association between cord blood PAEs levels and the incidence of preterm delivery in all pregnant women. The low cord blood PAEs concentration group and high cord blood PAEs concentration group were divided at the median phthalate level when the detection rate was more than 50% or at the LOD when the detection rate was less than 50%. **p<0.01.(TIF)Click here for additional data file.Figure S2The proportion of high cord blood PAEs levels in intravenous infusions therapy group and non- intravenous infusions therapy group. The high cord blood PAEs levels defines which phthalate concentration in cord blood were higher than the median phthalate level when the detection rate was more than 50% or the LOD when the detection rate was less than 50%. **p<0.01. *p<0.05.(TIF)Click here for additional data file."}
+{"text": "We report the case of an immunocompetent patient with an isolated tuberculoma of the liver, which was diagnosed by percutaneous US-guided liver biopsy. The patient received an antitubercular therapy, and there has been no relapse to date. Hepatic tuberculosis is an uncommon form of extrapulmonary tuberculosis. It is usually a disseminated disease associated with miliary tuberculosis, which is one of the most characteristic manifestations of tuberculosis (TB). Localized tuberculosis of the liver in the form of macronodular tuberculoma or an abscess is rare . HoweverA 44-year-old man with a history of benign gastroesophageal reflux disease (GERD), with irregular use of IPP, visited the emergency department with the chief complaint of intermittent fever for 3 months. Body weight loss (11 kg/month), abdominal fullness, poor appetite, and general malaise were also observed. He had no significant history of tuberculosis, and no relevant familial history. On direct questioning he admitted to have night sweats and abdominal pain. The patient said that he had received outpatient treatment followed by oral antibiotics. Unfortunately, his condition worsened with intermittent chills and body weight loss, so he was referred to our hospital. On admission, we found a mildly cachetic man. No jaundice was noted by examination of the palpebral or bulbar conjunctiva, but he was found to be pyrexial (38.7\u00b0C on admission). His heart rate was 91 beats/min, blood pressure was 135/89 mmHg, respiratory rate was 18 breaths/min, and oxygen saturation was 100% under normal conditions. Physical examination revealed mild right hypochondrial knocking pain with right hepatomegaly. Palpation did not reveal any abnormalities in the gallbladder or spleen, no abnormality was noted by chest percussion and there was no lymphadenopathy.The results of laboratory studies showed the following: an elevated C reactive protein (CRP) level of 152 . a mildly elevated erythrocyte sedimentation rate (ESR), and hyponatremia at 121mEq/L ; a normal potassium value at 4.16 mEq/L ; white blood cell count (WBC) of 8500/mL ; hemoglobin, 10.9 g/dL , platelet count, 280,000/\u03bcL ; aspartate aminotransferase (AST), 18 IU/L ; alanine aminotransferase (ALT), 25 IU/L ; total bilirubin, 2.2 mg/dL ; elevated creatinine, 3.2 mg/dL ; blood urea nitrogen (BUN), 21 mg/dL . Prothrombin time was within normal limits. Further evaluations, including tests for human immunodeficiency virus, carcinoembryonic antigen, cancer antigen 19-9, \u03b1-fetoprotein, prostate-specific antigen, serologic diagnosis of hydatid disease and amoebic hemagglutination were all negative. Several sets of blood and sputum cultures were negative for bacteria, fungus, and acid-fast bacilli. Abdominal ultrasound revealed marked hepatomegaly with a tumor with an unclear border in the VIII subsegment of the liver consisting partially of hypoechoic mass, with the hyperechoic area measuring 7 cm. No ascites or adenopathy was seen. Similarly, Abdominal computed tomography (CT) revealed a low-density lesion approximately 7 cm in diameter at the SVIII segment, showing various fat concentrations. Chest X-ray examination was normal . TherefoTuberculosis is a monumental health problem in the developing world and it remains a healthcare challenge in the developed world owing to immigration from endemic areas, increased prevalence of immunosuppression, and emergence of multidrug- and extensively drug-resistant strains of Mycobacterium tuberculosis . AbdominLiver TB is considered very rare among abdominal TB patients. Usually it is associated with foci of infection in the lungs or gastrointestinal tract. TB spreads to the liver through the hepatic artery, the portal system and the lymphatic system. Liver TB commonly involves the hepatic parenchyma and sometimes the biliary tree. The presentations of liver TB are divided into three types: Miliary hepatic TB which is haematogenously disseminated from the lungs via hepatic artery, and it is considerered the most common type of hepatic TB ; BiliaryNodular hepatic TB as in our case, which is most unusual and almost all publications have been single case reports. The localized form of tuberculosis of the liver, termed also a tuberculoma, was first described over a century ago by Bristowe 1858) who found the liver to be involved in 12 out of 167 cases of tuberculous ulceration of the intestines 58 who fo. ImagingOn ultrasonography, hepatic tuberculoma usually shows well marginated hypoechoic lesion with or without calcifications , and hypMycobacterium tuberculosis is increasing and may improve sensitivity rates [Otherwise, the rate of accurate pretreatment diagnosis by guided percutaneous liver biopsy has been reported to be low, and the presence of tubercle bacilli in the biopsy sample is rarely reported. Thus, the reliability of needle biopsy as a diagnostic method seems uncertain . Howeverty rates . For mosty rates . Hepaticty rates . Cumulatty rates , 9. The ty rates . Ultrasoty rates .Localized tuberculosis of the liver in the form of macronodular tuberculoma or an abscess is rare. However, because of the increasing incidence of pulmonary tuberculosis, clinicians should be aware of the possibility of tuberculous infection in all patients who have non-resolving liver abscesses, particularly in regions with high prevalence, such as Morocco. Since clinical features are protean and mimic neoplastic and infective hepatic disorders, the diagnosis requires a high index of suspicion. We suggest that tuberculous liver abscess should be considered in patients not showing typical features or who fail to respond to antibiotics. The demonstration of granulomas on liver biopsy remains the most sensitive diagnostic procedure, As for other forms of extra pulmonary tuberculosis, hepatic tuberculosis is a potentially curable disease. Good results have been obtained with four drug regimens without any added risk of hepatotoxicity as in our case."}
+{"text": "The frequency-dependent acoustic attenuation obeyed a power law; for native gel wax, it varied from 0.71 dB/cm at 3 MHz to 9.93 dB/cm at 12 MHz. The chosen oil-based inks, which have different optical absorption spectra in the range of 400 to 900 nm, were found to have good photostability under pulsed illumination with photoacoustic excitation light. Optically heterogeneous phantoms that comprised of inclusions with different concentrations of carbon black and coloured inks were fabricated, and multispectral photoacoustic imaging was performed with an optical parametric oscillator and a planar Fabry-P\u00e9rot sensor. We conclude that gel wax is well suited as a TMM for multispectral photoacoustic imaging.Tissue-mimicking phantoms are widely used for the calibration, evaluation and standardisation of medical imaging systems, and for clinical training. For photoacoustic imaging, tissue-mimicking materials (TMMs) that have tuneable optical and acoustic properties, high stability, and mechanical robustness are highly desired. In this study, gel wax is introduced as a TMM that satisfies these criteria for developing photoacoustic imaging phantoms. The reduced scattering and optical absorption coefficients were independently tuned with the addition of TiO Tissue-mimicking phantoms are widely used for evaluating new medical imaging systems and for training clinical practitioners. Traditionally, phantoms for optical modalities have been developed separately from those for ultrasonic modalities. For optical imaging modalities such as diffuse optical tomography and fluorescence imaging, controlling optical properties such as optical scattering and absorption coefficients has been important. Likewise, for ultrasound imaging, speed of sound, acoustic attenuation and acoustic impedance are often the most relevant parameters being looked at in phantoms. Hybrid imaging and sensing modalities, which rely on the propagation of both optical and ultrasound waves, present new challenges for phantom development. Photoacoustic (PA) imaging is a prominent example: ultrasound waves are generated with pulsed or modulated light \u20133. For tExisting TMMs that are widely used for optical and ultrasound phantoms have limitations for photoacoustic imaging. Aqueous phantoms such as those based on agar  and gelaet al.  varied from 0.71 dB/cm at 3 MHz to 9.93 dB/cm at 12 MHz. Differences in acoustic attenuation across the gel wax compounds increased with frequency, with a maximum variation of 22% at 12 MHz. These differences may reflect high sensitivities to handling, positioning, and spatial variations in thickness of the gel wax-based slabs. The speed of sound was found similar across the measured gel wax compounds with mean \u00b1 standard deviation values of 1445 \u00b1 2.7 m/s.The addition of TiOThe photoacoustic spectra of gel wax-based samples with inks  were in good agreement with the optical absorption spectra of the slabs within the measured range of 550 to 830 nm (t pulses .2 > 0.9) with the carbon black ink concentration  for photoacoustic imaging phantoms. Previous studies have shown that gel wax has favorable acoustic properties and mechanical robustness ,32. In t tissues , 40. Whi tissues ,42. Addi tissues .2 powder, the materials used in this study were proprietary; their chemical compositions were not known in detail. For consistency across future studies, it would be valuable to have recipes for gel wax that are openly available to the imaging community. This is also the case for optical absorbers for dispersion in gel wax. With these developments, long term studies of the optical, acoustic, and mechanical stability will be important.Apart from the TiOThere are several ways in which the phantom creation methods presented in this study could be extended. To improve the optical homogeneity of coloured inclusions, absorbers and scatterers that are pre-dispersed in oil solutions with lower viscosity than the inks used here may be useful. The acoustic properties could be tuned by including additives such as glass beads, paraffin wax, and co-polymers, as demonstrated in previous studies \u201329, 32. With controllable optical properties, favorable acoustic properties, temporal stability, and mechanical robustness, gel wax could be useful as a base TMM for many imaging and sensing modalities that involve light and/or ultrasound wave propagation. Additionally, it could be valuable as a spatially-localised constituent in photoacoustic imaging phantoms to mimic lipid rich tissue structures such as certain atherosclerotic plaques and nerves \u201348. Gel"}
+{"text": "We aimed to describe ethnic variations in infant mortality and explore the contribution of area deprivation, mother\u2019s country of birth, and prematurity to these variations.We analyzed routine birth and death data on singleton live births  in England and Wales, 2006\u20132012. Infant mortality by ethnic group was analyzed using logistic regression with adjustment for sociodemographic characteristics and gestational age.In the 4,634,932 births analyzed, crude infant mortality rates were higher in Pakistani, Black Caribbean, Black African, and Bangladeshi infants . Adjustment for maternal sociodemographic characteristics changed the results little. Further adjustment for gestational age strongly attenuated the risk in Black Caribbean  and Black African infants  but not in Pakistani , Bangladeshi , and Indian infants . Ethnic variations in infant mortality differed significantly between term and preterm infants. At term, South Asian groups had higher risks which cannot be explained by sociodemographic characteristics. In preterm infants, adjustment for degree of prematurity  fully explained increased risks in Black but not Pakistani and Bangladeshi infants. Sensitivity analyses with further adjustment for small for gestational age, or excluding deaths due to congenital anomalies did not fully explain the excess risk in South Asian groups.Higher infant mortality in South Asian and Black infants does not appear to be explained by sociodemographic characteristics. Higher proportions of very premature infants appear to explain increased risks in Black infants but not in South Asian groups. Strategies targeting the prevention and management of preterm birth in Black groups and suboptimal birthweight and modifiable risk factors for congenital anomalies in South Asian groups might help reduce ethnic inequalities in infant mortality. Marked ethnic disparities in infant mortality have been observed in many countries \u20136, incluA number of individual risk factors for infant mortality are well established, including preterm birth, low birth weight, intrauterine growth restriction, and socioeconomic status ,10,13\u201315Most previous analyses of ethnic variations in infant mortality in England and Wales have either used mother\u2019s country of birth or mother\u2019s/parents\u2019 ethnic group(s) rather than infant\u2019s ethnic group \u201326. AddiThe specific objectives of this study were (a) to describe ethnic variations in infant mortality in England and Wales; (b) to explore the individual and joint effects of infant\u2019s ethnic group, area deprivation and mother\u2019s country of birth, subdivided into UK and non-UK on infant mortality; and (c) to further explore the contribution of preterm birth to ethnic variations in infant mortality.The study was based on a retrospective population-based cohort using routinely collected and linked national data on all singleton live births at gestational ages of 22 weeks and over in England and Wales from 2006 to 2012 inclusive.Birth and death registration data are routinely collected and linked for all births and deaths in the first year of life in England and Wales. Since 2005, these data have been further linked with the National Health Service (NHS) number for babies (NN4B) electronic system. The NN4B system issues an NHS number to all births as soon as possible after birth and provides additional key birth details not collected at birth registration, such as gestational age at birth and the ethnic group of the infant ,29. All We cleaned the data extract provided by Office for National Statistics (ONS) by removing records with implausible or missing values in gestational age and birth weight. We sequentially excluded (a) births with an implausible gestational age, i.e. equal to or more than 43 weeks; (b) births with a missing birth weight; (c) births with an implausible birth weight for gestational age, defined as birth weight more than twice the interquartile range above or below the median, based on sex-gestation-ethnic group-specific birth weight centiles of the study cohort.The ethnic group data used for this study are the infant\u2019s ethnic group as reported in the NN4B birth notification system, guided by a \u2018pick list\u2019 with the ethnic categories used in the 2001 Census in England and Wales . In practh percentile for gestational age and sex derived from our study population.The main explanatory variables included area deprivation score of the mother\u2019s area of residence (as measured by quintiles of the 2015 English Index of Multiple Deprivation (IMD)  and the The primary outcome was infant death, defined as death before one year of age following a live birth. Infant mortality rates were calculated as the number of infant deaths per 1,000 live births. In some analyses, we considered infant mortality by cause and used the ONS cause groups . This isLogistic regression models were used to explore the association between ethnic group and infant mortality. We adjusted all models for sex of infant and their year of birth to account for differences between sexes and temporal changes in infant mortality (models adjusted for these two variables only are referred to as the \u2018base model\u2019 hereafter). A series of sequential adjustments were made to explore the association between infant mortality and ethnic group, IMD quintile, and mother\u2019s country of birth, both individually and jointly (objective b). First, these three exposure variables were added individually to the base model without mutual adjustment to explore the unadjusted associations between ethnic group, IMD quintile, mother\u2019s country of birth and infant mortality (objective b). Second, we adjusted for all maternal sociodemographic characteristics, including IMD quintile, mother\u2019s country of birth, age of mother and marital status/registration type (referred to as \u2018model A\u2019), to explore the joint effects of infant\u2019s ethnic group, area deprivation and mother\u2019s country of birth, taking account of other potential confounders (objective b). Finally, we additionally adjusted for gestational age (referred to as \u2018model B\u2019 or \u2018fully adjusted model\u2019 hereafter) to explore whether gestational age further explained ethnic variations in infant mortality (objective c). Because IMD scores are constructed differently in England and in Wales , we adjuTo further explore the joint effect of ethnic group, IMD and mother\u2019s country of birth, we tested the pairwise interaction between ethnic group and IMD, ethnic group and mother\u2019s country of birth, IMD and mother\u2019s country of birth, respectively using a Likelihood Ratio Test (LRT). We tested for an interaction between ethnic group and preterm birth using a LRT to further explore the contribution of preterm birth to ethnic variations in infant mortality.We conducted a sensitivity analysis in which we additionally adjusted for SGA (referred to as \u2018model C\u2019 hereafter). Because infant mortality attributed to congenital anomalies is known to be higher in Pakistani infants , a separAll analyses were conducted in STATA version 13  , with thThe linked dataset included 4,744,666 singleton live births at gestational ages of 22 weeks or more in England and Wales from 2006 to 2012. In total, we excluded 109,734 (2.3%) births, which included births with implausible gestational age , missing birth weight , or implausible sex-gestation-ethnic group-specific birth weight . The study population therefore consisted of 4,634,932 singleton live born infants at gestational ages of 22 weeks or more and not more than 42 weeks. Among them, approximately 65% were White British, 10% were South Asian , 7% were Other White, 5% were Black (including Black Caribbean and Black African), 4% were of the Mixed groups, and 2.5% were of other ethnic backgrounds. The ethnic group of around 6% of the study population was \u2018not stated\u2019.The characteristics of the mothers and the infants varied by ethnic group . CompareCompared with parents of White British infants, parents of Other White and South Asian infants were more likely to be married or jointly registered as living at the same address while parents of Black Caribbean and Black African infants were less likely to be married or jointly registered as living at the same address.In total, there were 15,001 infant deaths in the study population. The overall infant mortality rate was 3.24 per 1,000 live births in the whole study population and 2.87 in White British infants . Crude iInfants of all non-White ethnic groups had higher infant mortality rates compared with White British infants (ORs between 1.19\u20132.42) in the base model (adjusted only for infant\u2019s sex and year of birth) with Pakistani and Black Caribbean infants having the highest mortality . The lowHowever, after further adjustment for age of mother, IMD quintile, mother\u2019s country of birth, and marital status/registration type, the infant mortality rate was significantly lower for infants of mothers born outside the UK compared with infants of mothers born in the UK . The magnitude of the association between IMD and infant mortality decreased after adjustment but a decreasing gradient was still observed, i.e. lower rates in infants with mothers living in the less deprived areas. This multivariable adjustment changed the association between infant\u2019s ethnic group and infant mortality in different directions for different ethnic groups; the odds ratios increased in Indian, Pakistani, and Bangladeshi infants but decreased in Black Caribbean and Black African infants compared with the base model. After this multivariable adjustment, Other White infants no longer had a lower infant mortality rate than the White British infants .Further adjustment for gestational age  attenuated the association between ethnic group and infant mortality in all ethnic groups, especially in the Black Caribbean and Black African infants. The odds of infant death for Black Caribbean infants were no longer increased compared with White British infants  and the OR for Black African infants reduced to 1.17 (95%CI 1.06\u20131.29) in the fully adjusted model. The three South Asian groups all still had higher odds of infant death than White British infants, with a particularly high OR of 2.32 (95%CI 2.15\u20132.50) for Pakistani infants, and ORs of 1.24 (95%CI 1.11\u20131.38) for Indian infants and 1.47 (95%CI 1.28\u20131.69) for Bangladeshi infants. In this fully adjusted model, IMD remained inversely associated with infant mortality although the effect was attenuated whereas mother\u2019s country of birth was no longer statistically significantly associated with infant mortality . S3 TablWe found no significant interactions (results not shown) between infant\u2019s ethnic group and mother\u2019s country of birth, between infant\u2019s ethnic group and IMD, or between IMD and mother\u2019s country of birth.There was a significant interaction between gestational age dichotomized into term and preterm and infant\u2019s ethnic group (LRT p<0.001). We therefore stratified the study population into term and preterm births to further investigate how the association between ethnic group and infant mortality differed between these groups . To accoIn term infants, additional adjustment for SGA (model C) further reduced the odds in all three South Asian groups compared with model B , but all three groups still had higher odds of infant death compared with the term White British  and preterm infants . Term Indian infants also still had higher odds of infant death from all causes other than congenital anomalies compared with term White British infants  all have higher risks of death which are not explained by maternal sociodemographic characteristics. In preterm infants, adjustment for degree of prematurity fully explains the higher risks of infant death in preterm Black Caribbean and Black African infants compared with White British infants but not in preterm Pakistani and Bangladeshi infants, who have lower rates of very preterm birth. Additional adjustment for SGA appears to further explain some but not all of the residual excess risks in the South Asian groups. Furthermore, the higher rate of death due to congenital anomalies in Pakistani infants does not appear to explain all the excess risk of infant mortality in this group.Major strengths of our study are that the large sample size has given us the power to study smaller more homogeneous ethnic subgroups rather than broad grouping such as \u2018South Asians\u2019; and that we were able to study infant\u2019s ethnic group directly. Additionally the use of statutorily collected, linked birth and death registration data with cohort study design and low levels of missing data minimized potential biases such as selection bias, detection or observer bias and losses to follow-up, although we acknowledge that there may have been some losses to follow-up due to out migration.The comprehensive national coverage of the study also means the findings reflect ethnic disparities in infant mortality across England and Wales. The generalizability of our findings to populations in countries other than England and Wales is likely to depend on the extent to which the ethnic groups are comparable to those in the UK and to other local factors such as disparities in access to healthcare.Some misclassification of ethnic group seems possible, but it seems unlikely that this would have led to any systematic biases in our data except possibly in the mixed ethnic group where there is some limited data suggesting higher levels of misclassification . This isWe lacked information on potentially important modifiable risk factors such as smoking, alcohol use, drug use, diet, and physical activity, maternal obesity, breastfeeding, and uptake of antenatal care.The use of an area-based measure of deprivation (IMD) rather than an individual measure may be subject to the usual limitations of ecological summary measures . HoweverPrevious studies and routinely published data for England and Wales from the 1980s onwards have generally found that South Asian, Black Caribbean and Black African infants tend to have higher infant mortality rates than White infants \u201327,30. AAn important, new finding of our study is that the factors that may explain relatively high infant mortality rates compared with the White British differ for Black and South Asian groups. Gestational age appears to explain almost all the elevated risks in the Black Caribbean and Black African infants but not in the South Asian groups. Higher rates of preterm birth, especially moderately preterm and very preterm birth in the Black groups have been well documented in England and Wales, in other high-income countries and worldwide ,17,40\u201342The other novel finding of our study, the identification of an interaction between preterm birth and ethnic group has further demonstrated the importance of gestational age in the association between ethnic group and infant mortality, especially in explaining higher infant mortality in the Black groups. Further adjustment for degree of prematurity only appeared to explain the higher risks in preterm Black infants. This is broadly consistent with studies that show Black infants have similar or even lower gestational age-specific perinatal or infant mortality rates compared with the White infants only at very low gestational ages but not thereafter ,48\u201350 poOn the other hand, neither preterm birth nor area deprivation, together with other maternal sociodemographic characteristics, explained the increased risks observed in the South Asian groups, in particular the Pakistani group. International studies also show that Pakistani infants have one of the highest infant mortality rates in other host countries ,6. CongeOur study only looked at live births, but the finding of a higher risk of infant mortality in South Asian infants, whether term or preterm, is consistent with the patterns found by Balchin and colleagues that perinatal mortality rate  was highest in infants of South Asian women at all gestational ages . They alOur exploratory analyses adjusting for SGA using cut-offs derived from our own population suggested that differences in SGA or birth weight between ethnic groups might explain some but not all of the residual excess risks in South Asian groups. Low birth weight is an established risk factor for infant mortality  but furtAlthough the association between ethnic group and infant mortality in all the three South Asian groups changed in similar ways when adjusted for maternal sociodemographic characteristics and gestational age, there were still differences in infant mortality rates between the three groups, with Pakistani infants having particularly high risks and Indian and Bangladeshi infants having relatively lower and similar risks. Our data showed that fewer Indian than Pakistani and Bangladeshi infants had mothers living in deprived areas, however, this does not explain the much higher risks in Pakistani infants compared with Bangladeshi infants, nor the relatively small differences between Indian and Bangladeshi infants.Various explanations have been proposed and discussed elsewhere which might explain the differences that we observed between South Asian groups, including genetics, health-related behavior and access to healthcare , but we The identification of differing underlying factors for increased risks in mortality in South Asian and Black infants suggests that different prevention strategies may be required for these groups. The findings that gestational age, particularly more extreme preterm birth, appears to explain the increased risks in preterm Black infants, suggest a need for strategies to ensure that Black mothers have access to services for the prevention and management of preterm birth and for the conditions leading to iatrogenic preterm delivery. For South Asian groups, strategies targeting preventable risk factors for congenital anomalies and also ensuring access to antenatal screening for at risk groups, together with intervention strategies aimed at optimizing birth weight, may reduce some of the excess mortality, but further research is required to better understand other potentially preventable causes of excess mortality in these groups.In conclusion, South Asian and Black infants have higher infant mortality rates compared with White British infants which are not wholly explained by area deprivation, mother\u2019s country of birth or other maternal characteristics. A higher proportion of infants born at lower gestational ages appears to explain the increased risks in preterm Black Caribbean and Black African infants but not in the South Asian groups. An increased risk of infant mortality due to congenital anomalies explains some but not all of the excess risk of infant mortality in South Asian, particularly Pakistani infants.Our findings suggest that further research is needed into the causes of preterm birth in Black Caribbean and Black African mothers with the aim of developing strategies for the prevention and management of preterm birth and the causes of iatrogenic preterm delivery in these mothers and their infants. Excess risks in each of the three South Asian groups and the differences between these groups require further investigation, but strategies targeting suboptimal birth weight and modifiable risk factors for congenital anomalies in South Asian groups would be merited.S1 Table(DOCX)Click here for additional data file.S2 Table(DOCX)Click here for additional data file.S3 Table(DOCX)Click here for additional data file.S4 Table(DOCX)Click here for additional data file."}
+{"text": "HLA-B*27 as well as other human leukocyte antigen variants and at least 47 individual non-MHC-associated variants. However, substantial additional heritability remains as yet unexplained. To identify further genetic variants associated with the disease, we undertook an association study of AS in 5,040 patients and 21,133 healthy controls using the Illumina Exomechip microarray. A novel association achieving genome-wide significance was noted at CDKAL1. Suggestive associations were demonstrated with common variants in FAM118A, C7orf72 and FAM114A1 and with a low-frequency variant in PNPLA1. Two of the variants have been previously associated with inflammatory bowel disease . These findings further increase the evidence for the marked similarity of genetic risk factors for IBD and AS, consistent with the two diseases having similar aetiopathogenesis.Ankylosing spondylitis (AS) is a common chronic immune-mediated arthropathy affecting primarily the spine and pelvis. The condition is strongly associated with  For example, in the International Genetics of AS Consortium Immunochip study, which identified 37 AS-associated variants, the mean minor allele frequency (MAF) of the key-associated single-nucleotide polymorphism (SNP) was 32%, and only two low-frequency (MAF 1\u20135%) AS-associated alleles were identified and one rare allele (MAF<1%).11 As with many common heritable diseases, a large proportion of the heritability of AS remains unexplained. The vast majority of human genetic variation consists of low-frequency and rare variants with MAF<5%.12 Studies performed to date have not addressed such variants extensively due to the design of genome-wide association study SNP microarrays and the sample sizes of the studies not having power to detect association with low-frequency or rare variants.13 It is likely that there is a substantial numbers of additional variants in the low-frequency or rare variant range that remain undiscovered.Ankylosing spondylitis (AS) is an inflammatory arthropathy with a prevalence of 0.1\u20130.5% in populations of European or Asian descent. The condition primarily affects the sacroiliac joints and the spine, initially causing pain and reversible stiffness. Subsequent ankylosis of these leads to fixed spinal deformity and increasing disability. The condition also has extra-articular manifestations, most commonly in the eye (acute anterior uveitis), and rarely in the aorta, kidneys and lungs. Inflammatory bowel disease  is found in 5\u201310% of AS patients, and ~60% of AS patients have subclinical ileal inflammation. The disease is known to be highly heritable, and to date at least 47 independent genetic variants have been shown to be associated with AS susceptibility.14 Much of the variation found in exomes is also rare.16 Recent exome-sequencing analyses have demonstrated the huge number of rare and potentially damaging variants present in human exomes, with ~313 genes per person predicted to be adversely affected by exonic variants.14Exomes are proposed to carry a disproportionately high number of clinically important variants because of their potentially profound effects on the protein function.In this study, we sought to identify further AS-associated genetic variants focusing on exons using the Illumina HumanExome Beadchip microarray. In addition to covering common coding variants, this chip has extensive low-frequency and rare variant content, enabling us to perform a relatively low cost survey of the role of such variants in AS (compared with studies utilising whole-exome or -genome sequencing).After participant quality control (QC) there were 4,602 AS cases and 20,164 healthy controls. After SNP QC there were 207,193 SNPs. Principal components analysis was performed with 0\u201310 eigenvectors; the scree plot of genomic control-1000 statistics for different numbers of eigenvectors is shown in P<5\u00d710\u22126) with AS in the current study . SNP associations within ANTXR2, FCGR2A, IL1R1 and NOS2 were confirmed at suggestive levels of significance. At nine of the confirmed loci, association was seen with the primary associated SNP. At NOS2 the association was with rs2297518, which has been reported previously as a secondary signal at the NOS2 locus, the primary associated SNP rs253187511 not being included on the HumanExome Beadchip.Assays for 10 non-MHC SNPs previously associated with AS were included on the HumanExome Beadchip, and each one showed at least suggestive association  at a genome-wide level of significance =1.2), C7orf72  and FAM114A1 , and with a rare variant in patatin-like phospholipase domain containing 1 (PNPLA1) . SNPs in C7orf72, including the same SNP we have identified as being associated with AS (rs1456896), have been associated with Crohn\u2019s disease.18FAM118A variants have previously been associated with bone mineral density.22 Strong linkage disequilibrium exists between the bone mineral density-associated SNP (rs136564) and the AS-associated SNP, rs6007594 . Functional annotation for the variants is shown in Suggestive associations were seen with common variants in ERAP1 and IL23R  and rs10050860-C  1.27\u20131.43 and conditioned on rs30187 P=9.7\u00d710\u221210, allele C, OR=1.2, 95% CI 1.14\u20131.30). Moderate linkage disequilibrium was observed between rs30187 and rs10050860 (r2=0.156 and D\u02b9=1.00). As previously reported, after conditioning on rs30187, association was confirmed with the neighbouring ERAP2 gene . This association remains after controlling for both the rs30187 and rs10050860 ERAP1 variants . No additional associated variants were evident.After conditioning on the primary association signals there were second, independent signals observed in nd IL23R . The patIL23R, the intronic variant rs11465804 was slightly more significantly associated than the previously reported rs11209026 nonsynonymous SNP . These two SNPs are in high linkage disequilibrium , and conditioning for either SNP controlled for the association at the other SNP, suggesting that the rs11465804 variant tags this previously demonstrated functional SNP.24 As previously reported, a secondary association is seen at the IL23R locus, evident after conditioning on rs11465804 .3 No additional associated variants were evident.At FAM114A1 locus; these SNPs sit in the TLR10 gene  and rs6841698 .An independent signal supported by multiple SNPs was demonstrated after conditioning on the main SNP at the R10 gene . SNPs inUsing the SKAT-O test, no genes were significantly associated with AS at either a genome-wide or suggestive level of significance. This was the case for all the sets of variants incorporated into the burden equation including the following: (a) \u2018Damaging set\u2019, (b) \u2018Polyphen set\u2019 and (c) all variants. No association was identified even after removing the restrictive minor allele filter of 0.05 and including SNPs of all frequencies.\u22128, and equal MAF for the disease-causative and genotyped markers, the study had 100% power for MAF=0.05, but only 9% power for MAF=0.01 and close to zero (6.1\u00d710\u22125%) power for the median MAF in this study (2.0\u00d710\u22124).With sample sizes of 4,602 AS cases and 20,164 controls used in this study, the power to detect the association was good for low MAF variants, but low for rare variants . AssuminERAP1, IL23R, chromosome 2p15, chromosome 21q22, KIF21B-GPR25, GPR35, IL6R, ANTXR2, IL1R1, FCGR2A and NOS2). It also describes a common novel AS-associated non-MHC variant that achieved genome-wide significance (CDKAL1), and three novel common variants that achieved a suggestive level of significance . One new suggestive rare variant association in PNPLA1 was identified in single marker analyses, in addition to low-frequency associations observed with rs11465804 in IL23R and rs4349859 in HLA-B*27. However, no rare variant associations were noted using burden tests.This study re-demonstrates a number of known AS genetic associations both within and outside the MHC .33 Nonetheless, major differences exist between genetic associations of the two diseases pointing to differences in disease-specific aetiopathogenesis; for example, the absence of association of IBD with HLA-B*27, and the absence of association of AS with the major IBD loci NOD2/CARD15 and ATG16L1. The finding of three more concordant genetic associations further strengthens the evidence of shared aetiopathogenesis between these diseases.In the current study, we identify three AS-associated variants that have previously been associated with IBD, namely variants in or near FAM114A1. It may be responsible for the described association, but the corresponding protein NOXP20 contains a predicted caspase recruitment (CARD) domain suggesting that it may be involved in apoptosis.34CARD9 has previously been associated with AS and NOXP20 may have a similar role. In addition, neighbouring genes include the Toll-like receptors TLR1 and TLR6 that are intimately involved in innate immunity and so are strong biological candidates for involvement in AS. The second independent signal at the FAM114A1 locus was in the toll-like receptor 10 gene (TLR10). This missense variant (rs11096955) is predicted by Polyphen-2 to cause a benign change (isoleucine to leucine), but it may tag other more functional variants. This association implicates this important component of the innate immune system in AS aetiology. Toll-like receptors recognise evolutionary conserved sequences on pathogens and trigger immune responses. TLR10 has been recently identified to induce pro-inflammatory cytokine production and interferon in response to influenza infection.35 It has recently been suggested that immunodeficiency to gut organisms may trigger AS; if the association with TLR10 impairs innate immune responses, this would be consistent with this theory.36Little is known about FAM118A is a protein-coding gene of unknown function that encodes a single-pass transmembrane protein (www.uniprot.org). The AS-associated SNP (rs6007594) is a missense mutation causing an arginine to be replaced by a histidine. This missense change is predicted to be probably damaging with a Polyphen-2 score of 0.999 .37 Kwan et al.22 demonstrated that FAM118A is expressed in lymphoblastoid cell lines as well as human osteoblasts, and in both cell types showed major SNP effects on FAM118A expression levels. How this impacts on AS aetiology is not immediately evident and larger studies studying SNPs across this locus will be required to determine the genetic variant(s) responsible for the association observed here.PNPLA1 gene belongs to a family of genes, the members of which have diverse lipolytic and acyltransferase activities. The function of PNPLA1 itself is not well understood. It is expressed in epidermal keratinocytes, and has a role in glycerophospholipid metabolism in the cutaneous barrier. Variants in PNPLA1 are associated with the skin disorder ichthyosis.38 The rs141744967 variant is a missense polymorphism that causes a change from alanine to valine. The functional effect is not available from the Polyphen-2 server; both amino acids are non-polar but differ in size by 28\u2009Da. Further larger studies will be required to determine whether this gene is definitively associated with AS.The P=5\u00d710\u22128 using the participant numbers in this study, the study only had 80% power to detect variants with an additive relative risk of >1.8; other than human leukocyte antigen (HLA) associations, few such variants have been reported in common diseases. This demonstrates poor power to detect individual associations, and increasing the number of cases would improve this power. Nonetheless, if there are large numbers of rare variant associations contributing to AS, the study should have had good power to detect some of these, assuming its coverage of rare variants was good.This study has several potential limitations; the major limitation is power. The power to detect rare variants is a function of their allele frequency and their effect size along with the population frequency of disease and the required statistical significance threshold. While the study had excellent power to detect common variant associations, the power to detect rare variants was low. Considering variants with a frequency of 0.01 (1%), population disease frequency of 0.005 (0.5%) at a significance threshold of 12 This suggests that comprehensive rare variant microarray studies may not be feasible, although improvements in imputation methods raise the possibility that many rare but not unique variants may be addressable by this approach.39The coverage of the Exomechip microarray of rare variants is far from comprehensive, and this impacts both on the coverage of the study, and its ability to pinpoint genetic associations. Further fine-mapping and functional studies will be required to confirm whether the genes we have implicated at each locus are themselves directly involved in AS, or if the SNP associations observed operate by influences on other genes. Sequencing of whole genomes has demonstrated millions of low frequency and rare variants that are not covered on the chip, for example, in the low-coverage analysis of 1000 genomes 15.5 million variants were identified.Finally, rare variants do not share linkage disequilibrium with many other surrounding variants to the extent that common variants do. Therefore if identified in a study such as this, a good check of association is manual inspection of the genotype intensity clustering, and, in addition, considering the biological plausibility. However, probes can map to other areas of the genome without our knowledge giving well-clustered intensity plots. Biological plausibility is not necessarily a good measure of a true association as the association may be the first association in a pathway not previously known to be involved in disease aetiology. This makes independent replication studies essential, although it is particularly challenging for low-frequency or rare variants because of the sample size requirements.IBD genes increases the evidence of shared aetiopathogenesis between the diseases and the potentially important role of intestinal dysbiosis in AS.41 The major overlap between AS and IBD is also supported by another study showing similar genetic variants but differing effect sizes between variants associated with AS and anterior uveitis.42 Whether low-frequency and rare genetic variants are major contributors to the aetiopathogenesis of AS remains unclear and will likely require much larger studies with more comprehensive coverage of these variant types to resolve.This study has re-demonstrated many known AS risk loci, and also identified a novel common variant at a genome-wide level of significance, and four suggestive associations, including one rare variant association. The finding of further concordant associations with 43 from the United Kingdom, Australia and New Zealand were recruited . Healthy controls were provided by the following groups (1) 1958 British Birth Cohort ; (2) GoDarts type 2 diabetes cohort ; (3) Oxford Biobank ; (4) Twins UK cohort ; (5) Anglo-Australasian Osteoporosis Genetics Consortium . All patients gave written informed consent and ethical approval was provided by all appropriate institutional review boards.AS patients of European descent who met the modified New York criteria44 Each cohort had QC completed separately, assessing missingness by individual (threshold <3%), missingness by genotype (threshold<3%), Hardy-Weinberg equilibrium in controls (Chi-square test threshold P=0.01), extreme heterozygosity (threshold>3 standard deviations from mean) and identity by descent threshold of PI_HAT 0.20 was used. After laboratory QC that excluded ~3,000\u20135,000 SNPs per cohort, 20,714\u201322,864 SNPs were removed from each set to form a common SNP basis, 1,033 SNPs were removed due to excess missingness, 526 SNPs were removed from control sets due to not being in Hardy\u2013Weinberg equilibrium and 11,711 SNPs were removed due to allele and frequency inconsistencies between the cohorts. 979 subjects were removed due to excess relatedness, 207 were excluded due to extreme heterozygosity and 2 were excluded due to excessive missingness (after SNP QC).Each cohort was genotyped using the Illumina Infinium HumanExome BeadChip version 1.2. This Illumina microarray has ~240,000 markers, made up of exonic variants, splice variants, stop altering variants, ancestry informative markers and MHC tag SNPs. Genotype calling was completed with zCall.http://www.stats.ox.ac.uk/~davison/software/shellfish/shellfish.php). Unsupervised model-based clustering implemented in R with MCLUST was used to exclude patients deemed to be non-European after plotting with HapMap controls. This model assigns a cluster to each individual based on their principal component analysis values and in consideration to the weighted centre of each cluster and therefore assigns non membership status to those who don\u2019t cluster with core HapMap groups. This analysis identified 219 subjects who were removed due to non-European ethnicity. Shared genotyped SNPs between cohorts with MAF>0.05 were then used to perform principal component analysis for ethnicity identification using SHELLFISH (\u03bb) was calculated using the designer suggested random set of 5,000 synonymous variants (http://genome.sph.umich.edu/wiki/Exome_Chip_Design#Random_set_of_synonymous_variants_.28as_comparator.29).The genomic control measure  All variants, (b) \u2018Polyphen2 set\u2019: Polyphen-237 predicted possibly damaging or probably damaging, and (c) \u2018Damaging Set\u2019: Variants causing the following consequences: frameshift substitution, nonframeshift substitution, nonframeshift deletion, nonframeshift insertion, frameshift deletion, frameshift insertion, nonsynonymous single-nucleotide variant, stop-gain single-nucleotide variant, stoploss single-nucleotide variant, missense variant, splice acceptor variant, splice donor variant, splice region variant, initiator codon variant, stop retained variant and incomplete terminal codon variant.For low-frequency variant analysis we used the sequence kernel association test\u2013Optimal (SKAT-O) test that computes the SKAT test\u22124, reflecting Bonferroni correction for the 424 HLA alleles that were tested for association. Conditional analyses were completed by adding the allele being conditioned as a covariate to the logistic regression model. Power calculations were performed with the online Genetic Power Calculator.Cluster plots for reported SNPs were checked manually in the case cohort and the 1958 British Birth Cohort. Association of classical alleles was completed with imputed SNP2HLA classical alleles and one principal component by logistic regression in R. The level of significance for this analysis was 1.2\u00d710http://pngu.mgh.harvard.edu/~purcell/gpc/)Genetic Power Calculator: (http://www.broadinstitute.org/mpg/snap/)SNAP browser: ("}
+{"text": "Inherent differences between naturally-formed lakes and human-made reservoirs may play an important role in shaping zooplankton community structure. For example, because many reservoirs are created by impounding and managing lotic systems for specific human purposes, zooplankton communities may be affected by factors that are unique to reservoirs, such as shorter water residence times and a reservoir\u2019s management regime, compared to natural lakes. However, the environmental factors that structure zooplankton communities in natural lakes vs. reservoirs may vary at the continental scale and remain largely unknown. We analyzed data from the 2007 U.S. Environmental Protection Agency\u2019s National Lakes Assessment and the U.S. Army Corps of Engineers\u2019 National Inventory of Dams to compare large-bodied crustacean zooplankton communities  in natural lakes and reservoirs across the continental U.S. using multiple linear regressions and regression tree analyses. We found that large-bodied crustacean zooplankton density was overall higher in natural lakes compared to reservoirs when the effect of latitude was controlled. The difference between waterbody types was driven by calanoid copepods, which were also more likely to be dominant in the >0.243 mm zooplankton community in natural lakes than in reservoirs. Regression tree analyses revealed that water residence time was not a major driver of calanoid copepod density in natural lakes but was one of the most important drivers of calanoid copepod density in reservoirs, which had on average 0.5-year shorter water residence times than natural lakes. Reservoirs managed for purposes that resulted in shorter residence times  had lower zooplankton densities than reservoirs managed for purposes that resulted in longer residence times . Consequently, our results indicate that water residence time may be an important characteristic driving differing large-bodied zooplankton dynamics between reservoirs and natural lakes. Zooplankton are a vital component of aquatic food webs and ecosystem functioning. Zooplankton provide a crucial link between primary producers and higher trophic levels \u20133, are iWaterbody origin, i.e., if a waterbody is naturally-formed or human-constructed (a reservoir), may play an important role in structuring zooplankton communities because of inherent differences between the two waterbody types \u201316. For To the best of our knowledge, only two studies have directly examined how zooplankton communities vary between reservoirs and natural lakes , 24. BotDifferences in reservoir management  may influence zooplankton communities because reservoir purpose may affect many environmental characteristics, such as WRT , 23. ForDaphnia) differed between the two waterbody types and across reservoirs of different purposes in the continental U.S. First, we compared zooplankton communities and environmental drivers known to be important factors shaping zooplankton density and composition  between natural lakes and reservoirs at the continental U.S. scale while controlling for the effect of latitude. We predicted that zooplankton density and genera richness would be overall lower in reservoirs than in natural lakes. Second, we focused specifically on reservoirs and analyzed the effects of reservoir primary purpose  and environmental drivers on zooplankton density. We predicted that zooplankton densities would be lower in reservoirs used for purposes that generally result in shorter WRT  than those used for purposes that result in longer WRT .We analyzed data from the U.S. Environmental Protection Agency\u2019s (EPA) 2007 National Lakes Assessment (NLA) and the U.S. Army Corps of Engineers\u2019 National Inventory of Dams (NID) to test how environmental drivers of crustacean zooplankton density and genera richness ; therefore, there was likely a minimal effect of the time of year when natural lakes were sampled compared to when reservoirs were sampled. All sampled waterbodies were at least 0.04 km2 and 1 m deep and were chosen using a random stratified design based on surface area, ecoregion, and state , 8214, 8Reservoir WRT was significantly different across reservoirs of different primary purposes. Recreation, hydroelectric, and flood control reservoirs had half the WRT, on average, of reservoirs used for irrigation, which likely is related to differences in reservoir management operations. For example, hydroelectric reservoirs may have shorter WRT to meet electrical power demands , 84, and-1. High discharge reservoir purposes, such as hydroelectric reservoirs, may flush many of these taxa from the system before they become adults. Other factors certainly also play a role in these density differences across reservoirs of different purposes, as many other environmental factors varied across reservoir purposes. For example, chlorophyll a was lower in hydroelectric reservoirs versus some other reservoir types, which can result in lower zooplankton densities [a was also lower in reservoirs used for irrigation, which had the greatest crustacean zooplankton densities. Overall, our results suggested that reservoir purpose likely affects multiple environmental factors, which in turn can indirectly alter zooplankton densities and community structure.Our results suggest that zooplankton densities differed between reservoirs of different primary purposes as a result of differences in WRT: hydroelectric reservoirs (with shorter WRT) had the lowest zooplankton densities and irrigation reservoirs (with longer WRT) had the greatest zooplankton densities. Total copepod and calanoid zooplankton densities were especially lower in hydroelectric reservoirs versus most other reservoir purposes , with deensities , 38, 66.Daphnia in analyses, we were still able to compare calanoid copepods with one cladoceran taxa that was likely minimally affected by the sampling methods.The EPA\u2019s NLA had excellent spatial coverage of waterbodies. However, one limitation of our work is that most lakes and reservoirs were sampled only once and thus each zooplankton sample represents a single \u201csnapshot\u201d of the system. Consequently, other important factors such as plankton succession , 87, 88,This study constitutes one of the most comprehensive spatial datasets for the analysis of zooplankton, spanning more than 700 waterbodies in the U.S. Altogether, our findings indicated differences in zooplankton community structure between natural lakes and reservoirs. Our results also indicated that the relative importance of environmental drivers on zooplankton communities varied between reservoirs and natural lakes. Such zooplankton community and density differences may in turn alter freshwater food webs and water quality , 5\u20137. AsS1 Table-1), Chlorophyll a = chlorophyll a concentration (\u03bcg L-1), and WRT = water residence time (years). Also included are other environmental variables not included in our analyses because of correlations with r > 0.50 with the focal environmental variables: Max depth = maximum waterbody depth (m), Calcium = calcium concentration (mg L-1), and DOC = dissolved organic carbon concentration (mg L-1).\u201cNS\u201d refers to relationships that are not significant and \u201c.\u201d refers to empty cells. The focal environmental variables included in our analyses were: Temp max = maximum waterbody temperature (\u00b0C), DO mean = mean water column dissolved oxygen concentration (mg L(DOCX)Click here for additional data file.S2 TableN for each leaf, each leaf\u2019s response variable mean \u00b1 SE, and the description for node splits for each leaf group. The percentage of the waterbodies that consist of reservoirs are provided for each leaf split, for analyses across both natural lakes and reservoirs.Reported for each regression analysis is each individual leaf group, (DOCX)Click here for additional data file.S3 Table-1), Chlorophyll a = chlorophyll a concentration (\u03bcg L-1), and WRT = water residence time (years).All refers to aggregated analyses with both natural lakes and reservoirs. Temp max = maximum waterbody temperature (\u00b0C), DO mean = mean water column dissolved oxygen concentration (mg L(DOCX)Click here for additional data file.S1 FigN > 20 for each of these categories.Primary purpose was obtained for all reservoirs with available data from the U.S. Army Corps of Engineers\u2019 National Inventory of Dams database. (TIF)Click here for additional data file.S2 Fig(TIF)Click here for additional data file.S3 FigA refers to waterbodies that have pH \u2264 7.9; B refers to waterbodies with pH > 7.9 but \u2264 8.7; C refers to waterbodies with pH > 8.7 and water residence times > 0.338 years; D refers to waterbodies with pH > 8.7 and water residence times < 0.338 years.(TIF)Click here for additional data file.S4 FigA refers to waterbodies that have maximum temperatures > 27.5 \u00b0C; B refers to waterbodies with maximum temperature < 27.5 \u00b0C and pH < 8.38; C refers to waterbodies with maximum temperature < 27.5 \u00b0C, pH > 8.38, and water residence times > 0.177 years; D refers to waterbodies with maximum temperature < 27.5 \u00b0C, pH > 8.38, and water residence times < 0.177 years.(TIF)Click here for additional data file."}
+{"text": "Sperm production is one of the most complex biological processes in the body. In vitro production of sperm is one of the most important goals of researches in the field of male infertility treatment, which is very important in male cancer patients treated with gonadotoxic methods and drugs. In this study, we examine the progression of spermatogenesis after transplantation of spermatogonial stem cells under conditions of testicular tissue culture.PLZF protein. These cells, after being labeled with DiI, were transplanted in azoospermia adult mice model. The host testes were placed on agarose gel as tissue culture system. After 8\u00a0weeks, histomorphometric, immunohistochemical and molecular studies were performed. The results of histomorphometric studies showed that the mean number of spermatogonial cells, spermatocytes and spermatids in the experimental group was significantly more than the control group (without transplantation) (P\u2009<\u20090.05) and most of the cells responded positively to the detection of DiI. Immunohistochemical studies in host testes fragments in the experimental group express the PLZF, SCP3 and ACRBP proteins in spermatogonial cells, spermatocyte and spermatozoa, respectively, which confirmed the human nature of these cells. Also, in molecular studies of PLZF, Tekt1 and TP1, the results indicated that the genes were positive in the test group, while not in the control group.Testicular tissue samples from azoospermic patients were obtained and then these were freeze\u2013thawed. Spermatogonial stem cells were isolated by two enzymatic digestion steps and the identification of these cells was confirmed by detecting the These results suggest that the slow freezing of SSCs can support the induction of spermatogenesis to produce haploid cells under the 3-dimensional testicular tissue culture. It was reported in 2010 that 0.4% of young patient with childhood cancer can live for a long time . A surveFor each experimental group, 3\u20135 NMRI mouse  were used at the age of 4\u00a0weeks. These mice are treated with Busulfan 40\u00a0mg/kg  and after 4\u00a0weeks, the azoospermia model is developed .This study is based on 5 biopsy samples taken from different obstructive azoospermic patients [sperm positive in their testicular sperm extraction (TESE)] referred to Gandhi Infertility Clinic Center. After performing clinical procedure and confirming active spermatogenesis, and obtaining verbal informed consent from the patients, surplus testis samples were donated to the research laboratory. All stages of this research were based on the approval of the research ethics committee of Tarbiat Modares University with the registration ID IR.TMU.REC.1394.68.For the freezing and thawing of testicular tissues, the protocol of the Zeng et al.  was used2. The identification of isolated and purified SSCs was investigated by tracing the PLZF protein as a stem cell marker [SSCs were isolated by Mirzapour et al.  protocoll marker  in colonIn particular, 1.5% agarose  solution was prepared and sterilized. Segments with dimensions of 1\u00a0mm\u2009\u00d7\u20091\u00a0mm\u2009\u00d7\u20091.5\u00a0mm of agarose were arranged by scalp considering sterile condition . They weTo detect the transplanted cells and purify them from testicular endogenous cells, the cells cultured with a density of about 80% before transplantation for 5\u00a0min exposed to a 2\u00a0\u03bcg di-alkyl indocarbocyanine (DiI)  from a 1\u00a0ml preservative solution PBS  was placed at room temperature and then placed in a dark place for 20\u00a0min at 4\u00a0\u00b0C. After ensuring that the cells were stained under a fluorescent microscope, the cell surface was washed with PBS and then isolated from of petri dish by trypsin enzyme (25%) in 0.1% ethylenediamine tetraacetic acid (EDTA) . after 3 times washing in the medium, they were transplanted into the host testis. SSCs were transplanted into the host testes below the stereo microscope then they were cut into small pieces and placed under 3-D tissue culture conditions on the agarose support layer. For IVT of SSCs to exited host testes we are using Sato et al.  protocolA total of 5 sections, spaced equally apart, were selected from successive sections of the each testis. After hematoxylin and eosin (H&E) staining of each section, 10 seminiferous tubules with rounded or close-circle sections randomly selected were used to evaluate the testicular parameters. The number of spermatogonial cells, spermatocyte and spermatid per unit volume were measured in each testis by image-j software and previous studies , 19.PLZF protein [SCP3 protein [ACRBP protein [PLZF antibody , Rabbit anti SCP3 antibody , Rabbit anti ACRBP antibody . The secondary antibodies used were goat anti mouse IgG and goat ant rabbit igG, conjugated with Alexa 488 .The testicular tissue fragments of the experimental groups, in addition to the tracing of DiI, were subjected to immunohistochemistry after tissue processing. To confirm the nature of SSCs, spermatocyte and spermatozoa the  protein , SCP3 pr protein  and the  protein  were det protein . BrieflyPLZF, Tekt1 and Tnp1 were obtained from the NCBI database and the sequence of their exons and introns was determined. Primer design was done using the Primer3 online software. Designed primers are blasted to confirm their accuracy and reproduce only the genes\u2019 mRNA sequences. The sequences of the Real Time PCR primers of PLZF, Tekt1 and Tnp1 genes are shown in Table\u00a0\u2212\u2206\u2206CT) was used to determine the relative quantification of the target genes normalized to a housekeeping gene (\u03b2 actin). A validation experiment was performed to verify that target efficiencies and reference were approximately equal.In order to prove the presence of different classes of germ cells and to prove that these cells are not due to endogenous spermatogenesis of the mouse testis, testicular fragments of the experimental groups were studied in the of PLZF, Tekt1 and Tnp1 genes. The human specificity of primers designed to differentiate cells. In order to design the primers used in Real Time PCR, the gene sequences from All quantitative data in this study were presented as mean\u2009\u00b1\u2009standard deviation. One-way ANOVA, T-test and Tukey test were used for statistical analysis. The significance level was considered to be P\u2009<\u20090.05.PLZF protein in the colonies derived from the culture cell suspensions obtained . However, the expression of Tekt1 and Tnp1 genes in transplantation group was increase significantly compared to 0th day cell suspension (P\u2009<\u20090.05). The expression of Tnp1 in postmeiotic cells in host testis was very important because it shows human SSCs can subsided on mouse testis niche and initiate spermatogenesis to produce haploid germ cells as well as spermatids od sperm like cells.The identification of the isolated SSCs was confirmed by tracing the ned Fig.\u00a0. In the ned Fig.\u00a0. The recned Fig.\u00a0 that shoely Fig.\u00a0. Transcrup Chart\u00a0. The expThe transplantation of frozen\u2013thawed testicular tissue or SSCs of cancer patients after the completion of the cancer gonadotoxic treatment period can lead to the re-emergence of malignant cells . In an aThe transplantation of SSCs into the azoospermia testis model has been carried out by various researchers on various animal models . In the The results of this study indicate that this freezing setup hasn\u2019t major problem to support the progression and induction of spermatogenesis to obtaining haploid cells under IVT system and testicular tissue culture. However, the process of freezing testicular tissue, causing damage to the cells, is undeniable. But this method for cancer patients can only be a way of maintaining fertility. Therefore, the methods of testicular tissue freezing should be optimized to minimal damage possible."}
+{"text": "In vitro transplantation (IVT) of spermatogonial stem cells (SSCs) is one of the most recent methods intransplantation in recent decades. In this study, IVT and SSCs homing on seminiferous tubules of host testis in organ culturehave been studied.in vivo transplantation as transplantation groups, and testes without transplantation ofcells were assigned in the control group. Then histomorphometric, immunohistochemical and molecular studies were doneafter 2 weeks. In this experimental study, human SSCs were isolated and their identities were confirmed by trackingtheir promyelocytic leukemia zinc finger (PLZF) protein. These cells were transplanted to adult azoospermia mouse testesusing two methods, namely, IVT and in vivo groups were significantly more than those in thecontrol group (P<0.05). Immunohistochemical studies in the transplantation groups confirmed that the PLZF proteinwas expressed in the cells subsided on the seminiferous tubule. Quantitative reverse-transcription polymerase chainreaction (qRT-PCR) demonstrated that the PLZF gene expression was only positive in the transplantation groups, butit was not significantly different between the IVT group and the in vivo group (P>0.05). After two weeks, histomorphometric studies revealed that the number of subsided spermatogonial cells (SCs)and the percentage of tubules with subsided SCs in IVT and Testicular tissue culture conditions after SSC transplantation can help these cells subside on the seminiferoustubule basement membrane. Some male germ cells called spermatogonialstem cells (SSCs) exist on the basic membrane ofseminiferous tubules in testis and participate inspermatogenesis . These Sin vivo and in vitro conditions. In vivo transplantation has been extensively investigated and successful results have been obtained. Other researchers examined SSC transplantation in different species  to host testes. For these reasons, this procedure must be conducted in vitro,and rigorous follow-up sessions are required. Another limitation of in vivo transplantation is the immunesystem reaction of a recipient. So, it is resulted that IVT is a good choice for these situation in we cannottransplant in vivo. However, this issue is even more important in cancer patients who are exposed to chemotherapy and radiotherapytreatments because of the high risk of returning cells to cancer patients prior to treatment . In otheTherefore, Sato et al.  proposedin vitrospermatogenesis and establishing a tissue cultureenvironment, we aimed to develop a suitable in vitrorecipient testis for the homing and resumption ofspermatogenesis with human SSCs.Considering the importance of achieving In this experimental study, samples were obtained from five patients with obstructive azoospermia after therapeutic testicular sperm extraction (TESE) was completed, and the remaining samples were collected after informed consent was acquired. The testis tissue were washed with phosphate buffer serum  and subjected to two-step enzymatic digestion according to the technique suggested by Mirzapour et al.  with tryThe identity of isolated and purified SSCs was verified by tracking the PLZF protein  in the To set up an agarose support layer and a culture medium with specific compositions and growth factors, we used the method described by Yokonishi et al. . In parin vivo group  at room temperature and incubated them at 4\u00b0C in the dark for 20 minutes . Staininvo group  remainedvo group . To condvo group . In thisV were calculated by image-j software. An optical microscope equipped with an ophthalmologiceye lens and image-j software was used to measure variousstructural parameters in the sections prepared from the testesin the groups , 22-24. Two histological sections were prepared from each recipient testis with an interval of 12 \u00b5m to obtain the percent of tubules with SSCs subsiding on the seminiferous tubules , and all\u00df-actin) and the target genewere amplified in the same run. Comparative CT method(2-\u0394\u0394CT) was used to determine the relative quantification ofthe target gene normalized to a housekeeping gene (\u00df-actin).The primer sequences of PLZF gene is:PLZF as a pluripotency gene in a testicular tissue fragmentwas evaluated after two weeks. Total RNA was extracted from the tissue fragments of all of groups on day 14 by usingRNX-PlusTM  according to the manufacturer\u2019srecommendations. RNA concentration was then determined using a UV spectrophotometer . cDNAwas synthesized from 1000 ng RNAwith a Revert AidTM first-strand cDNAsynthesis kit  using oligo(dT) primers. PCRs were performed using Master Mix andCYBER Green I  in Applied BiosystemsStepOneTM instrument . The PCR program was started with an initial melting cycle at 94\u00b0C for4 minutes to activate the polymerase and followed by 40cycles of a melting step (20 seconds at 94\u00b0C), an annealingstep (30 seconds at 57\u00b0C), and an extension step (20 secondsat 72\u00b0C). After the PCR run was completed, the quality of thereactions was confirmed through melting curve analyses. Foreach sample, the reference gene . Data are presented as mean \u00b1 SE.Each data point represented the average of three separateexperiments, and five repeats were set up in each experiment. P<0.05 indicated statistical significance. The experimental stages in this research were in accordancewith the approval of the Ethics Committee of Tarbiat ModaresUniversity .The cell suspension, containing SCs and Sertoli cells,was obtained and placed under culture conditions.Immunocytochemistry revealed that the colonies derived from the SCs, including SSCs, expressed the PLZF protein . in vivo groups . The percent of tubules with SC homing in the IVT group was significantly higher than that in the control group  in the IVT and shown in . To confirm the nature of cells subsided on the basement membrane of seminiferous tubules, we detected PLZF protein in these cells, and the results revealed that this protein was positively expressed in the transplantation groups . in vivo group supported homing of transplanted cells. They transplanted human SSCs into the mouse testis and found that these cells adhere to the basement membrane of seminiferous tubules after 2 weeks in vivo condition.Autologous germ cell transplantation is a potential approach to restore fertility, especially for childhood cancer survivors who have become infertile due to cytotoxic therapies to treat cancer . At the in vitro to an immature azoospermic testis. As a result, transplanted cells were subsided on the basement membrane of seminiferous tubules after 7-14 days. They further labeled these cells with Acrosin Green Florescent Protein (GFP) to track them; after 7-14 days,these cells are detected. Our research emphasized that testicular tissue culturesystem could support homing of transplanted cells inthe testes of recipients. Our observations in the IVT andtissue culture of the recipient\u2019s testis were consistentwith those of Sato et al. , who werA major problem in studying SSCs homing is that it is difficult to track SSCs immediately after transplantation . This isOur results are also consistent with those of Illien-J\u00fcnger et al. , who traAccording to similar and confirmative results of other researchers, a testicular culture system can support not only the homing of SSCs on the basement membrane of seminiferous tubules, but also the resumption of spermatogenesis because of the secretion of proteins and materials from interstitial tissues as well as presence of the receptors and proteins on the membrane of SSCs . in vitro and homing of these cells in the testis was supported by tissue culture conditions. So, IVT and testis tissue culture system can be a good alternative to in vivo SSCs transplantation. It seems to be possible with this system to indicate that the in vitro conditions can be set up in a manner similar to the conditions in the body, so that we can do many goals that cannot be done within the body. Further studies should be performed to assay spermatogenesis after accomplishment of IVT and testis tissue culture. Our results indicated that human SSCs were successfully transplanted in azoospermic mouse testis"}
+{"text": "The effects of low-dose radiation are being increasingly investigated in biological, epidemiological, and clinical studies. Many recent studies have indicated the beneficial effects of low doses of radiation, whereas some studies have suggested harmful effects even at low doses. This review article introduces various studies reporting both the beneficial and harmful effects of low-dose radiation, with a critique on the extent to which respective studies are reliable. Epidemiological studies are inherently associated with large biases, and it should be evaluated whether the observed differences are due to radiation or other confounding factors. On the other hand, well-controlled laboratory studies may be more appropriate to evaluate the effects of low-dose radiation. Since the number of such laboratory studies is steadily increasing, it will be concluded in the near future whether low-dose radiation is harmful or beneficial and whether the linear-no-threshold (LNT) theory is appropriate. Many recent biological studies have suggested the induction of biopositive responses such as increases in immunity and antioxidants by low-dose radiation. Based on recent as well as classical studies, the LNT theory may be out of date, and low-dose radiation may have beneficial effects depending on the conditions; otherwise, it may have no effects. Radiation exposure at high-dose levels  is considered to be harmful and it increases the incidence of cancer. On the other hand, the effects of lower-dose exposure remain controversial. Some consider radiation exposure below 200 mSv to be hazardous based on the linear-no-threshold (LNT) hypothesis, whereas others consider low-dose exposure to have beneficial effects, known as radiation hormesis and the radioadaptive response ,4,5. OthStudies addressing the effects of low-dose radiation are divided into biological, epidemiological, and clinical (human) ones. Well-designed and well-controlled biological studies would be reliable, but it should be noted that large numbers of subjects are usually necessary in view of the relative weakness of the effects compared with the high-dose effects. While influences on humans need to be inferred from the results on animals, insects, and other living organisms in biological studies, the results of epidemiological studies are directly related to human health. However, epidemiological studies inherently contain many large biases, and the results must be interpreted much more cautiously, with special attention to the confounding factors. In this context, prospective studies on humans may be optimal to investigate the issue of low-dose radiation effects, although a large-scale study is not easy to conduct.There have already been numerous studies addressing the issue of low-dose radiation effects; many suggest beneficial or otherwise no effects, whereas some others suggest harmful effects. It is impossible to review all such articles. The purpose of this review is to introduce and criticize studies in which we are particularly interested from our viewpoints as radiologists, and also to introduce our own results and unpublished data from the group associated with the authors. Based on the data, we discuss the effects of low-dose radiation. Since the issue is closely related to whether the LNT hypothesis is true, let us start with the LNT hypothesis. Drosophilia melanogaster in the high-dose range, and suggested that chromosome aberrations increased nearly in proportion to the radiation dose [In this hypothesis, the relationship between radiation doses and the probability of stochastic effects of radiation, i.e., cancer incidence and genetic effects, is assumed to be linear and thus, radiation increases the incidence of cancer even at low doses. There is no threshold for such effects. This hypothesis was proposed in the 1940s\u20131950s, and Hermann Muller, a Nobel Prize winner, markedly contributed to the establishment of the hypothesis. He conducted an experiment using ion dose . He insiion dose . At thation dose . Thus, tion dose ,10, and ion dose .Drosophilia melanogaster, the dose\u2013response relationship was not linear but rather U-shaped, with a decrease in the mutation frequency at low doses, while it was considered linear in a repair-defective mutant strain [Drosophilia [After Muller\u2019s work, some succeeding studies also supported the linear relationship, but more recently, it was shown that the DNA repair capacity was closely related to the existence of a threshold in the dose\u2013response curve . In DNA-t strain . A thressophilia . In chrosophilia . Since mClassical and more recent data on the LNT theory were summarized in a review by Tubiana et al. , includiHormesis is a term used in toxicology; even highly toxic substances may exert stimulatory and beneficial effects at low doses or concentrations. All toxic compounds may have such hormetic effects, and it was found that carcinogens had effects to suppress cancer at low drug doses ,18. AccoBecause of the increasing interest in the effects of low-dose radiation, numerous laboratory studies have been carried out to date. Reviewing all of them is impossible; recently, a comprehensive review article was published, which summarized many but still only a small proportion of all previous studies . This reThe radioadaptive response is a phenomenon whereby small conditioning doses of ionizing radiation reduce the detrimental effects of subsequent higher radiation doses. Olivieri et al.  reportedRecently, Feinendegen  summarizThree major cellular defense systems have been proposed to explain the adaptive response: (1) protection against reactive oxygen species by antioxidants such as glutathione and detoxifying enzymes such as catalase and superoxide dismutase (SOD); (2) DNA repair, particularly for double-strand breaks, owing to the induction of repair enzymes; and (3) elimination of genomically damaged cells by immune defense mechanisms and apoptosis. They are considered to be common to the radiation hormesis phenomenon, and so it is reasonable that hormetic responses appear under adequate conditions. Reported mechanisms and phenomena of radioadaptive response (and radiation hormesis) at the molecular and cellular levels are summarized in Our group is interested in the growth-stimulating effects of low-dose radiation. Various stimuli are known to accelerate the growth of insects and plants. These stimuli include ultraviolet radiation, magnetic and electromagnetic fields, microwaves, ultrasound, and low-dose radiation ,38,39,40Tribolium) was reported more than 40 years ago [Drosophilia melanogaster [Extension of the lifespan of irradiated flour beetles  has been of concern in clinics. Epidemiological studies on this issue are discussed later. Simulating the clinical situations, biological experiments to evaluate the negative or positive effects were investigated. Miller et al.  investigTrp53+/\u2212 mice exposed to a single 10-mGy CT scan or gamma irradiation. CT-scanned mice lived longer than the control mice, and CT caused a significant increase in the latency of sarcoma and carcinoma. In another experiment from the same group, 4 Gy was administered first to the same mice and weekly CT scans were repeated 10 times [More recently, contradictory data have been reported. Lemon et al.  investigWhile the above-mentioned biological studies on low-dose effects comparing different groups of animals are considered to be associated with relatively large experimental errors, measurements of levels of enzymes, cytokines, and immunological parameters before and after radiation are less prone to such errors if the same animals and individuals are examined serially. Many studies reported increased activities of antioxidants such as SOD, glutathione, and catalase after low-dose radiation ,57,58,59The DNA repair capacity is increased by irradiation owing to the induction of DNA repair enzymes . At low In addition, increases in various immunological parameters, which enhance host immunity, have been reported. These include increases in CD4+ T cells and CD8 molecule expression , T-cell Survivors of the atomic bombings of Hiroshima and Nagasaki comprise one of the largest cohorts to study the effect of radiation, with data on about 120 thousand individuals available. There are many published data on the cohort ,69,70,71To add to the complicated and confusing situation, a recent report suggested that the doses received by atomic bomb survivors have been largely underestimated . HistoriMany studies have examined this issue. Most of them are anecdotal and some should be criticized. Several studies investigated the relationship between cancer incidence or mortality and amounts of natural background radiation ,75. TheyIn Japan, there is a report that inhabitants of the Misasa spa area, famous for radon production, had low cancer mortality rates; in particular, lung cancer mortality was much lower than that of the Japanese average . Such deIn Taiwan, a low cancer mortality rate was reported in residents of apartments contaminated with Cobalt-60, but in that study, the control group was not matched to the residents in the building . A subseA number of studies investigated the cancer incidence or mortality in people occupationally exposed to low-dose radiation. A report on nuclear industry workers from 15 countries suggested an overall increase in cancer mortality, but when the studied countries were analyzed separately, only the data from Canada showed an exceptionally high mortality rate; data from the other 14 countries did not show significant increases in cancer mortality . In addiIt is well known that high in the atmosphere, radiation levels from cosmic rays are marked, and pilots and cabin attendants are exposed to excessive natural radiation. A study of 19,184 male European pilots showed that cancer mortality of the pilots was lower than that of age-matched controls and the decrease was more marked in those receiving higher levels of radiation . They weIn the UK, the mortality of radiologists who registered with the radiological society since 1897 was studied . BritishAt least two papers have been published suggesting an increase in cancer (including benign tumors) incidence in individuals undergoing CT during childhood ,93. SoonThere is a well-known American study  that investigated the efficacy of lung screening by CT in former heavy smokers . The stuAn interesting case report was recently published . A patie228Ac and 77Br. The dose rate was 5 \u03bcGy/h for \u03b3 rays (measured by a survey meter) on the surface of the mats. Control (placebo) mats with the same physical property but no radioisotopes were also manufactured.In the last section of this article, we introduce unpublished data from Prof. Norinaga Shimizu  on a human study investigating the effects of a low-dose-radiation-emitting mat (hormesis mat), with permission from Dr. Shimizu. This was presented at the Japanese Society for Radiation Oncology Symposium for Cancer Control  but it will not be published in English. Low-dose-radiation-emitting mats were manufactured by Aoyama Stein Co., Ltd. . The raw materials are the same as those used for the sheets employed in the experiments involving silkworms , and thep < 0.05). Sleep latency and the physical, psychological, and neurosensory status were all improved in the hormesis mat group compared with the placebo group.Sixty healthy volunteers (30 men and 30 women) with a median age of 32 years  were randomly divided into a hormesis mat group (15 men and women) and placebo group (15 men and women). They were instructed to sleep on the mats every day. The volunteers underwent medical and physical checkups, and their serum reactive oxygen species levels were measured. The reactive oxygen levels at 3 months after starting the experiment were 3.1 and 9.4% lower on average than the initial levels in the placebo and hormesis mat groups, respectively, in men, and 3.1 and 8.5% lower, respectively, in women  period were observed in the hormesis mat group. Thus, the studies by Dr. Shimizu and colleagues demonstrated that continuous low-dose radiation during sleep yielded beneficial effects from various aspects.There have been numerous studies on the effects of low-dose radiation. Among the types of studies, the least reliable type may be epidemiological studies, since they are subject to many biases. Biological studies with sufficient numbers of subjects are more reliable. In particular, for studies comparing lifespan and cancer incidence of low-dose-irradiated subjects and controls to be reliable, large numbers of subjects are necessary. On the other hand, studies that investigate biological, biochemical, and/or physiological changes in the same subjects or individuals before and after low-dose radiation may be the most reliable. Many such studies have suggested that by low-dose radiation, host immunity, levels of radioprotective substances, and the DNA repair capacity are increased. These phenomena should lead to beneficial effects for living organisms. Data suggesting the beneficial effects of low-dose radiation are steadily increasing, and we believe that in the near future, it will be confirmed that there is at least no harm from low-dose radiation, and that low-dose radiation is beneficial to living organisms under specific conditions. Recently, the concept of exposome is spreading to assess life-course environmental exposures ,100, and"}
+{"text": "There continues to be significant debate regarding the most effective mode of ex situ machine perfusion of livers for transplantation. Subnormothermic (SNMP) and normothermic machine perfusion (NMP) are two methods with different benefits. We examined the metabolomic profiles of discarded steatotic human livers during three hours of subnormothermic or normothermic machine perfusion. Steatotic livers regenerate higher stores of ATP during SNMP than NMP. However, there is a significant depletion of available glutathione during SNMP, likely due to an inability to overcome the high energy threshold needed to synthesize glutathione. This highlights the increased oxidative stress apparent in steatotic livers. Rescue of discarded steatotic livers with machine perfusion may require the optimization of redox status through repletion or supplementation of reducing agents. Liver transplantation remains the only definitive cure for end-stage liver disease. Despite significant success and innovation in the field, there continues to be a severe shortage of organs that is unable to meet the need of transplant candidates. Machine perfusion technology has the potential to alleviate a significant portion of the shortage, with initial clinical trial results demonstrating lower organ discard rates in livers preserved by normothermic machine perfusion (NMP) compared to conventional static cold storage (SCS) .Machine perfusion has been incorporated into clinical practice by transplant centers around the world. However, there is a significant range of perfusion practices among different centers. Groups have reported their experience with NMP ,3,4, subFurther elucidation of these subsets of donor grafts and their physiology during machine perfusion is required to allow informed decision making by transplant surgeons. We compared the metabolomic profiles of steatotic livers perfused at subnormothermic versus normothermic temperatures to evaluate the advantages and disadvantages of each.p = 0.35) and 180 min  of perfusion, though they did not reach significance  at 180 min compared to pre-perfusion concentrations. Subsequently, principle component analysis (PCA) demonstrated similar composition of metabolites in pre-perfusion biopsies of steatotic livers, with divergence after initiation of SNMP or NMP. Interestingly, PCA of lipids clustered by mode of perfusion , the main cysteine donor involved in glutathione synthesis. NMP livers, however, demonstrated a gain in N-Ac stores during three hours of perfusion . Notably, there was marked depletion of tissue reduced glutathione (GSH) levels in SNMP livers , suggesting the immediate utilization of available glutathione after initiation of SNMP. Oxidized glutathione (GSSG) levels decreased to a ratio of 0.02 at 60 min of perfusion from pre-perfusion levels (p < 0.05), indicating degradation and utilization of glutathione components for other cellular processes. NMP livers were able to sustain their anti-oxidative capacity comparatively; GSH levels decreased after the initiation of perfusion  but to a lesser degree than SNMP livers (A significantly different anti-oxidative capacity was seen in the two groups. SNMP livers demonstrated a reduction in tissue N-acetylcysteine (N-Ac) levels  at 120 and 180 min of perfusion, respectively. Metabolite concentrations involved in the tricyclic acid cycle generally showed increased fold change ratios in SNMP livers and decreased ratios in NMP livers, consistent with the temperature-dependent rate of metabolic activity , and dicarboxylic fatty acids demonstrated significantly increased tissue concentrations with perfusion compared to NMP livers. Ceramides and dihydroceramides were significantly increased in both groups, while sphingolipids and monoacylglycerols were significantly decreased in SNMP livers .The debate regarding the best modality of machine perfusion continues. This study provides a metabolomic profile of discarded steatotic human livers during three hours of subnormothermic versus normothermic machine perfusion to help inform future perfusion studies. Our most significant finding is that steatotic livers undergoing SNMP are able to increase ATP and energy charge ratios; however, this appears to hinder the anti-oxidative capacity that would be required to overcome the oxidative injury encountered during surgical reperfusion at the time of implantation . Given tOne potential way to overcome glutathione depletion while capitalizing on ATP production is to combine perfusion modalities. Several groups have reported their experience with combined hypothermic and normothermic machine perfusion of livers with results indicating improved liver function and viability compared to NMP or SCS alone ,20. ThisWith respect to lipid metabolism, it is notable that steatotic livers appear to synthesize ceramides regardless of perfusion modality. Ceramides can be generated through the hydrolysis of sphingomyelin by the enzyme acid sphingomyelinase (ASMase) or through de novo synthesis in the endoplasmic reticulum. ASMase appears to be linked to key mechanisms involved in the regulation of steatosis, fibrosis, and lipotoxicity . The inhIn contrast, SNMP livers demonstrate significantly larger fold increases in cytochrome P450 (CYP450) eicosanoid products, 14,15-dihydroxyeicosatrienoic acid  and 8,9-dihydroxyeicosatrienoic acid . DHETs are the breakdown product of epoxyeicosatrienoic acids (EETs), which have been shown to possess significant vasodilatory and anti-inflammatory properties and play an important role in liver physiology. One limitation of our metabolomic analysis is that it does not include the concentrations of EETs or its counterpart 20-hydroxyeicosatraenoic acid (20-HETE), which has vasoconstrictive and pro-inflammatory properties , but if Several limitations are worth discussion. First, there is inherent heterogeneity in discarded organ research. Livers with similar profiles were submitted for metabolomic analysis in an attempt to control for these various factors, though variability was unavoidable. Notably, the two groups differed in macrosteatosis (more severe macrosteatosis in SNMP group compared to NMP), which could impact the functional and metabolomic changes observed. In addition, baseline comparison of lipidomic profiles prior to initiation of perfusion demonstrated pathway enrichment in phospholipid and secondary bile acid metabolism. Though the majority of the lipid subclasses demonstrated no enrichment, these pre-perfusion differences could certainly impact the observed changes during perfusion . This stThe use of metabolomics has numerous clinically relevant applications for transplantation. In this study, metabolomic profiling using mass spectrometry was used to examine differences between discarded livers undergoing SNMP versus NMP in order to assist in designing optimized perfusion conditions for steatotic grafts. Faitot et al. reported the use of real-time high-resolution nuclear magnetic resonance (NMR) metabolomic analysis in patients undergoing liver transplantation and demonstrated the ability to accurately predict EAD using lactate and phosphocholine (L + PC) levels . NotablyFourteen human donor livers with moderate (30\u201359%) or severe (>60%) macrosteatosis, declined for transplantation by all transplant centers in our donation service area, with consent for research, between August 2015 and April 2018 were included in this study. All donor livers were received through New England Donor Services (NEDS); no organs were procured from prisoners. Donors or their surrogates  provided informed consent for use of donor organs in research. The Massachusetts General Hospital Institutional Review Board (IRB) and the NEDS approved this study (No. 2011P001496), and all studies were carried out in accordance with IRB and NEDS approved guidelines. All donor livers were procured based on the standard technique of in situ cold flush using University of Wisconsin (UW) preservation solution. Procurement techniques for donation after brain death (DBD) or donation after circulatory death are previously described . Warm isUpon arrival to our perfusion lab, back table preparation of the livers was performed as described previously . 2 Therapeutics LLC, Souderton, PA, USA) were added to the base perfusate. Detailed composition is provided in the Liver grafts were perfused for three hours using a pressure and temperature-controlled perfusion device, Liver Assist . During SNMP, livers were perfused with a portal venous pressure (PVP) of 3\u20137 mmHg and mean hepatic arterial pressure (HAP) of 30\u201360 mmHg. The temperature of the perfusate was maintained between 20\u201322 \u00b0C. The perfusate consisted of Williams\u2019 medium E supplemented with hydrocortisone, insulin, and penicillin/streptomycin [1 min1 mL\u22121). Alanine aminotransferase (ALT) was determined using Piccolo Xpress Chemistry Analyzer . The volume of bile produced was recorded at 1-h intervals. Two wedge liver biopsies were collected immediately prior to perfusion and then hourly during perfusion. One tissue sample was snap-frozen in liquid nitrogen for future analysis and the other preserved in formalin for histology. Perfusion dynamics were recorded every 30 min. Vascular resistance was defined as perfusion pressure divided by flow rate . Tissue slides were evaluated for macrosteatosis content and preservation injury by a blinded expert pathologist (EOAH). Frozen liver biopsies from each time point (~25 mg) were pulverized in liquid nitrogen and analyzed for metabolic cofactors using a targeted multiple reaction monitoring analysis on a Sciex TripleTOF 6600 Quadruple Time-Of-Flight system, performed at the principle research institution. Metabolites were extracted using the protocol provided by Yuan et al. . ConcentTissue biopsies of three livers from each group were analyzed for 1600 compounds of known identity by Metabolon, Inc. . Principal component analysis, pathway enrichment scores, detailed methods, and statistical approach are provided in the p < 0.05. Wilcoxon\u2019s rank-sum (Mann-Whitney U) test and Fischer\u2019s exact test were used for continuous and categorical comparisons, respectively. Repeated measures data were analyzed using a random intercept mixed model with a categorical effect of time. If between-group comparisons were made, the categorical effect of group and the group by time interactions were added to the model. Statistical analysis was performed using Stata 15.1 . Demographic and perfusion data are presented as the mean \u00b1 standard error of the mean (SEM), unless otherwise specified, with statistical significance defined as Subnormothermic and normothermic machine perfusion of liver grafts are techniques of organ resuscitation that produce significantly different metabolomic profiles. Perfusion protocols that combine sequential temperature variation, guided by evidence-based metabolite replacement, can maximize the potential of both methods to improve organ quality for successful transplantation."}
+{"text": "P = 0.04; partial pressure of carbon dioxide perfusate, P = 0.04) and increased adenosine triphosphate levels 1.8\u2010fold. Following normothermic perfusion, those organs achieved lower tissue expression of markers of oxidative injury  and inflammation  compared with livers that had normothermic perfusion alone. All livers in the combined group achieved viability criteria, whereas 40% (2/5) in the normothermic group failed (P = 0.22). In conclusion, this study suggests that a combined protocol of hypothermic oxygenated and normothermic perfusions might attenuate oxidative stress, tissue inflammation, and improve metabolic recovery of the highest\u2010risk donor livers compared with normothermic perfusion alone.Hypothermic oxygenated perfusion (HOPE) and normothermic perfusion are seen as distinct techniques of ex situ machine perfusion of the liver. We aimed to demonstrate the feasibility of combining both techniques and whether it would improve functional parameters of donor livers into transplant standards. Ten discarded human donor livers had either 6 hours of normothermic perfusion (n = 5) or 2 hours of HOPE followed by 4 hours of normothermic perfusion (n = 5). Liver function was assessed according to our viability criteria; markers of tissue injury and hepatic metabolic activity were compared between groups. Donor characteristics were comparable. During the hypothermic perfusion phase, livers down\u2010regulated mitochondrial respiration (oxygen uptake,  Potential beneficial protective mechanisms of machine perfusion have been demonstrated for both hypothermic and normothermic perfusion techniques during preclinical experiments and pilot clinical studies.Hypothermic oxygenated perfusion (HOPE) may permit mitochondrial functional recovery, increasing cellular adenosine triphosphate (ATP) levels, and mitigate the injury to the tissue that occurs during rewarming.Data from our research on NMP perfusions showed that a proportion of poor\u2010quality livers exposed to prolonged cold storage do not recover their function and fail our viability criteria (unpublished observations). Although hypothermic perfusion and normothermic perfusion were developed as distinct strategies, we hypothesized that HOPE might be seen as a beneficial therapeutic intervention by restoring liver metabolism prior to a period of normothermic perfusion, which permits liver viability testing. The present study aimed to assess the feasibility of a protocol combining HOPE with NMP and to investigate its potential benefits over NMP alone.The study was designed to compare 2 perfusion strategies to restore function of high\u2010risk ECD livers, following a period of SCS, within the certified 6\u2010hour timeframe allowed by the used perfusion device.The study endpoints were evaluation of hepatocellular injury and liver function assessment. Ten discarded human donor livers were consecutively assigned to 2 study groups, each consisting of 5 organs and all perfused for 6 hours. Livers in 1 group were exposed to normothermic perfusion alone , while the other group underwent 2 hours of HOPE, followed by 4 hours of normothermic perfusion (HOPE + NMP group). Figure All included livers were procured with the intention of transplantation according to the National Organ Retrieval Service standards. The organs were declined for clinical use by all the UK liver transplant centers and subsequently offered for research purposes. The livers were preserved in University of Wisconsin fluid under standard clinical practice of SCS prior to commencing perfusion. The study protocol was approved by the appropriate institutional review committee. Ethical approval for the study was obtained by the London\u2010Surrey Borders National Research Ethics Service and the National Health Service Blood and Transplant Ethics Committee .The liver preparation for the perfusion was carried out using a standard clinical back\u2010table procedure as described elsewhere.HOPE was performed via PV only, using 3 L of Belzer MPS University of Wisconsin Machine Perfusion Solution  with the temperature set at 10\u00b0C. The target flow was 0.1 mL/minute/g of liver with a maximum pressure of 3 mm Hg. The target oxygen perfusate pressure was 80\u2010100 kPa. After 2 hours of HOPE, the perfusion was stopped and the liver temporarily placed on ice. The system was then drained and subsequently refilled with the NMP perfusion solution. The HA and common bile duct were cannulated and then the normothermic perfusion commenced. The perfusate exchange, from the end of HOPE to the start of NMP, took an average of 20 minutes.The perfusion fluid for NMP consisted of 1000 mL (4 units) of an acellular, polymerized bovine hemoglobin\u2010based oxygen carrier Hemopure  complemented with human albumin solution and additional supplements as described in Supporting Table The target flow was 0.25 mL/minute/g liver tissue on the arterial side and 0.75 mL/minute/g liver tissue in the venous circuit. To achieve these flows, perfusion pressures on the device were adjusted between 30\u201050 mm Hg (mean pressure) on the arterial side and 8\u201010 mm Hg on the PV. The temperature was initially set at 20\u00b0C and increased incrementally to 37\u00b0C within 30 minutes of starting NMP. The target perfusate oxygen pressure was 40 kPa.Liver biopsies were taken before commencing the perfusion (t = 0), after finishing the HOPE perfusion (t = 2 hours), and on completion of NMP , pH, base excess, bicarbonate, O2 saturation, hemoglobin, hematocrit, sodium, potassium, chloride, calcium, glucose, and lactate concentrations.Arterial and venous perfusates were assessed using a Cobas b 221  point\u2010of\u2010care system blood gas analyzer. Parameters included partial pressure of oxygen and partial pressure of carbon dioxide  as described elsewhere.The viability of the organ was assessed at the end of the perfusion by our unit\u2019s clinical criteria, based on perfusate lactate levels falling to concentrations of <2.5 mmol/L within 6 hours, in combination with evidence of bile production, stable vascular flows, and homogeneous parenchymal perfusion.Menghini needle and wedge biopsies obtained prior to perfusion and at the end were fixed in formalin, processed, and embedded in paraffin. Thereafter, 4\u2010\u03bcm sections were cut and stained with hematoxylin\u2010eosin and periodic acid\u2013Schiff (PAS). Hematoxylin\u2010eosin sections were semiquantitatively graded for ischemic\u2010type coagulative necrosis, large\u2010 and small\u2010droplet macrovesicular steatosis, and preexisting acute or chronic liver disease. The PAS\u2010stained sections were scored for percentage of hepatocytes depleted of glycogen, and the variation between the beginning and the end of the perfusion compared across groups. Histological assessment was conducted by an experienced liver transplant pathologist without prior knowledge of the designated perfusion category or outcome.Immunohistochemistry was performed on formalin\u2010fixed paraffin\u2010embedded sections to assess surrogate markers of oxidative injury and tissue inflammation.Uncoupling protein 2 (UCP\u20102), a mitochondrial inner membrane protein that uncouples the electron transport chain from oxidative phosphorylation. Elevated uncoupling protein 2 expression is associated with increased reactive oxygen species (ROS) production.4\u2010hydroxynonenal (4\u2010HNE) as a marker of cell membrane phospholipid peroxidation.For oxidative injury, we assessed expression of the following:Cluster of differentiation (CD)14, a lipopolysaccharide receptor, which is part of the toll\u2010like receptor 4 signalosome. It is essential for activation of the toll\u2010like receptor 4 via the recognition of ligands such as damage\u2010associated molecular patterns (DAMPs) known to be up\u2010regulated during ischemia/reperfusion injury.CD11b is an integrin on the surface of leukocytes; up\u2010regulation on its expression indicates activation of the cells by substances including ROS.Vascular cell adhesion molecule 1 (VCAM1) expression is up\u2010regulated on vascular endothelial cells and Kupffer cells when activated by ROS and proinflammatory cytokines during ischemia/reperfusion injury.For the assessment of tissue inflammation, the following markers were analyzed:All primary antibodies were detected using specific ImmPRESS Excel Amplified horseradish peroxidase Polymer Staining Kit specific to the respective mouse or rabbit immunoglobulin isotype. A list of primary antibodies and the dilution used is provided in the Supporting Materials.Four pictures of each section excluding the edges were randomly selected for analysis (\u00d7400 magnification). For UCP\u20102, 4\u2010HNE, CD14, and CD11b, a semiquantitative scoring system, the modified immunoreactive score (IRS),VCAM1 tissue expression was assessed by image analysis using an established system of color differentiation , and the variation in the percentage of the positive area of staining (\u0394%VCAM1) over time was compared between groups.Quantification of ATP levels was done by homogenization of liver tissue with the concentration determined using the ATP Bioluminescent Assay kit . More details are provided in the Supporting Materials.P < 0.05. GraphPad Prism  software was used for all statistical analyses and graph creation.Continuous variables were expressed as median with interquartile range (IQR) and categorical variables as absolute numbers with percentage frequencies. Comparisons between groups were performed using Fisher\u2019s exact test for categorical variables, Mann\u2010Whitney U test for independent continuous variables, and Wilcoxon signed rank test for repeated measurements over time on the same sample. The statistical level of significance was set at 2 (21\u201031 kg/m2). Median cold ischemia time (CIT) was 510 minutes (446\u2010682 minutes) for DCD, and 491 minutes (454\u2010586 minutes) for donation after brain death (DBD). The median donor risk index (DRI) was 2.2 (1.9\u20102.6). There were no significant differences in donor demographics between the groups as shown in Table P = 0.78). There was a trend toward NMP livers having shorter SCS preservation times for DCD . The detailed donor and liver data are presented in Tables Seven (70%) livers were from donation after circulatory death (DCD). The median donor age of the entire cohort was 52 years , and the body mass index was 25 kg/m2 2\u201031 kg/m2P = 0.03). In contrast, the arterial flows in the NMP livers increased more rapidly during the rewarming phase and remained higher throughout the perfusion , while the vascular resistance patterns and levels were similar. The detailed data are presented in Fig. Portal flow patterns and end\u2010perfusion volumes differed between the groups. During the initial 60 minutes of the NMP group perfusion, the livers\u2019 portal flow rate increased rapidly and plateaued after 180 minutes. For the HOPE + NMP group, steady low\u2010volume flow rates approximating 300 mL/minute were achieved during the HOPE phase. The flow rate subsequently increased for the first 2 hours of rewarming during the NMP phase. The median flow rates in the HOPE + NMP group exceeded the NMP group after 210 minutes and continued to increase until the end of the perfusion  and then declined steadily (0.006 [0.001\u20100.007] kPa/g liver/L perfusate) until the end of the 2\u2010hour perfusion (P = 0.04). There was a concomitant decrease in the median PCO2 dissolved in the perfusate from time 0 to 2 hours of hypothermic perfusion (1.3 [1.2\u20101.9] to 0.7 [0.6\u20100.8] kPa); P = 0.04). This change in mitochondrial respiration was associated with a median 1.8\u2010fold  increase in tissue ATP levels.During the HOPE phase, livers demonstrated a decrease in the oxygen uptake rate and COP = 0.19).The oxygen consumption in the NMP group increased sharply within the initial 60 minutes with distinct patterns in viable compared with nonviable groups Fig. . In the P = 0.31). However, there was a significant difference in the incremental rate between viable and nonviable livers in the NMP group . Data are shown in Fig. Both groups had similar ATP stores at the end of the 6 hours of perfusion (P = 0.17). In the nonviable livers, lactate levels peaked after 60 minutes with minimum levels achieved by the end of the perfusion of 8.5 mmol/L. A total of 3 (60%) livers in the NMP group and 5 (100%) in the HOPE + NMP were deemed viable (P = 0.22). Details of the achievement of viability parameters are presented in Table P = 0.13).The lactate levels and clearance dynamics differed between the groups. In the NMP group, lactate levels peaked after 60 minutes, reached the viability criteria within 150 minutes, and subsequently remained low. In the HOPE + NMP livers, it did not change through the hypothermic phase, and it slowly increased in the rewarming phase with a subsequent rapid clearance just after 2 hours of NMP. The end\u2010perfusion lactate levels were similar in viable livers from the NMP group and the HOPE + NMP group .The glucose levels slowly increased within the initial 120 minutes of perfusion in the NMP group, followed by slow clearance until the end of the perfusion. In the HOPE + NMP group, they slowly increased through the hypothermic perfusion, followed by a drop (related to the perfusate fluid replacement) with a subsequent slow increase, and then decreased, reaching similar figures to the NMP group at the end of the perfusion . Bile pH  and bile glucose  were comparable between the groups at the end of the perfusion. Data are represented in Fig. Bile production at 6 hours of the perfusion was similar between both groups , and only 1 liver from each group presented with steatohepatitis. The levels of hepatocyte glycogen depletion from the start to the end of the perfusion period were similar in both groups  with a positive value reflecting an increase in the percentage of hepatocytes depleted of glycogen. Data are presented in Fig. Routine histological assessment showed no observable differences between the groups on the biopsy prior to the perfusion. At the end of the perfusion, none of the livers presented with areas of parenchymal necrosis, congestion, or cytoplasmic vacuolization. The median large\u2010droplet macrovesicular steatosis percentage was not significantly different between the groups  and oxidative stress\u2013mediated cellular injury (4\u2010HNE) expression reflecting lipid peroxidation . Both proteins were present predominantly in hepatocytes. Similarly, they showed decreased expression of CD14 at the end of the perfusion compared with commencement. A statistically significant opposite trend was seen for the livers perfused using NMP only . The intensity of the tissue inflammatory response was shown by different patterns of nonparenchymal cell staining for CD11b, which was present on polymorphonuclear leukocytes (neutrophils and monocytes/macrophages). There was a significant difference in its level of expression between the 2 groups throughout the perfusion , suggesting a potential beneficial effect of the combined protocol. Intrahepatic endothelial cells play a key role in reperfusion injury by promoting leukocyte adhesion and retention. To evaluate the (cytokine\u2010mediated) activation of endothelial cells, the percentages positive areas of VCAM1 staining were determined. VCAM1 expression decreased over time in both groups, but this was more pronounced in livers that had undergone HOPE . Data are presented in Fig. Immunohistochemical analysis showed that livers subjected to NMP alone expressed higher levels of tissue markers of oxidative injury and inflammation compared with those subjected to HOPE + NMP by the end of the perfusion run. Livers that had HOPE showed a significant reduction in expression of markers associated with mitochondrial ROS production (UCP\u20102; NMP \u0394IRS versus HOPE + NMP \u0394IRS; 1 versus \u20132; r injury \u2010HNE exprMachine perfusion was developed to minimize damage during organ preservation, and early clinical experience proved its superiority over SCS and positive impact on extended criteria liver utilization.Our team became an early adopter of the NMP technique, and we observed that a proportion of livers failed to recover their function.Although the use of HOPE with subsequent NMP was reported by the Groningen group previously, their experiment was aimed at assessing benefit of HOPE compared with SCS and used the NMP phase to simulate in vivo reperfusion.Our study represents a novel approach and shows unique data, as it assesses the viability of human donor livers using clinically validated criteria.Although there has been extensive mechanistic research using animal models showing a down\u2010regulation in the tissue inflammatory responses, including activation of endothelial and resident immune cells of the liver, the evidence from human livers, especially those from the highest\u2010risk donor pool, is very scarce.NMP provides a near\u2010physiological environment for the liver by supplying oxygen and nutrients at normothermic condition, enabling the organ functional assessment. Our previous work on end\u2010ischemic NMP of declined human livers allowed us to define the viability criteria employed for transplantation of discarded human livers.The use of an acellular hemoglobin\u2010based oxygen carrier was shown by our group and others to be a suitable alternative to the use of packed red cells in the context of NMP.The feasibility of combining variant perfusion techniques has been suggested before, though the advantages, caveats, and logistics aspects of different combinations are yet to be seen.The present study has some caveats. It is not a transplant model because this would not be clinically possible in the United Kingdom at this early investigative stage. Although the reperfusion injury could be simulated by a subsequent NMP with whole blood, we opted to study the organ functional recovery only, and we have applied our current viability criteria to define its potential transplantability. Those criteria have been used by our team in clinical trials to safely transplant discarded human livers, allowing us to compare the findings and predict transplantability and clinical relevance of our preclinical proof of concept studies.Using an acellular oxygen carrier fluid also precludes any direct comparison of our data with frequently published machine perfusion studies\u2019 endpoints, including perfusate transaminases.In conclusion, this proof of concept study demonstrated that the combination of sequential HOPE and NMP is not only feasible, but that it may potentially improve the functional recovery of high\u2010risk ECD livers compared with NMP alone. Although we do not suggest that this is an optimized protocol, this novel approach might be particularly beneficial for DCD organs. Further studies are needed to explore whether the combined protocol confers other benefits, such as the reduced biliary complications of HOPE or the safety of prolonged perfusions of NMP.\u00a0Click here for additional data file."}
+{"text": "Eriosoma lanigerum [Hausmann]; Hemiptera: Aphididae), a worldwide pest of apple , occur both below- and aboveground. These spatially subdivided subpopulations encounter different abiotic conditions, natural enemies, and control tactics. Restricting movement between them might be an effective management tactic to decrease woolly apple aphid persistence and abundance. We examined this possibility in the field, using sticky barriers to restrict upward woolly apple aphid movement to tree canopies, and in the greenhouse, using mulches and sand amendments to restrict downward movement to roots. In the field, blocking aphid movement up tree trunks did not decrease the number of colonies in tree canopies. Instead, sticky-banded apple trees had higher aphid colony counts late in the study. Earwigs, which are woolly apple aphid predators, were excluded from tree canopies by sticky bands. In the greenhouse, fewer root galls (indicative of aphid feeding) occurred on trees in sandy potting media and on those with mulch (wood chips or paper slurry). Our results suggest that upward movement is less important than other factors that affect aboveground aerial woolly apple aphid population dynamics. In addition, apple orchards planted in sandier soils or with mulches may be partially protected from woolly apple aphid root feeding.Movement of insect pests between spatially subdivided populations can allow them to recolonize areas where local extinction has occurred, increasing pest persistence. Populations of woolly apple aphid ( Many pest species move into crops from habitats that provide spatial or temporal refuge from control tactics, such as neighboring fields or natural habitats ,2. MovemFor pests that move into and out of the soil, restricting movement might reduce their abundance in both environments. This may be an effective tactic, as underground (edaphic) pest populations are often difficult to manage and can be a source of aboveground  populations . BarrierEriosoma lanigerum [Hausmann]; Hemiptera: Aphididae) is a worldwide apple  pest with nymphs (\u2018crawlers\u2019) that can move into and out of the soil year-round, although movement is reduced during winter ), a pest on grape roots, are thought to be similarly suppressed by sandy soil [Soil sand content and mulches could affect woolly apple aphid edaphic abundance in the field, but their management utility is currently unclear. Marcovitch  tested endy soil ,50,51. Ondy soil . Wood chndy soil  found thWe conclude that apple orchards on sandy soils or ones which use mulches are potentially less susceptible to woolly apple aphid root infestation, but aerial woolly apple aphid populations are typically not strongly affected by movement from edaphic populations. Recently released woolly apple aphid resistant rootstocks from the Geneva breeding program  are a prS1 Table(CSV)Click here for additional data file.S2 Table(CSV)Click here for additional data file.S3 Table(CSV)Click here for additional data file."}
+{"text": "This paper considers multiple structural designs for photonic crystal surface emitting lasers operating at key wavelengths. Initially a structure from Williams et al. is modelled, the structure is modified to include an additional GaAs waveguide layer  and to include an additional PC layer (termed double decker). These structures are modelled by a combination of coupling calculation and waveguide modelling and are compared to the original structure. We show that both of these schemes give an increase in coupling, but present fabrication challenges. Next, we model standard laser structures operating at key wavelengths  where a photonic crystal is located above the active region and explore the effect of increasing thickness of photonic crystal. We find that increasing the thickness increases the coupling coefficient but not true for the full range of thicknesses considered. This study allows a more universal comparison of the use of all-semiconductor, or void containing PCSELs to be conducted and we find that the realisation of all semiconductor PCSELs covering a wide range of material and wavelengths are possible. There has been considerable recent interest in photonic crystal surface emitting lasers  above the PCSEL, and the use of a PC above and below the active element. Trends are assessed, and comparisons are made between all-semiconductor and void-containing PCSELs. General guidelines are deduced for PCSEL waveguide design. We show that the inclusion of a ballast layer increases the PC coupling for both all-semiconductor and void containing structures but that all-semiconductor PCSELs still have a higher coupling. The double decker structure is shown to increase the PC coupling and is the first structure in this study to give a higher coupling for void containing PCSELs than for all-semiconductor PCSELs, however we highlight the complexity in manufacturing such a device.We go on to consider PCSEL designs for different material systems and wavelengths. The wavelengths considered are 405\u00a0nm  method and models relying on coupled mode theory. FDTD method is time and computer memory consuming. Models based on coupled mode theory were initially developed for modelling of distributed feedback (DFB) lasers. These models are less demanding and in recent years they have been successfully applied to PCSELs  (Johnson and Joannopoulos rom Eqs.\u00a0 and 2)  a,b,c,d rom Eqs.\u00a0, where K1 (black) and K3 (red), for a PC consisting of InGaP/GaAs on a square lattice with a circular lattice, for a range of atom radii from 0 to 0.45 r/a. The dependence of K3 has two local maxima at r/a of 0.2 and 0.4, they correspond to band diagrams with largest frequency differences between bands shown in Fig.\u00a0Figure\u00a01, the coupling coefficient generally increases as atom radius increases until a radius of 0.4, when the coupling decreases again. For K3, the coupling coefficient has a double peak at r\u00a0=\u00a00.15a and r\u00a0=\u00a00.4a. The coupling coefficients are not plotted at a radius of ~0.3a as this corresponds to a change in character of the band structure and the assignment bands is complicated as the bands cross . Figures a\u2013c show the mode profile of an all-semiconductor PCSEL (dashed line) and void containing PCSEL (solid line) where the atom radius of the PC is 0.1a, 0.3a and 0.45a respectively. The mode of the void containing structure are distorted away from the PC, which is attributed to the low refractive index of the PC in the void containing case, which results in low mode overlap with the PC.Figure\u00a0PC is mode overlap with the PC, \u2206n is the refractive index contrast and \u03bb is vacuum wavelength). Figure\u00a0From the mode profile the coupling coefficient is calculated using 1 (black) and K3 (red) for (a) void containing and (b) all-semiconductor PCSEL, for a range of atom radius from 0.05 to 0.45a. For the void containing structure, K3 has two peaks one at 0.15a and one at 0.4a, while K1 has a global peak at 0.175a. The all semiconductor PCSEL again has two peaks for K3, at 0.2a and 0.45a, while the K1 has a peak at 0.45a. The maximum peak in K1 is 500\u00a0cm\u22121 for the void containing structure and 1300\u00a0cm\u22121 for the all-semiconductor case. In the case of Fig.\u00a0av pushes the mode away which dominates, giving a relatively low coupling coefficient. For low r/a void containing PCSELs have higher coupling coefficients, while all semiconductor PCSELs have a higher coupling at large r/a.In this section we combine the two models mentioned previously to consider the effect of both the PC and the waveguide on the coupling coefficient. Here the coupling coefficient calculated for the PC from the band-structure and Eqs.\u00a0 and 2)  2) is mu1 of (a) void containing and (b) all-semiconductor PCSEL where the atom radius is increased from 0.05 to 0.45a, for PC thickness (L) of 50\u00a0nm (black), 150\u00a0nm (red), 300\u00a0nm (green) and 400\u00a0nm (orange). For the void containing structure the peak coupling occurs at either 0.15a or 0.25a, and coupling decreases as PC thickness increases, this is due to the low refractive index of the PC layer further distorting the mode away from the PC layer. The all-semiconductor structure has a peak at 0.4a, increasing PC layer thickness (L) increases the coupling, this can be attributed to the PC layer having increased volume for the mode to couple to.It is expected that the thickness of the PC layer will have an effect on the coupling coefficient of a PCSEL due to the change in modal structure and overlap integral. For the moment we ignore the need for the PC to be of specific thickness to ensure constructive interference in the far-field. Figure\u00a01, in this case the void containing structure has two peaks at 0.125a and 0.4a, the peak coupling occurs for PC layer thickness of 150\u00a0nm. The all-semiconductor structure has a local peak in coupling at 0.2a and a global peak in coupling at 0.45a, as the PC thickness increases the coupling also increases.Figure\u00a0av of the PC layer being similar to the refractive index of the waveguide. In the void containing PC case, a thicker PC layer has detrimental effects due to the low nav. Our observed maxima in coupling at r/a\u00a0=\u00a00.2 for 150\u00a0nm PC thickness, is in agreement with observed values in the literature.In general, for this structure, we find that an increase in PC layer thickness has a monotonical improvement at an r/a of 0.4, this is enabled by the nAs discussed previously, PCSEL structures containing voids have lower coupling of the optical mode to the photonic crystal as compared to their all-semiconductor counterparts. This is attributed to the low refractive index of the PC layer \u201cpushing\u201d the optical mode away from the PC region. In order to address this, we consider a structure which includes an additional p-type GaAs waveguide region above the PC region  and an all-semiconductor PCSEL (b), for ballast layer thicknesses from T\u00a0=\u00a00\u2013500\u00a0nm. For the void containing PCSEL peak coupling occurs at r\u00a0=\u00a00.2a. As ballast layer thickness (T) increases the coupling increases to a maximum of ~800\u00a0cm\u22121 at T\u00a0=\u00a0100\u00a0nm, as T is increased beyond 100\u00a0nm coupling decreases. For the all-semiconductor PCSEL the ballast layer reduces the coupling coefficient, and as ballast layer thickness is increased the coupling decreases.Figure\u00a03, in both cases a double peak is observed and coupling is increased as T is increased from 0 to 100\u00a0nm then decreases as the ballast layer thickness is increased further. For the void containing PCSEL peaks occur at r\u00a0=\u00a00.15a and r\u00a0=\u00a00.4a, while for the all-semiconductor PCSEL peaks occur at 0.2a and 0.45a.Figure\u00a0The ballast layer has been shown to distort the mode profile of the PCSEL, for both the all-semiconductor and the void containing PCSEL a ballast layer of 100\u00a0nm gives the highest coupling. A ballast layer of suitable thickness has a significantly advantageous effect for void containing PCSELs at an r/a of 0.2. For all-semiconductor PCSEL, a marginal effect is observed with no detriment to the structural design. This bodes well for a one step epitaxial process in Williams et al.  where T The final structure considered in this section, consists of two PC regions located above and below the active region, first proposed by Kurosaka et al.  The inteFigure\u00a01 as atom radius increases from 0.05a to 0.45a, for a void containing (a) and an all-semiconductor (b) double decker PCSEL, for separation distance (L) from 0 to 200\u00a0nm. For the void containing PCSEL with separation of 0 and 50\u00a0nm the coupling is 0 above 0.2a and 0.3a, respectively, because there is no bound mode in this case, for separation distance >50\u00a0nm coupling is optimum at r\u00a0=\u00a00.25a and decreases as separation decreases. For the all-semiconductor PCSEL the coupling is greatest at r\u00a0=\u00a00.4a, as separation (L) increases the coupling decreases.Figure\u00a03, in both cases a double peak is observed and coupling is decreased as L is increased from 0 to 200\u00a0nm. For the void containing PCSEL peaks occur at r\u00a0=\u00a00.15a and r\u00a0=\u00a00.4a, while for the all-semiconductor PCSEL peaks occur at 0.2a and 0.45a. The double decker void containing PCSEL has peak coupling (K3) of 1000\u00a0cm\u22121 at r\u00a0=\u00a00.15a, this is a two fold increase in coupling compared with the original structure.Figure\u00a01 to K3, the effect of comparative ratio has been discussed elsewhere  can be maintained. For void containing structures there is a tendency for optimal coupling to occur at radius to be at ~0.2 r/a, this is in contrast to the all-semiconductor PCSEL where optimal coupling occurs at ~0.4 r/a. The addition of a ballast layer increases the peak coupling values for both all-semiconductor and void containing PCSELs, with the largest effect for a ballast layer thickness of 100\u00a0nm. The inclusion of a second PC layer (double decker PCSEL) leads to an increase in coupling coefficients, particularly with void containing structures where the low average refractive index of the PC layer strongly confines the mode between the PC layers within the active region. However, it is worth noting that the realisation of a double decker structure poses significant fabrication issues.In this Section\u00a0980\u00a0nm PCSELs based on GaAs were simulated by combining waveguide and band structure modelling. We find that nnav (where \u03bb is the vacuum wavelength and nav is the average refractive index in the PC).In this section we consider PCSEL designs for different material systems and wavelengths. The wavelengths considered are 405\u00a0nm (AlInGaN) , an 80-nm-thick undoped GaN layer, a 20-nm-thick p-A0.16Ga0.84N electron-blocking layer (EBL), a 115-nm-thick p-GaN layer, a PC layer, a 40-nm-thick p-GaN layer and a 115-nm-thick p+-GaN contact layer. The MQW active zone consists of three 2.5-nm-thick In0.09Ga0.91N well layers and 7.5-nm-thick GaN barrier layers. The PC region is 220\u00a0nm thick and consists of GaN/Al0.11Ga0.89N with a 50% fill factor. The mode profile is shown overlaid on the device structure  have shown promise in a range of applications including security applications such as target illumination and industrial applications such as gas sensing . The void containing structures all had peak coupling at atom radius r\u00a0~\u00a00.2a, while the all-semiconductor structures had peak coupling at r\u00a0=\u00a00.4a. By including a ballast layer within the structure the coupling was increased for ballast layer thickness <100\u00a0nm. The double decker structure gave the largest increase in the coupling, but fabricating such a structure would be problematic.In this paper we have considered three structures, a structure from Williams et al. , a ballaThree PCSEL structures with emission spanning the UV to mid-IR  have been considered. To a first approximation the device design is an existing edge emitting laser structure, with a PC layer in the upper waveguide cladding. All three structures have been modelled as a 1D waveguide and PC coupling coefficients have been calculated. We show that in each case it is possible to obtain strong mode overlap with both the photonic crystal and the active regions. The realisation of PCSELs spanning a full range of key laser wavelengths will allow the many advantages of PCSELs to be implemented into a range of laser systems."}
+{"text": "Carbon nanotube yarns (CNY) are a novel carbonaceous material and have received a great deal of interest since the beginning of the 21st century. CNY are of particular interest due to their useful heat conducting, electrical conducting, and mechanical properties. The electrical conductivity of carbon nanotube yarns can also be influenced by functionalization and annealing. A systematical study of this post synthetic treatment will assist in understanding what factors influences the conductivity of these materials. In this investigation, it is shown that the electrical conductivity can be increased by a factor of 2 and 5.5 through functionalization with acids and high temperature annealing respectively. The scale of the enhancement is dependent on the reducing of intertube space in case of functionalization. For annealing, not only is the highly graphitic structure of the carbon nanotubes (CNT) important, but it is also shown to influence the residual amorphous carbon in the structure. The promising results of this study can help to utilize CNY as a replacement for common materials in the field of electrical wiring. To fulfill future claims on everyday applications for higher efficiency, new materials with improved physical properties and lower production costs are necessary. One of the most promising candidates for these materials is carbon nanotubes (CNT). These one-dimensional tubular carbon structures have been shown to possess an impressive array of physical properties on the scale of individual tubes , includiThe main factors defining the electrical conductivity of CNY are the intrinsic electrical properties of the CNT and the contact resistant between adjacent CNT. Different groups have shown that adjusting the CNT type used for spinning can improve the electrical properties of these yarns. An ideal CNY would consist purely out of long metallic single walled CNT and reach conductivity as high as an individual CNT . Other wIn this work, we provide a systematical study on the effects of annealing and functionalization by acid treatment to CNT yarns. Hereby different acids were tested for functionalization as well as annealing temperatures up to 2500 \u00b0C.The pure multiwalled CNT (MWCNT) array was shown to consist of CNT with 2 to 6 walls with the majority of CNT possessing 2 to 4 walls . The diaFor improvement of the properties of CNY and therefore the electrical conductivity the CNY were annealed under an Argon atmosphere at 1000, 1500, 2000 and 2500 \u00b0C for 30, 60 and 120 min . The appD/IG ratio drops from 0.83 to 0.23 for samples annealed at 2500 \u00b0C for 2 h , HCl (37%), HNO3 (65%) and half concentrated HNO3 (32%).Oxidative acids like HNOrify CNT  whereas rify CNT . The eff2O2 treatment. Even after 216 h, the yarn structure does not differ significantly from the original sample .In addition to the effect of acid treatment on the surface and the appearance of the yarn, the effects on the structure, diameter and electrical conductivity of the yarn were investigated. 3 is different from the other tested acids. There is a strong variation of the ID/IG ratio but only a relatively small fluctuation of the average yarn diameter observed with this treatment. However, it should be noted that the measured diameter of the yarn is subject to high fluctuations, as indicated by the large error bars. These variations are due to the severe deformation and partial detachment of parts of the outer yarn layer. Over the considered experimental period (3 to 216 h), the ID/IG ratio is subject to large variations ranging from 0.68 to 0.88. This, in turn, shows the strong impact of concentrated HNO3 on the structure of CNT materials. Electrical conductivity drops after the strong increase at 3 h and drops to 300 S/cm after 216 h. The maximum value for the conductivity is 642 S/cm, which represents an increase of more than 110%. For verification, a trial was carried out with only one hour of treatment, which only resulted in a value of 416 S/cm. With this, it appears that 3 h represents an optimal treatment time with concentrated HNO3 for the yarn used in this work. The drop of conductivity after 3 h treatment could be explained by the strong oxidative nature of concentrated HNO3, this results in a strong defect introducing behavior. This appears to be proof that the action of introducing functional groups on the surface of CNY is not as effective as claimed as increasing treatment time should result in a noticeable positive effect on conductivity by increasing the charge carrier density between the CNT. In addition to this, a stable densification of the yarn through the functional groups and introduced stronger dipol-dipol and H-bridge bonds should be observable [The behavior of the yarn after treatment with concentrated HNOservable .2O2, HCl and half concentrated HNO3) is less dramatic than with concentrated HNO3, with the ID/IG ratio being only minimally affected. It is even decreased by the influence of HCl. After 216 h, the values are in the range of 0.72 to 0.78. This decrease is explained in literature by the removal of carbonaceous impurities during acid treatment [3 treated sample with the electrical conductivity increases with decreasing diameter and vice versa. The maximum values reached after 3 h are for H2O2, HCl and half concentrated HNO3 587, 551 and 541 S/cm, respectively. In the case of H2O2 a rise in electrical conductivity (518 S/cm) can be observed after a treatment time of 216 h. However this effect has not yet been explained and requires further investigation. It might be assumed that temperature fluctuations and the influence of light during the experiment leads to a decomposition of H2O2 and formation of hydroxyl radicals that can promote the formation of functional groups on the surface of the CNT.The influence of the other acids  array. The MWCNT array were grown by chemical vapor deposition as described by Iijima et al. . The parAnnealing of CNY was conducted in a high temperature furnace  under an Argon atmosphere and normal pressure. For this treatment, CNY were put into closed graphite vessels. Sections of the yarns were annealed at 1000, 1500, 2000 and 2500 \u00b0C for 30, 60 and 120 min.For functionalization, a variety of different acids were used including concentrated  and half concentrated  nitric acid, conc. hydrochloric acid  and conc. hydrogen peroxide . Pieces of 3 cm long CNY were put into glass vials with an excess amount of acid (5\u201310 mL). The acid treatment was carried out at 25 \u00b0C for 3 to 216 h for each of the acid treatment processes. Directly after each treatment, the samples were washed with water and dried over night at 108 \u00b0C.CNY were characterized by SEM  and TEM  before and after each annealing and acid treatment. SEM and TEM investigations were conducted using a cathode voltage of 15 kV and 80 kV respectively. For TEM investigations we spread the CNY in a mechanical way using tweezers to individualize several CNT. These CNY samples were fixed on a copper grid with special TEM glue. We investigated serval parts of the samples on the basis of more than 20 SEM images and about 75 TEM images. For statistical analysis of the CNT diameter a well as the number of walls we evaluated were min. 100 CNT per sample .\u22121 using a wavelength of 514.5 nm . ID/IG ratios were calculated from Raman spectra by dividing the intensity of the D-band through the intensity of the G-band. For each sample, 5 Raman measurements were conducted. Electrical conductivity was measured between 5 and 295 K with a Nanovoltmeter  using the four-point measurement method. Here the current will be operated by two contacts at the ends of the CNY whereas the voltage will be measured by two additional contacts, which are located between the two current supply contacts (\u03c3) was calculated by the equation l is the length between the inner contacts of the four-point set up, A is the cross-sectional area of the CNY calculated from the diameter measured from SEM images and R is the measured resistance.Characterization with a Micro-Raman Spectrometer  was performed over between 1000 and 1800 cmcontacts . SpecifiIn this work, we show a systematic study of the influence of annealing and functionalization by treatment with highly concentrated acids on the electrical conductivity and structure of CNY. Annealing enhances electrical conductivity by a factor of more than 5.5. A high graphitization of the CNT at 2500 \u00b0C leads to enhanced transport of electrons through the individual CNT. It was found the amorphous carbon resulting from the synthesis of the CNT plays an important role by connecting the CNT in the yarn structure and helps to reduce the contact resistance between adjacent CNT. Acid treatment over longer times and with different kinds of acids leads to an increase of electrical conductivity with a treatment time of 3 h resulting in the optimal increase. With this method, an increase of more than two times can be achieved. Furthermore, it was found that this increase is less dependent on forming functional groups on the surface of the CNT, but on the compression of the yarns and reduction of the intertube space. These results for annealing and functionalization help to understand the influences on CNT yarns by different post synthetic treatments and reveal key factors that could assist in producing highly conductive CNT yarns."}
+{"text": "The intensity of small-angle X-ray scattering from GaN nanowires on Si(111) depends on the orientation of the side facets with respect to the incident beam. This reminiscence of truncation rod scattering gives rise to a deviation from Porod\u2019s law. A roughness of just 3\u20134 atomic steps per micrometre-long side facet notably changes the intensity curves. I(q) \u221d q\u22124. The intensity depends on the orientation of the side facets with respect to the incident X-ray beam. It is maximum when the scattering vector is directed along a facet normal, reminiscent of surface truncation rod scattering. At large wavevectors q, the scattering intensity is reduced by surface roughness. A root-mean-square roughness of 0.9\u2005nm, which is the height of just 3\u20134 atomic steps per micrometre-long facet, already gives rise to a strong intensity reduction.Small-angle X-ray scattering from GaN nanowires grown on Si(111) is measured in the grazing-incidence geometry and modelled by means of a Monte Carlo simulation that takes into account the orientational distribution of the faceted nanowires and the roughness of their side facets. It is found that the scattering intensity at large wavevectors does not follow Porod\u2019s law  From the standpoint of small-angle scattering, GaN NWs are long hexagonal prisms with a substantial distribution of their cross-sectional sizes and orientations. Since these NWs are, on average, aligned along the substrate surface normal, the incident X-ray beam must be directed at a grazing incidence to the substrate surface. Grazing-incidence small-angle X-ray scattering (GISAXS) has been employed to study Si  from particles with sharp boundaries (i.e. possessing an abrupt change in the electron density at the surface) follows a universal asymptotic law I(q) \u221d q\u22124. Sinha et al.  has a dimensionality of q\u22121. Sinha et al.  as a model example. Since GaN NWs are faceted crystals  roughness of about 0.9\u2005nm, corresponding to the height of a few atomic steps on a micrometre-long NW sidewall facet. Both the facet truncation rod scattering and the surface roughness cause deviations from Porod\u2019s law.In this paper, we show that the GISAXS intensity from GaN NWs at large wavevectors depends on the azimuthal orientation of the NW ensemble with respect to the incident X-ray beam. The intensity is maximum when the scattering vector is directed along the facet normal, and minimum when the scattering vector is parallel to the facet. In other words, the azimuthal dependence of the GISAXS intensity reveals the facet truncation rods. We also show that the intensity at large 2.et al. (2016a2 plasma source for generating active N. The samples were grown on Si(111) substrates, which were preliminarily etched in dilute HF (5%), outgassed above 1173\u2005K for 30\u2005min to remove any residual SixOy from the surface, and exposed to the N plasma for 10\u2005min. The substrate growth temperature was approximately 1073\u2005K, as measured with an optical pyrometer. The Ga and N fluxes, calibrated by determining the thickness of GaN films grown under N- and Ga-rich conditions  and 1e). The NWs remain thin (and possess larger density) in their bottom parts and merge in their top parts. By merging, they reduce the surface energy of side facets at the cost of bending energy  of the GaN(0002) reflection to determine the tilt range with respect to the substrate surface normal, and the GaN using an X-ray energy of 22\u2005keV (wavelength \u03bb = 0.5636\u2005\u00c5). The incident beam was directed at grazing incidence to the substrate. The chosen grazing-incidence angle was 0.2\u00b0, qx. The scattering around the transmitted beam possesses a larger intensity. For that reason, the T streak is chosen here for further analysis. Besides the T and R streaks, the intensity distribution in Fig. 2Fig. 23.I(qx) from the measured maps. Since a single NW is a needle-like object, its scattering intensity in reciprocal space is concentrated in the plane perpendicular to the long axis of the NW. A random tilt of a NW results in a corresponding tilt of the intensity plane. Hence, one can expect that the spread of 4\u20135\u00b0 in the directions of the long axes of the NWs results in a sector of intensity in Fig. 2qz increasing proportional to qx.We use the specific features of the NWs as oriented long prisms to improve the accuracy of the determination of the GISAXS intensity i.e. scans at constant values of qx. These profiles are fitted by a Gaussian plus a background that may linearly depend on qz. The FWHMs of these profiles \u0394qz are plotted in Fig. 4a). As expected, \u0394qz increases linearly with qx. One can also see that the straight lines through the data points for each sample do not pass through the origin. The additional broadening can be attributed to the effect of the finite NW length L, so that, in the first approximation, the intersection point at qx = 0 is \u0394qz = 2\u03c0/L. For a more accurate determination of the NW lengths, we performed Monte Carlo simulations of the \u0394qz versus qx curves. The Monte Carlo simulations are described below in Section 4a) by green lines.Fig. 3qz/qx of the respective curves in Fig. 4a). These values are close to  the widths of the orientational distributions measured by Bragg diffraction, as described in Section 2The angular ranges of the NW orientations are 5.9\u00b0, 5.1\u00b0 and 4.6\u00b0 for samples 1, 2 and 3, respectively. They can be obtained from the slopes \u0394a). The average lengths for samples 2 and 3 are 350 and 400\u2005nm, respectively, notably smaller than the NW lengths in Figs. 1c) and 1e). The difference can be explained by NW bundling. The NW length measured in the GISAXS experiment is not its total length but an effective length of the NW segment between the merging joints. From the ratio of the total NW length to the segment length  at both small and large momenta qx. At small qx, the intensity profiles are narrow and the peak intensity has to be determined from just a few data points. At large qx, the intensity is low and the background is comparable to the signal. After performing the fits of the cross-sectional profiles  shown in Fig. 3qz on qx, we make one more step to reduce the noise of the data. Linear fits are made for the \u0394qz on qx dependencies plotted in Fig.\u00a04a). Then, the fits of the qz profiles shown in Fig. 3I(qx) are smoothed. These curves are used in the subsequent analysis.The fits in Fig. 3b) presents the GISAXS intensity I(qx) measured on sample 1 at varying azimuthal orientation \u03c8. The sample orientation \u03c8 = 0 corresponds to the incident X-ray beam along a GaNx-axis direction) is along b) comprises the measurements obtained on the rotation of sample 1 about the normal to the substrate surface (i.e. about the direction of the long axes of the NWs) from \u03c8 = 0\u00b0 to 30\u00b0 with a step of 5\u00b0. Since the sample has a rectangular shape and the illuminated area varies on rotation, the curves are scaled to obtain the same intensity in the small-qx range. The scaling factors differ by a factor of less than 2. The azimuthal dependence of the intensity at large qx is evident from the plot.Fig. 4b). Thus, we observe the same azimuthal dependence of the intensity but with a smaller total intensity and a higher level of noise. For this reason, for the further analysis presented here we exclusively consider the intensity distributions around the transmitted beam.In the case of the reflected beam  \u221d qx, we plot in Fig. 4c) the same data as qx, which would tend to a constant value for large qx. Surprisingly, a strong deviation from Porod\u2019s law is observed. Furthermore, the data not only deviate from Porod\u2019s law, but also exhibit a strong azimuthal dependence. In order to explain this un\u00adexpected behaviour, in the next section we develop a Monte Carlo method to calculate the scattering intensity.Since we expect Porod\u2019s law 4.4.1.A(q) in a coordinate system linked to the NW, i.e. with the z axis in the direction of the long axis of the NW. Hence, the cross section of the NW is in the xy plane. Next, we will consider in Section 4.3A(q)|2 over different NW orientations.We calculate first the scattering amplitude (form factor) of a NW V. Since the NW is a prism, the scattering amplitude can be represented as a product of the components along the NW axis and in the plane perpendicular to it, A(q) = A\u2225(q\u2225)A\u22a5(\u22a5q). The longitudinal component is simply L is the NW length.The scattering amplitude of a NW is given by its form factor A\u22a5(\u22a5q) can be reduced to a sum over the vertices, as was initially shown for faceted crystals by von Laue , rj are coordinates of the vertices, and lj and nj are unit vectors along the polygon side between the vertices rj and rj+1 and normal to it, respectively. Lee & Mittra /|rj+1 \u2212 rj|, we have in the limit q\u22a5\u2005\u00b7\u2005lj \u2192 0 For a planar polygon, the form factor readsttra 1983 proposedttke 2017 explicita) shows the intensity distribution calculated by equation (3b) shows a Monte Carlo calculation of the average intensity from hexagons of different sizes. A log-normal distribution of the lengths of the hexagon sides is taken with the same mean value of 12\u2005nm and a standard deviation of 4\u2005nm. This choice of parameters corresponds approximately to the respective values for sample 1.Fig. 6f) by the black line. The intensity distribution is presented as the product qx for an ensemble of randomly oriented hexagons  after averaging over all possible orientations. As stated above, the intensity maxima in Fig. 6b) correspond to the directions normal to the sides of the hexagon. They possess, at large qx, an f), we observe a linear increase in the intensity for large values of qx (black line).The radial intensity distribution in the direction along the intensity maximum is presented in Fig. 6qm = 0.175\u2005nm\u22121 in Fig. 6f) is related to the mean length of the side facet of the hexagons a = 12\u2005nm as a \u2243 2.1/qm, which allows us to determine the hexagon size directly from plots of qx  is the Bessel function. The first maximum of J1(x) at x \u2243 1.84 gives the circle radius R \u2243 1.84/qm. We can relate a hexagon and a circle even closer, by defining an effective radius Ra of a circle possessing the same area as a hexagon with a side length a. Then, we have Ra = a and Ra \u2243 1.9/qm, with the proportionality coefficient very close to the case of a circle.The local maximum at a): random heights hj are taken in the directions normal to the facets. Then, we check that the generated hexagon is convex, and discard it otherwise. Fig. 5b) presents examples of randomly generated hexagons with the same orientation of their sides. The distribution of the hexagon shapes is chosen to simulate sample 1 and is further described in Section 5c). The respective radial intensity distribution is presented in Fig. 6f) as the blue line. One can see that the black and blue lines in Fig. 6f) are remarkably different. In particular, the hexagon shape distribution notably reduces the dip in intensity. Therefore, the distortion of the hexagons can be deduced from the intensity plots.With the form factor defined by the positions of the vertices according to either equation , where the function R(q) is defined as pm are the probabilities of the shift of the side facet by m steps. Hence, the function R(q) is the characteristic function of the probabilities pm.A step of height pm = (1 \u2212 \u03b2)\u03b2m with the parameter \u03b2 < 1. It describes a flat surface with a fraction \u03b2 one step higher, its fraction \u03b2 is, in turn, one step higher, and so on /(1 \u2212 \u03b2) and the corresponding characteristic function is Consider the geometric probability distribution d0(\u03bc1/2) and the characteristic function is The Poisson probability distribution jth term in sum (3Rj = R(q\u22a5\u2005\u00b7\u2005nj) that depends on the orientation of the respective facet. This is different from a common treatment of surface roughness, which involves a single factor |R|2. In particular, the Poisson probability distribution gives for R|2 = et al.  shows the scattering intensity distribution obtained with the roughness factors given by equation  corresponds to prisms with perfectly aligned long axes.Fig. 6f) shows the radial intensity distribution obtained from the map shown in Fig. 6d) in the direction of maximum intensity, calculated using the roughness factors for the geometric probability distribution given by equation (7f) shows the intensity from the same distribution of hexagons but calculated using the roughness factors derived from the Poisson probability distribution, equation  presents a Monte Carlo calculation of the intensity for the distribution of distorted hexagons described above and sketched in Fig. 5b), after averaging over the orientations \u03c8 uniformly distributed from 0\u00b0 to 360\u00b0. The corresponding radial intensity distribution, shown in Fig. 6f) by a grey line, follows the two-dimensional Porod law I(q) \u221d q\u22123 at large q. At small q, it coincides with the intensity distribution for the oriented hexagons.Fig. 6q\u2032 for a given NW by applying three rotations about the orthogonal axes x, y and z to the scattering vector q in the laboratory frame, q\u2032 = Mz(\u03c8)My(\u03d5)Mx(\u03b8)q. Here the scattering vector in the laboratory frame is shown in Fig. 2qx axis is normal to the mean direction of the long axes of the NWs, the qz axis is along this direction and qy = 0. The matrices Mj  are the rotation matrices about the respective axes, and the angles \u03b8, \u03d5 and \u03c8 defining the NW orientation are the nautical angles, as shown in Fig. 7For the three-dimensional distribution of the NW orientations, inherent to the spontaneous formation of GaN NWs on Si(111)  \u221d q\u22124. However, since the NWs are long prisms, the scattering intensity from a single NW of length L with its long axis in the z direction concentrates in reciprocal space in a disc of width \u0394qz = 2\u03c0/L. We have seen in Section 3qz/qx < \u0394\u03b8, where \u0394\u03b8 is the angular range of orientations. As long as 2\u03c0/(Lqx) < \u0394\u03b8, oriented NWs give the same intensity in the x direction as fully randomly oriented ones. Therefore, Porod\u2019s law is satisfied for qx > 2\u03c0/(L\u0394\u03b8).If the NW orientations are completely random, L = 200\u2005nm, the condition derived above reads qx > 0.7\u2005nm\u22121, which is in good agreement with the region of constant Fig. 85.qx by a Gaussian, as shown in Fig. 3qz scans obtained in this fit provided the intensity I(qx).Figs. 9The measurements performed at different azimuthal orientations \u03c8 of the sample are presented in Fig. 9qx, to reveal deviations from Porod\u2019s law. The data are represented in more detail: one can see that the sixfold symmetry is quantitatively proven for samples 2 and 3, while sample 1 reveals some difference between the curves at \u03c8 = 0\u00b0 and 60\u00b0. This difference is presumably due to the illumination of different parts of the sample during its rotation.The same data are presented in Fig. 10We calculate the scattering intensity by the Monte Carlo method. It enables a simultaneous integration over the distributions of the NW lengths, their cross-sectional sizes and shapes, and the orientations of both the NW long axis and of the side facets. The calculations take a fairly short time. It takes less than a minute on a single CPU core of a standard PC to calculate an intensity curve to an accuracy sufficient to make estimates. The smooth curves presented in this paper took less than an hour of CPU time each.qz scans, similar to the ones presented in Fig. 3qz as a function of qx. The results of these calculations are presented in Fig. 4a) as green lines. For sample 1, agreement between the measurements and the simulations is found at a mean NW length of 230\u2005nm, coinciding with the mean NW lengths in the scanning electron micrograph in Fig. 1a). For samples 2 and 3, the simulations of the \u0394qz versus qx curves give mean NW lengths of 350 and 400\u2005nm, respectively, values notably smaller than the real NW lengths in Figs. 1c) and 1e). This difference is explained, as we have already discussed in Section 3Table 1Taking into account the sixfold symmetry of the intensity distributions in Fig. 9a, its variation and the variation in the cross-sectional shapes of the NWs, and the roughness of the side facets. We have seen in Section 3a determines the position of the local maximum of the curves at qx \u2243 0.17\u2005nm\u22121, which corresponds to a side facet width of about 12\u2005nm. The depth of the dip between this maximum and the rise of the curves at larger qx is controlled by the width of the facet size distribution and the shape distribution of the cross sections. The decrease in qx is caused by the roughness of the side facets.The parameters of the NW ensemble to be determined from the Monte Carlo simulations are the mean width of the side facets hj shown in Fig. 5a) were generated at random around a mean value. Fig. 5b) exemplifies the shapes of the NWs used in the simulation of sample 1. The right-most column in Fig. 10The distorted cross sections of the NWs were modelled in the Monte Carlo study as described in Section 4.1A and the perimeter P. Then, we determine out of these parameters the radius R from A = \u03c0R2 and the circularity C = 4\u03c0A/P2. The circularity thus defined is C = 1 for a circle, C = et al., 2014For each generated NW, we calculate the cross-sectional area a), we estimate the accuracy of the determination of the tilt angle distribution to be \u00b1\u20050.1\u00b0, while the accuracy of the NW length distribution can be estimated as \u00b1\u200550\u2005nm.The mean NW radii, the standard deviations of the radius distributions and the roughnesses obtained in the Monte Carlo simulations for samples 1\u20133 are presented in Table 1d)\u20131f). The analysis was performed using the open-source software ImageJ  is fairly close to the distribution derived from the micrographs. The circularity distribution obtained from the micrographs is, however, extended towards smaller values, indicating a higher density of NWs with elongated cross-sectional shapes. Such a discrepancy can be attributed to an artefact caused by the NW tilt. Specifically, the scanning electron micrographs exhibit very little difference in brightness between the top facet and the top part of the side facet of the NW, so that ImageJ treats both regions together, i.e. as extended intensity spots.The distributions of the cross-sectional radii and circularities of the NW ensembles were also interdependently obtained by analysing top-view scanning electron micrographs similar to those shown in Figs. 1l. 2016a in theirc) and 11e)] are smaller than those derived from analysis of the scanning electron micrographs, and the discrepancy increases with increasing NW length. We remind the reader that the mean NW radius can be derived directly from the position qx of the local maximum in the experimental curves presented in Fig. 10qx \u2243 0.17\u2005nm\u22121 and shifts only slightly to smaller values (and hence to larger radii) as the NW length increases from sample 1 to sample 3.In contrast with sample 1, the NW radii obtained from the Monte Carlo simulations of the scattering intensity from samples 2 and 3 [Figs. 11et al., 2016aThe origin of the discrepancy between the NW radii determined from the scanning electron micrographs and from the modelling of the GISAXS intensity is in the bundling of the NWs. Bundling is almost absent for sample 1, and the cross sections of the NWs obtained from the micrographs characterize the NWs along their full length. As the NWs grow in length, they bundle together, which causes an apparent radial growth. Simultaneously, the NW density decreases, so that the fraction of the surface covered by the NWs remains constant  and 11f) are due to the finite resolution of the micrographs, as well as to the algorithm used by ImageJ that tends to round faceted objects.The widths of the circularity distributions in the right-hand column of Fig. 11f) that, when the scattering vector is oriented normal to the side facets of the NWs (\u03c8 = 0\u00b0) and the facets are atomically flat, facet truncation rod scattering would result in a linear increase in the intensity on the qx plot at large qx. The decrease in the experimental curves indicates a roughness of the side facets. We obtain in the Monte Carlo modelling r.m.s. roughnesses of \u03c3 = 0.9, 0.95 and 0.85\u2005nm for samples 1, 2 and 3, respectively. According to the height of the atomic steps on a GaN(d0 = a0a0 = 0.319\u2005nm is the GaN lattice spacing), the r.m.s. roughness is less than 3.5\u2005steps.We have seen in Section 4.16.GaN NWs nucleate spontaneously on Si(111) and grow with a substantial disorder with respect to their orientations. Their growth is, nevertheless, epitaxial: the NWs inherit the out-of-plane and in-plane orientations of the substrate. Since these NWs are typically long (from hundreds of nanometres to a few micrometres) and thin (tens of nanometres), the range of orientations of their long axes of 3\u20135\u00b0 is sufficient to provide the same average in the small-angle scattering intensity as if they would have all orientations in space. However, an angular range of orientations of the side facets of 3\u00b0 gives rise to features in the GISAXS intensity distribution that are reminiscent of the crystal truncation rod scattering from flat surfaces of single crystals.We have found that the GISAXS intensity depends on the orientation of the side facets with respect to the incident X-ray beam direction. In our experiment, the incident beam is kept normal to the average direction of the long axes of the NWs. The orientation of the incident beam with respect to the side facets is varied by rotating the sample about the substrate surface normal. The scattering intensity is maximum when the incident beam is along the facets, or in other words, when the scattering vector is in the direction of the facet normal.q\u22122. Porod\u2019s law (q\u22124) is a result of a full average over all orientations of the plane , the range of orientations of the long axes is large enough to provide an integration over the tilt angle and give rise to a q\u22123 dependence when the scattering vector is along the facet normal. In the Iq4 versus q plots shown in Fig. 10i.e. in the direction between facets.The X-ray scattering intensity from a planar surface is proportional to q\u03c3 \u2273 1, where \u03c3 is the r.m.s. roughness. The Monte Carlo modelling of the experimental curves in Fig. 10q.The surface roughness gives rise to a decrease in the intensity at et al., 2016aThe GISAXS curves vary fairly little from one sample to another, despite the large difference between the cross-sectional sizes of the NWs observed in the scanning electron micrographs shown in Fig. 1Finally, we conclude that GISAXS, together with Monte Carlo modelling of the intensity curves, is well suited for the determination of the distributions of the cross-sectional sizes of NWs. The methods developed in the present paper are not specific to GaN NWs on Si(111) and can be applied to other NW distributions and material systems. In particular, they will be applied in a separate study to assess the radius distributions of GaN NW ensembles grown on TiN, which exhibit a much lower density that hinders analysis of the NW cross-sectional shapes by scanning electron microscopy."}
+{"text": "We assessed the clinical outcomes and toxicities following hypofractionation with helical tomographic intensity-modulated radiotherapy technology (tomotherapy) in patients with stage III non-small cell lung cancer (NSCLC) who were not candidates for surgery or concurrent chemoradiation.  Forty-three patients with stage III NSCLC who were treated between 2011 and 2017 were enrolled. The prescription doses for gross target volume and clinical target volume were 70\u2009Gy and 60\u2009Gy (respectively) delivered in 15\u201325 fractions over 3\u20135 weeks. p < 0.05). Seven patients developed grade 1-2 acute radiation pneumonia (RP), 5 patients developed grade 1-2 late RP, while 3 patients developed grade 3 late RP. None of the patients developed grade 4-5 radiation lung injury.  The median overall survival (OS) time was 34.23 (range 11.33\u201399.33) months. The estimated 1-, 2-, and 3-year OS rates were 97.7%, 74.4%, and 55.9%, respectively; the corresponding progression-free survival (PFS) rates were 79.1%, 53.5%, and 36.1%, respectively. The local disease recurrence, regional disease recurrence, and distant metastasis rates at 3 years were 4.7%, 11.62%, and 55.81%, respectively. On multivariate analysis, dose regimen (<19\u2009f vs. \u226519\u2009f) was an independent prognostic factor affecting OS, PFS, and DM ( Tomotherapy may be an effective treatment option for patients with stage III NSCLC. It may be a viable alternative to surgery with lower incidence of side effects. Currently, stereotactic radiotherapy is becoming a viable alternative to surgical treatment of early-stage non-small cell lung cancer (NSCLC). The clinical application of molecular targeted therapies and immunotherapy have significantly prolonged the survival time of patients with advanced-stage NSCLC. For most patients with locally advanced NSCLC, the current standard is chemoradiotherapy , which has not changed for more than 40 years ; the assIn a previous study conducted at our department, the 3-year survival rate of patients with stage I NSCLC was 91% . Chang eTherefore, the purpose of this study was to explore the feasibility and effectiveness of hypofractional tomotherapy in patients with stage III NSCLC who are not eligible for surgery or concurrent chemoradiation. The primary end points were overall survival (OS) rate and progression-free survival (PFS) rate. Local control rate, regional control rate, distant metastasis rate (DM), and incidence of radiation pneumonitis (RP) were regarded as secondary end points.A total of 43 patients with stage III NSCLC who were treated between December 2011 and March 2017 were enrolled in this study. The inclusion criteria were as follows: (1) patients with histologically or cytologically confirmed NSCLC who were inoperable or refused surgery; (2) Karnofsky performance score (KPS) >70; (3) patients who were able to lay on the bed for more than 30 minutes; (4) use of 18-Fludeoxyglucose positron emission tomography (18F-FDG PET)/CT and CT scans for stage III patients within 1 month before tomotherapy; (5) union for International Cancer Control (UICC) TNM classification system (2002) was used for staging; (6) patients who had previously received chemotherapy, except bleomycin, were also allowed in this study. Genetic testing was recommended for patients with adenocarcinoma and targeted therapy (such as gefitinib or erlotinib) was recommended as maintenance therapy in patients with EGFR-mutant or EML4-ALK fusion gene. Patients who received other treatment for NSCLC, those with history of any cancer in the 5-year period immediately preceding the confirmation of NSCLC, those with a history of chest radiotherapy, or those with any serious medical conditions were excluded from this study.All patients were instructed to breathe normally without control. Using a body net to fix the supine patients, an enhanced CT scan was used for location. Scanning range was from the mandible to 3\u2009cm below the diaphragm, including the entire lung tissue. Slice thickness and slice gap were all 5\u2009mm. Scanning image was sent directly onto the HT Hi-Art version 5.1.4 Accuray planning system  through the network with 4\u2009s scanning speed for each level. Monaco version 5.11.01 software  was used to draw the target region and organs at risk (OARs). The pulmonary tumors were performed in the lung windows  and the mediastinal lymph nodes were delineated in the mediastinal windows . The gross target volume (GTV) was defined as the primary tumor and lymph nodes greater than 1.5\u2009cm in the short axis in the enhanced CT or PET/CT; the clinical target volume (CTV) included extra 5\u2009mm around the GTV; the planning target volume (PTV) included an extra 5\u2009mm margin around the CTV. Image guidance was performed every time before treatment. We did not consider the impact of respiratory movements on inside target volume (ITV). Respiratory gating system was not used in the study. The prescription dose for GTV and CTV were 70\u2009Gy and 60\u2009Gy administered in 15\u201325 fractions over a period of 3\u20135 weeks. Patients with older age, severe multiple comorbidities, higher level of lymph node metastases, and large tumor volumes received more fractions treatment in order to reduce higher dose volume in the normal lung tissues and OARs.The relative volume of the main bronchi, esophagus, and trachea receiving more than 55\u2009Gy was required not to exceed 30%; the maximum dose to the spinal cord was not more than 45\u2009Gy; the acceptable dose of the V20 for total lungs was \u226430%; the relative volume of heart receiving more than 50\u2009Gy did not exceed 50%.rd, 6th, 9th, and 12th in the first year, and every 6 months for the next two years. PET/CT was also recommended in all patients between 3\u20136 months in the first year or if tumor relapse was suspected. Local disease recurrence (LR) was defined as CT evidence of tumor progression in the same lobe or if the PET/CT images showed SUVmax >5. Regional disease recurrence (RR) was defined as occurrence of any intrapulmonary lymph node metastasis. Distant metastasis (DM) was defined as occurrence of any metastasis outside the lungs or any tumor seeding in a different lung lobe. The first date as the time of tumor progression was the time when the PET/CT or CT image demonstrated abnormality. More details are available in our previously published article [Patients were assessed by physical examination and chest CT scan. The first follow-up was at 4\u20136 weeks after treatment; subsequent follow-ups were at 3 article . Acute rp values <0.2 in univariate analyses were included in the multivariate analyses. All significance tests were 2-tailed and p values <0.05 were considered indicative of statistical significance.OS was measured from the first day of tomotherapy to the day of death or last follow-up. PFS, LR, RR, and DM were calculated from the first day of tomotherapy to the first day of the tumor progression or death. All statistical analyses were performed using the SPSS statistical software version 20.0  or Graphpad Prism 8.3 software . Pearson chi-square test and Fisher's exact test were used to compare the characteristics of patients. The Kaplan-Meier method was used to generate the OS and PFS curves; the optimal cutoff values of continuous variables were obtained by receiver operating characteristic (ROC) curve analysis. Univariate analyses and multivariate analyses were performed using the Cox proportional hazard model. Factors considered to influence the prognosis (such as history of chemotherapy and targeted drug therapy) or those associated with The median age of patients was 69 years (range 50\u201386). Twenty-one patients had confirmed squamous cell carcinoma, 12 patients had confirmed adenocarcinoma, and the rest had other types. In 22 patients, the tumor was located in the central part, while 21 patients had peripheral tumors. Fifteen patients had undergone chemotherapy before tomotherapy, while 28 patients had not undergone chemotherapy .The median OS time was 34.23  months, while the median PFS time was 25.00  months. The estimated 1-, 2-, and 3-year OS rates were 97.7%, 74.4%, and 55.9%, respectively, while the 1-, 2-, and 3-year PFS rates were 79.1%, 53.5%, and 36.1%, respectively . Accordip < 0.05), while the dose regimen was the only factor associated with poorer PFS and DM (p < 0.05). Chemotherapy and targeted therapy were associated with poorer RR (p > 0.05) in univariate analysis; however, no significant association was observed in multivariate analysis. None of the factors showed an association with LR in univariate or multivariate analyses.p < 0.05, data not shown).In this study, hypofractionation with tomotherapy prolonged the overall survival time of patients with stage III NSCLC compared with traditional treatment. Moreover, this treatment reduced the occurrence of various complications during the perioperative period, thereby reducing the risk of death. Therefore, our findings suggest that hypofractionation with tomotherapy may be an effective alternative to surgery.Currently, concurrent chemoradiotherapy is the standard treatment for patients with stage III NSCLC who cannot be operated . However\u03b3-SBRT technique with the mode of GTV/CTV/70\u2009Gy/60\u2009Gy/10\u2009f (BED: 119\u2009Gy) for patients with stage I/II NSCLC; the 3-year LC rate was 95% and the 5-year OS rate was 60.3% [The modern radiotherapy technology has made it possible to deliver high-dose radiation to the target tissue while ensuring low dose to the surrounding normal tissues; this has made the SBRT as the standard treatment for patients with early-stage NSCLC \u20138. In a as 60.3% . The prop = 0.322) and regional control rates . Only two patients developed local recurrence at 5.63 months and 7.76 months after treatment, while five patients developed regional recurrence within 3 years. We further found that the therapeutic effect in the <19\u2009f group was better with higher BED (102.69\u2009Gy) as compared to that in the \u226519\u2009f group. Moreover, the dose regimen was the only independent prognostic factor for OS, PFS, and DM. This phenomenon is likely attributable to the activation of immunogenic system and the occurrence of \u201cabscopal effect\u201d after hypofractional radiation [The main cause of tumor progression is either poor local control or distant metastasis. Both study groups (<19f vs. \u226519f) showed excellent local control rates . The incidence of late RP is similar to that reported by Yan et al. . Shen etSome limitations of our study should be improved. In several studies, tumor volume, or even volume changes during treatment were associated with the overall survival of patients \u201322. OwinOur findings suggest that hypofractionation with tomotherapy is an effective treatment option for patients with stage III NSCLC who were medically inoperable or refuse concurrent chemotherapy; these patients may also benefit from the reduced incidence of toxicity."}
+{"text": "In spite of the efforts in developing and maintaining accurate variant databases, a large number of disease-associated variants are still hidden in the biomedical literature. Curation of the biomedical literature in an effort to extract this information is a challenging task due to: (i) the complexity of natural language processing, (ii) inconsistent use of standard recommendations for variant description, and (iii) the lack of clarity and consistency in describing the variant-genotype-phenotype associations in the biomedical literature. In this article, we employ text mining and word cloud analysis techniques to address these challenges. The proposed framework extracts the variant-gene-disease associations from the full-length biomedical literature and designs an evidence-based variant-driven gene panel for a given condition. We validate the identified genes by showing their diagnostic abilities to predict the patients\u2019 clinical outcome on several independent validation cohorts. As representative examples, we present our results for acute myeloid leukemia (AML), breast cancer and prostate cancer. We compare these panels with other variant-driven gene panels obtained from Clinvar, Mastermind and others from literature, as well as with a panel identified with a classical differentially expressed genes (DEGs) approach. The results show that the panels obtained by the proposed framework yield better results than the other gene panels currently available in the literature. There are several publicly available databases contain the disease-associated variants such as Clinvar2, SNPedia3, OMIM4, Swiss-Prot5, COSMIC6, BioMuta7, HGMD8, UMD9, HGVbaseG2P10, MutDB11, dbSNP12, PharmGKB13 and InSiGHT14. All these databases are manually curated by human experts. While this manual curation ensures a high quality of the annotations, the manual extraction of this type of information from the biomedical literature takes an enormous amount of time and effort. The current rate with which new variants are published is simply too high for any manual annotation process. As an additional challenge, despite the HGVS (Human Genome Variation Society) standard recommendations for the description of the variants, many variants are still reported in literature in non-standard formats. A number of automatic mutation indexing tools have been developed. Such tools process biomedical literature and produce a list of mutations that appear in these papers. These include MutationFinder15, EMU16, MEMA17, MuteXt18, Mutation GraB19 and MutationMiner20. The most recent such tool, tmVar 2.021 extracts variants from an article and normalizes them to their unique dbSNP identifiers. The next step is to develop software tools to extract variants-disease associations from the biomedical literature. Several methods have been proposed for this purpose such as MuGeX22, OSIRIS23, EnzyMiner24 and the methods proposed by Singhal\u00a0et al.25. All these methods have been applied to only the title and the abstract section of biomedical articles. However, a comprehensive study showed that a significant number of genetic variants are only included in the full text and the supplementary materials of the articles26. These will be missed if the variants are only extracted from titles and abstracts. Doughty\u00a0et al.16 also proposed a tool named EMU for extracting the disease-associated mutations from biomedical literature. Although this tool automatically extracts the mutations and their corresponding genes from an article, it still requires human curation to discover the mutation-disease associations.One crucial step in understanding the biological mechanism underlying a disease condition is to capture the relationship between the variants and the disease riskHere we propose an automated framework to extract disease-associated variants from the full-length biomedical literature and design a variant-driven gene panel for a given disease phenotype. As the first step, the proposed framework employs word cloud analysis to identify the variant-relevant articles. The variant-gene-disease associations are then extracted from these articles. An evidence-based variant-driven gene panel is then generated based on the mined triplet information. A comprehensive validation procedure illustrates the capabilities of the proposed framework. We validate the proposed variant-driven gene panels by showing their abilities to predict the patients\u2019 clinical conditions  on multiple independent validation datasets.Figure\u00a021. One solution to address this challenge is to mine only the variant-relevant articles. We compare the performances of two different approaches for detecting the variant-relevant articles. The first approach considers only the articles that mention any disease or gene or any of their synonyms in the title and abstract sections. In the second approach, we employ the word cloud analysis and generate a weighted list of variant-relevant keywords. In particular, we first generate a weighted list of words (referred to as variant-relevant keywords) that appear frequently in the full-body text of 10,000 random articles with at least one mentioned variant (using tmVar 2.0). Subsequently, an article is considered to be relevant to variants if at least 10% of these keywords appear in the full-body of the article. We apply both approaches on a set of 10,000 random full-length articles. Figure\u00a0The input of the proposed framework consists of 3,322,746 full-length articles downloaded from the PMC database on January 2020. The variant indexing procedure from a full-length article is challenging because any chemical formulae, figure numbers, etc. that are represented in \u201cCharacter-Number-Character\u201d format could be identified as a variant27 to identify the appropriate genes. The tmVar 2.021 is the tool we employ for extracting the variants and normalizing those which are included in dbSNP to their unique identifiers (dbSNP RSIDs). We use DNorm28 to identify all the disease phenotypes mentioned in an article.We use the publicly available and well-known entity recognition tools to extract the variant, gene and disease phenotype from each input article. In particular, we use GNormPlus21 to find the associated gene. Then, we map each retrieved variant-gene pair to the corresponding genomic coordinates  using the Variant Recoder30 tool. We eliminate the variant-gene pairs\u00a0with no matched genomic coordinates .Once a variant is extracted from an input article, we follow the steps provided by Wei\u00a0et al.1. Let V and the closest (based on the word counts) mentioned diseases in an article, where k is the number of times this variant is mentioned in that article. The disease association score is calculated for each appearance of variant\u00a0V and the closest mentioned disease DV and the disease\u00a0The Same Sentence Occurrence (SSO) is a binary score which is 1 when the variant\u00a0V and the disease\u00a0The Same Paragraph Occurrence (SPO) is a binary score which is 1 when the variant\u00a0V and the disease\u00a031 for this analysis.The sentiment score (SS) calculates the polarity sentiment value for the text mentioned between the variant\u00a0The variant V is considered to be associated with disease\u00a0We use a set of features to capture the variant-disease associations from an input article adapted from the method proposed by Singhal et al.16. These datasets contains variant-disease pairs extracted from 29 and 129 PubMed articles for prostate cancer and breast cancer, respectively. We use these datasets and report the standard evaluation metrics  for both methods. As shown in Table\u00a0We also perform an experiment to compare the performance of the proposed scoring method for extracting the variant-disease associations with the simple sentence co-occurrence scoring method (baseline method). In this experiment, we use the\u00a0two manually curated benchmark datasets provided by Doughty\u00a0et al.In this step, we first generate a variant-gene-disease panel which includes all the associations between the gene, variant and disease entities extracted from the input corpus  of the receiver-operator characteristic (ROC) to assess the performance of the classifier. We repeat this procedure n times (where n is the number of available gene expression datasets). An average of the AUCs is calculated over the n rounds of sampling. This procedure is used to compare the diagnostic quality of the proposed gene panel with the current available variant-relevant gene panels\u00a0obtained from\u00a0literature.In this section we describe the\u00a0two experiments performed to assess the diagnostic value of the proposed variant-driven gene panels. In the first experiment, we use the genes present in the proposed panel to predict the patients\u2019 clinical condition  from several independent patient cohorts Fig.\u00a0. The hyp33. The detailed descriptions of the enrichment pathway analysis method are included in the Supplementary Materials. A \u201ctarget pathway\u201d refers to the pathway that was created to explain the mechanism of the given disease (e.g. the acute myeloid leukemia KEGG pathway (hsa05221) is the target pathway for acute myeloid leukemia). The expectation here is that a gene panel that is relevant to the given disease would rank the target pathway at the very top of the ranked list of pathways. This validation method was widely adopted by others, such as42. In addition, not only the target pathway but also the other identified significantly enriched pathways provide crucial information to assess the performance of the proposed gene panel. We\u00a0also provide the top 10 significantly enriched pathways and the references explaining the association of the respective pathways to the disease case study for each gene panel in the Supplementary Materials.In the second experiment, we assess the relevance of the proposed gene panel to the given disease based on the rank of target pathway when an enrichment pathway analysis is performed. For each signaling pathway, the enrichment pathway analysis method calculates the probability of finding a center number of gene overlaps between the proposed gene panel and the presented genes in each pathway just by chance and then ranks the pathways based on this probability43. Dataset summaries are described in the Supplementary Materials. For each disease case study, we also calculate the percentage of the genes in the proposed gene panel that overlap with the genes in each gene expression dataset. We perform this experiment as a quality check to ensure that the majority of the genes in the proposed gene panel are contributing to the validation analysis , breast cancer and prostate cancer. The resulted gene panel proposed for each case study are included in the Supplementary Materials Table . All the44 demonstrated that the AML patients with TP53 alterations have lower response\u00a0rate to the intensive chemotherapy and therefore have inferior survival rate. FLT3 and C-KIT are known to be associated with poor AML prognosis discovered by Pratz et al.45 and Yang et al.46, respectively. Ley et al.47 investigated the role of DNMT3A and found that there is a direct link between the presence of mutations in this gene and the intermediate risk of AML. Chaturvedi et al.48 also reported the therapeutic role of mutant IDH1 in AML. Gaidzik et al.49 have shown that the therapy-resistance and inferior outcomes are the main genetic characteristics of the AML patients with RUNX1 mutations. The presence of mutations in TYMS and NPM1 are also discovered in AML patients51. SLC29A1 mutations are also\u00a0found to be associated with poor therapy outcome in AML patients52.First, we extract all the genes with at least one mentioned variant discovered to be associated with AML by the proposed framework. The top 10 genes that have the highest number of variants are TP53, FLT3, KIT, DNMT3A, IDH1, COX8A, RUNX1, TYMS, NPM1 and SLC29A1. These genes play significant roles in the underlying mechanism of AML. For instance, Kadia et al.43. The other variant-driven gene panels which are available for AML are obtained from Clinvar2, Mastermind29 and the panel proposed by Singhal et al.25. Clinvar is a repository for mutations and their associated disease phenotypes which are manually curated from the biomedical literature. The Mastermind search engine provides literature-based variant-genotype-phenotype association information. We also include the results when using only the differentially expressed genes  plays important role in prostate cancer cell proliferation as demonstrated by Balk\u00a0et al.25 in terms of the ability to predict the patients\u2019 clinical outcome on several independent validation cohorts  Singhal et al.25, iii) Doughty et al.16 and iv) the classical DEGs. The classification results demonstrate that the gene panel proposed here performs better than the other gene panels in terms of the ability to predict the patients\u2019 clinical outcome on several independent validation datasets  Clinvar25, could be the reason for the gene differences between these two panels. We also investigate the percentage of the identified AML-related variants which are mentioned in the title and abstract sections of the articles, and compared them with those that are mentioned in the full body of the articles but not in the title and the abstract. Figure\u00a0We investigate the novelty of our identified genes by checking their overlap with other available variant-driven gene panels for AML Fig. . AlthougThe number of articles describing the disease-related variants is rapidly increasing. This highlights the pressing need for the development of automated tools that are able to extract the variant-disease associations from literature. In this article, we implement an automated framework to extract the variant-gene-disease associations from the full-length biomedical literature and design an evidence-based variant-driven gene panel for a given disease. The identification of the variant-relevant articles using word cloud analysis, and the consideration of the full-length articles are the main contributions of the proposed framework. We illustrate the utilities of the proposed variant-driven gene panels in capturing the mechanisms involved in AML, prostate cancer, and breast cancer using 27 independent gene expression datasets containing a total 2,109 patients. The results show that the proposed gene panel outperforms the other published gene panels in terms of the ability to predict the patients\u2019 clinical outcome.Supplementary informationSupplementary Table S1Supplementary Table S2Supplementary Table S3"}
+{"text": "Goserelin is an effective alternative to surgery or estrogen therapy in prostate cancer palliation, and possibly to ovariectomy in premenopausal breast cancer. However, not all users of goserelin can benefit from it, or some patients are not sensitive to goserelin. The advent of network pharmacology has highlighted the need for accurate treatment and predictive biomarkers. In this study, we successfully to identify 76 potential targets related to the compound of goserelin through network pharmacology approach. We also identified 18 DEGs in breast cancer tissues and 5 DEGs in cells, and 6 DEGs in prostate cancer tissues and 9 DEGs in cells. CRABP2 is the common DEG both in breast and prostate cancer. The risk prediction models constructed with potential prognostic targets of goserelin can successfully predict the prognosis in breast and prostate cancer, especially for very young breast cancer patients. Moreover, seven subgroups in breast cancer and six subgroups in prostate cancer were respectively identified based on consensus clustering using potential prognostic targets of goserelin that significantly influenced survival. The expression of representative genes including CORO1A and ANXA5 in breast and DPP4 in prostate showed strong correlations with clinic-pathological factors. Taken together, the novel signature can facilitate identification of new biomarkers which sensitive to goserelin, increase the using accuracy of goserelin and clarify the classification of disease molecular subtypes in breast and prostate cancer. Goserelin is a synthetic analogue of gonadotrophin-releasing hormone (GnRH) which stimulates gonadotrophin and sex hormone release in the short term, and then causes suppression with continued administration , 2. BreaRecently, network pharmacology (NP) was proposed as a promising approach to discover drugs from a systems perspective and at the molecular level. It combines bioavailability prediction, multiple drug target prediction and network analysis to understand the active compounds and therapeutic targets of drug , 7. Lee Currently, clinicians mainly use the clinic-pathological factors and clinical stages to assess the risk of breast and prostate cancer in patients and guide clinical treatment , 10. Thehttps://cancergenome.nih.gov/). It contains 1164 breast samples  and 551 prostate samples . The clinical data of above samples, such as gender, age, tumor grade, clinical stage, and survival time, were also downloaded from the TCGA database. The RNA expression data of cells with breast and prostate cancer were downloaded from the Gene Expression Omnibus . Data from the GSE 62410, GSE 107209 datasets were used for differential expression analysis. GSE 62410  contains 6 samples, including 3 PrEC cells and 3 LNcaP cells. GSE 107209  contains 6 samples, including 3 MCF-10A cells and 3 UFH-001 cells. The gene expression data and clinical data of very young breast cancer patients were derived from international cancer genome consortium (ICGC) portal (https://dcc.icgc.org/). Genotype-Tissue Expression (GTEx) program (https://commonfund.nih.gov/gtex) established a data resource and tissue bank of healthy people. Data of GTEx were used to plot the expression of genes in multiple normal tissues in our study, and we used the R language \u201cggpubr\u201d package to drawing boxplot to distinguish the expression of genes between male and female. PharmMapper Server is designed to identify potential target candidates for the given probe small molecules  using pharmacophore mapping approach [https://pubchem.ncbi.nlm.nih.gov/), and finding the best interaction mode between the potential target candidates and compound by pharmacophore database (http://www.lilab-ecust.cn/pharmmapper/). R software (3.6.2) was used for data extraction and sorting to obtain the gene expression matrices and clinical data.The RNA-Seq gene expression profiles (FPKM format) of patients with breast and prostate cancer were downloaded from the cancer genome atlas (TCGA) portal  between tumor and normal tissues/cells, we used the R language \u201climma\u201d package to screen the DEGs. Mann Whitney test was performed to determine differential expression levels of genes between tumor samples and corresponding control samples. |log2 fold change (FC)|>\u20091, and false discovery rate (FDR) values\u2009<\u20090.05 were considered to be statistically significant.https://stringdb.org/) [http://www.cytoscape.org/) [A protein protein interaction (PPI) network of related goserelin target genes were constructed using the STRING online database (db.org/) , and thepe.org/) . Gene onIn order to build a prognostic model applicable to breast and prostate cancer patients and relate to target genes of goserelin, all target genes were used to conduct a univariate Cox survival regression, and P value\u2009<\u20090.05 was used to create a lasso regression model. This method was applied to reduce potential over-fit and implemented through the \u201cglmnet\u201d R package. Univariate and multivariate Cox analysis were conducted with R language \u201cforest\u201d and \u201csurvival\u201d package. The risk scores of each patient were calculated through the prediction formula of the risk prediction model. We calculated the cut-off value used to determine whether the patient is at high or low risk. The formula used for this model was:P\u2009<\u20090.05 was considered statistically significant for all tests.Recurrence-free survival (RFS) was collected and defined as the interval from the data of surgery to the end of follow-up results or death. Overall survival (OS) was calculated from the date of diagnosis to the date of death or last follow-up. Survival curves were estimated using the Kaplan\u2013Meier method, and the log-rank test was used to test for differences between groups. The time-dependent receiver operating characteristic (tdROC) curve analysis was first applied to evaluate the predictive accuracy of the model for cancer-specific death or biochemical recurrence based on the risk scores, with the help of the \u201csurvivalROC\u201d R package. Chi-squared test or Fisher\u2019s exact test were used to investigate the correlation between risk model and clinicopathologic variables, and draw the heatmap through the \u201cpheatmap\u201d R package, 2 test or Fisher\u2019s exact test, and draw the heatmap through the \u201cpheatmap\u201d R package, P\u2009<\u20090.05 was considered statistically significant for all tests.Consensus clustering was performed using the \u201cConsensusClusterPlus\u201d package in R to identify subgroups based on the target genes of goserelin. This algorithm determined consensus clustering by measuring the stability of clustering results from the application of a given clustering method to random subsets of data. In each iteration, 80% of the tumors were sampled, and the k-means algorithm with the Euclidean squared distance metric was used. These results were compiled over 100 iterations. After executing ConsensusClusterPlus, we obtained the cluster consensus and item-consensus results. Graphical output results included heatmaps of the consensus matrices, which displayed the clustering results, consensus cumulative distribution function (CDF) plots, and delta area plots, and which allowed us to determine an approximate number of clusters. Numbers of clusters were determined according to the following criteria: relatively high consistency within the cluster, relatively low coefficient of variation, and no appreciable increase in the area under the CDF curve. Associations between both clinicopathologic characteristics and clustering were analyzed using the \u03c7http://bioinformatics.psb.ugent.be/webtools/Venn/). We inputted the retained candidate target genes into the UALCAN  database. Wilcoxon signed-rank test was used to generate a P-value for expression of age, gender, stages, different molecular subtypes, nodal metastasis status, menopause status, or gleason sorces analysis, all P-values\u2009<\u20090.05 were considered statistically significant.We are screening the representative target genes through the Venn diagram , we identified 202 specific BP, 15 CC, and 21 MF of GO terms were enriched in these genes  and 551 prostate samples . The genes that met the cutoff criteria of a fold change\u2009>\u20091 and an adjusted The RNA expression profiling data of cells were downloaded from the GEO database. Heatmap analysis showed that genes have differential expression profiles between breast MCF-10A cells and 3 UFH-001 cells  and peri-menopause (P\u2009=\u20090.01), N1 was higher than N0 (P\u2009=\u20090.02), luminal subtype was higher than triple negative breast cancer (TNBC) (P\u2009=\u20090.005) and HER-2 positive subtype (P\u2009<\u20090.001). However, the expression levels of CRABP2 based on gleason score group, nodal metastasis group, and molecular signature group in prostate cancer were lower than normal controls , and stage M (P\u2009<\u20090.05). While survival status (P\u2009<\u20090.05) was the only difference between high risk and low risk groups in the prostate cancer risk model.In order to investigate the prognostic value of these 76 target genes in breast and prostate cancer, respectively. Univariate Cox survival regression analysis was performed based on the expression levels of the genes from TCGA Figs.\u00a0a and 7a.P\u2009<\u20090.001) of breast cancer patients in univariate analysis and only risk score and age at diagnosis were still significantly related to OS  in multivariate Cox regression analysis of prostate cancer patients in both univariate and multivariate Cox regression analysis  and age (P\u2009=\u20090.004), CD3E expression levels (P\u2009=\u20090.017) were only related to age, GSTZ1 (P\u2009=\u20090.040), GZMM (P\u2009=\u20090.006), GRHPR (P\u2009<\u20090.001), UBTF (P\u2009=\u20090.004) and CHP1 (P\u2009=\u20090.003) were significantly related to stage M  and stage N (P\u2009<\u20090.001), , cluster 7 had the best prognoses, while cluster 2 had the worst , cluster 5 had the best prognoses, while cluster 1 had the worst  curve. We calculated average cluster consensus and the coefficient of variation among clusters depending on category number Figs.\u00a0a and 9a.rst Fig.\u00a0c. Kaplanrst Fig.\u00a0d and 9d.Women younger than 35\u00a0years are more demanding for fertility. Goserelin is the most commonly ovarian suppression strategies among premenopausal women with hormone positive breast cancer owing mainly to its non-invasiveness and reversibility . The genP value\u2009<\u20090.05 was shown in Fig.\u00a0Kaplan\u2013Meier survival analysis of OS and RFS were performed based on ICGC survival data, and only log-rank Venn diagram summarize amount of predicted composed gene of risk prediction models links for DEGs, OS, hub genes or clinical factors. Eventually, CORO1A and ANXA5 as the representative target genes for breast cancer Fig.\u00a0a and DPPP\u2009<\u20090.05, Fig.\u00a0P\u2009<\u20090.001), post-menopause was higher than peri-menopause (P\u2009=\u20090.041), N1 was higher than N0 (P\u2009=\u20090.048), triple negative breast cancer (TNBC) was higher than luminal subtype (P\u2009<\u20090.001). The expression of ANXA5 was significantly lower in breast cancer patients than normal controls in subgroup analysis based on gender, age, menopause status, disease stage, nodal metastasis status, and molecular subtypes  and HER2 postitive (P\u2009=\u20090.003) subtypes, peri-menopause was higher than per-menopause (P\u2009=\u20090.028) and post-menopause (P\u2009=\u20090.002), and as the age progressed, the expression of ANXA5 decreased , ERG fusion (P\u2009<\u20090.001), SPOP mutation (P\u2009=\u20090.001) groups  due to its advantage of being easy to administer, noninvasive and reversible. As palliative treatment for premenopausal women with ER-positive breast cancer, goserelin may similarly rival ovariectomy . AndrogeIn the present study, we identified 18 DEGs in breast cancer tissues and 5 DEGs in cells, and 6 DEGs in prostate cancer tissues and 9 DEGs in cells. CRABP2 is the common DEG both in breast and prostate cancer. Previous reports state that\u00a0abnormal expression of CRABP2 is associated with malignant cancers in the human being . In our On the basis of various prospective and retrospective studies of breast cancer, age is an independent prognostic factor with worse survival \u201326. ReceAlthough a combination of multiple genes manifests more efficiently than a single gene as a diagnostic or prognostic biomarker, the predictive role of single gene was also explored in our study. CORO1A is highly expressed in cells of the haematopoietic lineage, where it has been predominantly investigated in lymphocytes, macrophages, mast cells and neutrophils , but hasIn conclusion, this study presents a novel signature with demonstrated prognostic value similar in magnitude to that of clinical staging of breast and prostate cancer, and having added value in very young breast cancer patients. This signature can facilitate identification of new biomarkers which sensitive to goserelin, increase the using accuracy of goserelin and clarify the classification of disease molecular subtypes in breast and prostate cancer. Future experimental and clinical studies are necessary to produce a solid confirmation of our results.Additional file 1: Figure S1. The compound and compound target genes of goserelin. (a) Chemical structures of goserelin. (b) Protein\u2013protein interaction network of target genes of goserelin. (c) 10 hub genes in protein\u2013protein interaction network. Figure S2. Construction and evaluation of the very young breast cancer risk prediction model in ICGC. (a) Overall survival (OS) in univariate Cox regression of target genes. (b) Recurrence-free survival (RFS) in univariate Cox regression of target genes. (c) Lasso regression for genes in univariate Cox regression of OS. (d) Lasso regression for genes in univariate Cox regression of RFS. (e) Kaplan\u2013Meier survival curve (OS and RFS) for patients with high-risk group and low-risk group. (f) ROC curve showed the predictive efficiency (OS and RFS) of the risk score. Figure S3. The differentially expression of representative target genes between male and female in multiple normal human tissues by GTEx data, *P\u2009<\u20090.05, **P\u2009<\u20090.01 and ***P\u2009<\u20090.001. Table S1. GO analysis of target genes of goserelin. Table S2. KEGG pathways enrichment analysis of target genes of goserelin."}
+{"text": "Reinforced concrete is used worldwide in the construction industry. In past eras, extensive research has been conducted and has clearly shown the performance of stress\u2013strain behaviour and ductility design for high-, standard-, and normal-strength concrete (NSC) in axial compression. Limited research has been conducted on the experimental and analytical investigation of low-strength concrete (LSC) confinement behaviour under axial compression and relative ductility. Meanwhile, analytical equations are not investigated experimentally for the confinement behaviour of LSC by transverse reinforcement. The current study experimentally investigates the concrete confinement behaviour under axial compression and relative ductility of NSC and LSC using volumetric transverse reinforcement (VTR), and comparison with several analytical models such as Mander, Kent, and Park, and Saatcioglu. In this study, a total of 44 reinforced-column specimens at a length of 18 in with a cross-section of 7 in \u00d7 7 in were used for uniaxial monotonic loading of NSC and LSC. Three columns of each set were confined with 2 in, 4 in, 6 in, and 8 in c/c lateral ties spacing. The experimental results show that the central concrete stresses are significantly affected by decreasing the spacing between the transverse steel. In the case of the LSC, the core stresses are double the central stress of NSC. However, increasing the VTR, the capacity and the ductility of NSC and LSC increases. Reducing the spacing between the ties from 8 in to 2 in center to center can affect the concrete column\u2019s strength by 60% in LSC, but 25% in the NSC. The VTR and the spacing between the ties greatly affected the LSC compared to NSC. It was found that the relative ductility of the confined column samples was almost twice that of the unrestrained column samples. Regarding different models, the Manders model best represents the performance before the ultimate strength, whereas Kent and Park represents post-peak behaviour. Concrete is the most widely used construction material in all civil engineering projects across the world. A manufactured product, concrete is composed of cement, fine and coarse aggregate, water, and admixture(s) . ConcretA column is a compressive structural member that takes all the vertical load from the slabs and beams and safely transfers it to the ground. Columns are also designed to resist lateral forces that come from the wind and earthquakes. Additionally, columns are frequently used to support beams, slabs, or arches on which the upper part of the wall or ceilings rests . A columThe work on confinement concrete was started in the early 1900s. For the first time, the concrete strength improvement with the closing effect was initially reported by the Consideration in 1903 for concrete with NSC ,13. AfteWhen a column is loaded with an external force parallel to its central axis, the external force is called the axial loading force. Axial loads are classified into concentric and eccentric axial loads . This stDuring a concentric axial compressive load, the centre concrete shortens in the longitudinal direction and expands in the lateral direction due to the effect/action of poison ratio . Core coHSCs have been used in multi-story buildings and long-span bridges to reduce structural element size and take up little space. However, the stress\u2013strain curve of the HSC is a relatively steep and short post-peak branch which become more brittle than the NSC and LSC. Most HSC columns with a concrete cylindrical strength of 100 MPa could be very brittle if sufficient confinement reinforcement was not provided . Most enThe confinement effect of lateral transverse steel cannot be judged based on analytical models for low strength concrete.  suggesteIn previous decades, the utmost scientists and researchers have worked on various methods to advance concrete performance and ductility ,42. HowePrevious research has experimentally investigated SSC and HSC\u2019s confinement behaviour and validated it analytically with several analytical models . HoweverMost of the research work is performed on the analytical model of SSC and HSC performance, but minimal work is carried out on LSC (2000 psi) and NSC (3000 psi). With the absence of experimental work, it is impossible to use the same theoretical model as standard and HSC to determine low-strength concrete performance subjected to axial compression. An in-depth study/research is required to find LSC confined performance under compression restricted with the least spacing of transverse reinforcement. The current research mainly focuses on tie spacing, volumetric transverse ratio, relative ductility, and concrete strength. The flowchart shown in 3, whereas the specific gravity is 2.69, and the absorption rate was 0.53 per unit by weight as per ASTM C128-15 [Different materials were used in the mix design and concrete column specimens, such as fine and coarse aggregate, hydraulic cement, and reinforcement. As per ASTM C136-14, the results obtained from the sieve analysis of fine aggregate are presented in  C128-15 . However C128-15 . As per  C128-15 , that waIn this research, deformed bars were used as longitudinal and transverse reinforcement. Before using steel as a reinforcement, it is necessary to evaluate and know a material\u2019s properties. UTM has a capacity of 200 t was used to find the reinforcement\u2019s tensile and bending properties. In all column specimens, the deformed steel bars of 4 bars #4 bar with a grade of 60 were used as longitudinal reinforcement. Two different randomly deformed steel bars of #3 and #2 bars were used for the transverse reinforcement in the concrete columns. For the tensile strength of the randomly selected transverse and longitudinal reinforcement, the UTM capacity of 200 t was used. The results obtained from the UTM are represented in 3, whereas the mix proportion of cement is 238.41 kg/m3 (3.43 kg), fine aggregate is 603.96 kg/m3 (8.64 kg), and coarse aggregate is 822 kg/m3 (11.84 kg) for a single column. Using a 0.57 w/c ration, the proportion of water is 1.955 L. Twenty-two NSC and LSC casing specimens were cast in two sets of three, each with the same size, having the same longitudinal but different transverse reinforcement. The slump ranged from 1\u20132 in, and the absorption rate of coarse aggregate was 0.53 per unit by weight. In contrast, the fine aggregate\u2019s specific gravity and fineness modulus were 2.73 and 2.69, respectively. As per ASTM C39/C39M-20 [Mixture proportioning is a process of selecting suitable ingredients and determining their relative proportions with the objective of producing concrete with a certain minimum workability, strength, and durability as economically as possible. The theoretical experimental method was used to determine the recipe of the concrete mix. The design of the concrete mixture began with determining the qualitative features of components and determining their basic properties. On the basis of general formulas, the proportions of aggregates were selected, followed by the amount of individual concrete components. Using the properties of fine and coarse aggregate and cement as mentioned above, the mix design of concrete was carried out for 28 days of 3000 psi strength. The concrete mix ratio was taken by its weight of 1:2.28:2.76 (cement: fine aggregate: coarse aggregate), with a w/c of 0.57. The volume of one cylinder is 0.0144 m/C39M-20 , three cA column specimen with a length of 18 in and a cross-section of 7 in \u00d7 7 in was designated for the concentric axial compression. Each sample is reinforced with longitudinal/vertical bars of 4 bars #4 bars. The horizontal/transverse reinforcement is provided in square hoops with variable ties spacing #3 and #2 bars. Due to the limited height available between the two jaws of the Universal Testing Machine (UTM) loading unit, the height of the column specimen was kept at 18 in. Column specimens were fabricated and cast for mix designs of 3000 psi (NSC) and 2000 psi (LSC) cylinder compressive strength. The column specimen details and terminology are given in Mixed designs were used for concrete strength of 2000 and 3000 psi. The steel cages were placed in the formwork with a transparent cover of 3/4 in between the centre line of rectangular hoops at the formworks\u2019 inner surface. The columns were cast in three layers, and each layer was vibrated before placing the next layer through a vibrator. The casting of the concrete column specimens is shown in After casting the sample, all concrete columns and cylinders are covered with a wet hessian sack for thoroughly moist curing. After the curing, the concrete columns were instrumentally set up and placed in the UTM for testing.The testing setup arrangement is a critical stage to acquire accurate data from testing column specimens. Each column sample was fully equipped and arranged to collect the desired data. For longitudinal deformation of the column sample, two displacement transducers with a capacity of 60 mm were connected on the front and rear face sides of the column sample to achieve the deformation on both sides. The calibration length was 9 in on both sides of the sample. Displacement sensors were fixed in each face\u2019s middle, as shown in As shown in Both sides of the column specimen were caped with plaster of Paris with a depth of 12 mm to distribute the load on the column specimen uniformly. Furthermore, for the uniform distribution of loads, a steel plate of cross-sectional 8 in \u00d7 8 in was placed on the column specimen\u2019s upper surface. The deformation gauges with a 60 mm capacity were installed on both sides to record the deformation before spalling the concrete cover. The gauge length for each displacement was kept at 9 in and was calibrated and connected with the data acquisition system.The samples were strictly aligned with the UTM loading unit to certify concentric axial loading on the sample. The loading rate was 20\u201350 psi and remained constant throughout the test. The load continues until the column sample collapses due to the longitudinal bars\u2019 compliance or rupture of the transverse hoops. After testing the column samples, the data were documented and saved in a system for further analysis.W = load in a pound (lb), A = average of the gross area of the upper and lower bearing surfaces of the specimens (in2).As the load advanced to the peak/ultimate value, the cracks appeared on all sides of the column specimen, as shown in A column\u2019s ductility is greatly influenced by the concrete strength, the ratio of steel in longitudinal and transverse directions, axial load, and the distance between the ties. All of these factors must be considered before evaluating the ductility of columns. When the axial load level is higher than the balanced axial load level, the column becomes a compression failure.The researcher has proposed several analytical equations based on the experimental results to forecast concrete confinement behaviour. The work on the confined concrete was started in the early 1900 century. Considere initially reported the strength enhancement in concrete with the confinement effect in 1903 for the normal strength of concrete. Later on, Richart et al., in 1929 , experim\u03b5co, \u03b5cc, \u03b5cu show the strain, whereas ccf and 0.85cf\u2032 show the stress as shown in ccf is the maximum concrete stress and \u03b5cc/\u03b5cu is the corresponding strain in the Blume et al. model [cf is the longitudinal compressive concrete stress, and cf\u2032 is the compressive strength of confined concrete, whereas c\u03b5 is the longitudinal compressive concrete strain.Blume et al., 1961 , introdul. model . HoweverIn 1967, Soliman and Yu  introducEquation (6) is for the ascending:Equation (7) is for the descending:Manders et al., 1988 , proposeExperimental results illustrate that decreasing the amount of spacing between the ties will increase the axial CS in both the NSC and LSC. Furthermore, volumetric transverse reinforcement (VTR) also affects axial compression and relative ductility. Changing the bar number from #2 to #3 positively affects and increases the concrete columns\u2019 CS and ductility. The ratio of crushing strain to the peak strain is defined as relative ductility. The bar number and spacing between ties affect concrete columns\u2019 relative ductility. As the number of bars increases and the spacing between them decreases, it increases the relative ductility in concrete columns. The relative ductility at VTR #3 and #2 bar of LSC is more than the NSC in 8 in and 6 in c/c, but the opposite in 4 in c/c due to the effect of ties spacing and VTR. The spacing between the ties and VTR greatly affect the LSC relative ductility than that of the NSC. However, reducing the spacing between the ties at a maximum (4 and 2 in c/c) greatly affects the relative ductility of NSC, as shown in In comparing NSC and LSC at different VTR, the NSC shows better ductility in both #2 and #3 bars than LSC. The highest relative ductility was found in the LSC with 2 in c/c of VTR #3 bar, which means that the VTR and the bar spacing greatly affected the LSC. However, overall, the NSC\u2019s relative ductility at both #2 and #3 bar shows high ductility, as shown in In order to study the effect of confinement and ties spacing on confined concrete behaviour, we compare the response curves of the confined concrete, as shown in The core concrete stresses are greatly affected by decreasing transverse steel spacing compared with the bar\u2019s size. The core stresses in LSC are twice the core stresses of NSC. In contrast, the ductility of NSC and LSC remains approximately the same.After testing the concrete columns, the experimental results were compared with the analytical models introduced by Mander et al. in 1988 and Kent and Park in 1972. As different types of confinement were used with different ties spacing, the graphs are consequently different. A comparison was made among the specimens with 8-, 6-, 4-, and 2-in tie spacing column specimens. It has been observed that the relationship suggested by various analytical models are to address the ultimate strength and strain increment of the concrete stress\u2013strain curve. The comparisons conducted here with numerous experimental data provide an adequate correlation with a wide range of relevant parameters. It turns out that a well-generalized relationship works, especially for confined-core concrete with low concrete strength.Compared to 8 and 6 in c/c, the 4- and 2-inch c/c spacings show a large change in the original length and sustain for a long time without fracture. The lateral expansion in the 2 in is meagre due to the internal microcracking and the ties\u2019 spacing. Test results show that significant strength and ductility can be achieved if lateral reinforcement is provided.Likewise, the NSC, the different confinement, and the VTR of LSC were also compared with the analytical models. In LSC, the Kent and Park model best represents the core concrete strength\u2019s performance before, during, and after the peak/ultimate strength until the fracture, as shown in Less lateral expansion due to the close confinement in 2 in c/c provides a high strength and modulus of elasticity. The spacing between the ties increasing will provide low strength and cause buckling in the column. The columns with closed ties of 2 in c/c go to the plastic limit and give warring before the fracture. In contrast, the columns with 6 and 8 in c/c suddenly fracture after achieving the peak/ultimate load, which is more dangerous in civil engineering projects. The lateral restraint pressure on the core concrete is directly related to the concrete columns\u2019 lateral reinforcement. Thus, the larger confining pressure results in better confining efficiency. Compared with NSC, the LSC shows higher strength in 2 in c/c, whereas the rest are the opposite. Moreover, the NSC\u2019s Manders model shows better performance before the ultimate strength, whereas the Kent and Park model offers the best performance after the peak strength. However, the Kent and Park model shows the best performance before and after LSC\u2019s ultimate strength. Overall, it can be seen that there is a slight variation in the experimental and analytical model. In addition, both the models are validated experimentally.The current research experimentally evaluates the compressive strength and relative ductility of confined and unconfined NSC and LSC with varying VTR and ties spacing. In contrast, the experimental results were compared with the analytical models and select the best analytical model that best described low-strength concrete\u2019s detention behaviour with low-pitch cross-sections. Based on the experimental results, the following conclusions can be drawn.Experimental results show that the concrete core stresses are greatly influenced by reducing the transverse steel spacing. As the spacing between the ties increases, the strength of the concrete-reinforced column become decreases. The enhancement in the concrete core\u2019s compressive capacity due to confinement was almost double in LSC confinement than in the NSC column specimens. Keeping VTR the same and by reducing the transverse steel spacing, the axial core compressive strength increases. Therefore, the confinement effect due to transverse steel is more in LSC compared to NSC.Additionally, the axial compressive strength and relative ductility were greatly affected by the spacing between the ties in the reinforced concrete column. Moreover, the reinforcement grade used for the ties also affects the concrete compressive strength and relative ductility. Change in the bar number from #2 to #3 bar increases the compressive strength. Therefore, bar #3 shows the highest compressive strength than the #2 bar.In terms of relative ductility, there was a slight variation in the concrete core\u2019s relative ductility in NSC and LSC. The relative ductility was found to be almost equal for NSC and LSC by comparing the same VTR column specimens. It was found that the relative ductility of the confined column samples was almost twice that of the unrestrained column samples.In comparing experimental results with the analytical models, for NSC columns, the behaviour of the concrete column before the peak is better described by the Manders model, whereas the Kent and Park model better describes the post-peak behaviour of the concrete column. However, there is a variation in behaviour of LSC columns. The Manders model shows high strength, whereas the Kent and Park model shows better behaviour before and after the peak. Overall, it can be seen that there is a slight variation in the experimental results and analytical models. In addition, both the Mander and Kent models are validated experimentally."}
+{"text": "Rehabilitation and palliative care are health care fields with separate histories but some recent convergences. Both have been identified as components within universal health coverage and each is the subject of a supportive World Health Assembly Resolution. We draw on the historiography of the two specialties, a recent systematic review of their engagement with each other as described in 62 studies, and critical policy perspectives to examine how rehabilitation and palliative care have been framed as potential partners in care. We examine the changing patient groups served by each field and the organizational forms that combined rehabilitation and palliative care (CRPC) may take. We explore the implications of such collaboration for the underlying goals and values of the two specialties, where each is the subject of changing definitions with differing responsibilities for regulating access to services as well as assuring and documenting quality. We conclude that to be effective CRPC must adapt to the highly segmented and specialized systems in which it is required to operate, recognizing that rehabilitation and palliative care are themselves co-constructors of such segmentation and specialization, but also potential agents for change. Palliation as a field emerged in the decade after world war two and concentrated at first on those who had lost the \u2018battle\u2019 against cancer. Beyond these militaristic dimensions, they seemed to have little in common.Rehabilitation and palliative care emerged as fields within modern healthcare during the course of the 20th century. Their histories have been mainly separate, asynchronous and unconnected. Rehabilitation as a field gained traction after World War I and was initially focused on military personnel returning from conflictAs they have evolved, however, both rehabilitation and palliation have encountered shared milestones and benchmarks. Both are characterized as interdisciplinary fields and have coalesced around professional societies and groups to build capacity and promote their work. Both have been defined by the World Health Organization in ways that have enhanced their significance and credibility. Each has sought, with some difficulty, to build an evidence base to underpin practice. Both have had to engage with a changing \u2018target group\u2019 of people who might benefit from their services. Both have quality of life as an over-arching goal. Each can be seen within the architecture of current health care policy.In most recent times there has been evidence of connections between rehabilitation and palliation in relation to people with significant burdens of illness, whether or not death is expected. A measure of dialogue has developed between the two specialties. Collaborations have begun that seek to forge joint approaches to service delivery and clinical care. The conceptual language of the two fields has found points of overlap leading to new terminologies and \u2018framings\u2019. Whilst these endeavours are in the main still at an early stage, we believe they merit examination, and as we shall see, there is a body of literature, which now illustrates the encounter between rehabilitation and palliative care. In this paper, we place these collaborative endeavours in a historical and societal context, we explore the evidence on how and using which arguments rehabilitation and palliative care are combining, then, using a more critical lens, we go on to consider the circumstances and perspectives that are shaping their mutual development.. We therefore examine the \u2018problems\u2019 that have been described in order to make an argument for rehabilitation and for palliative care and the emergent argument for bringing the two fields together in combined rehabilitation and palliative care (CRPC).In attending to the latter, we are inspired by a post-structural approach to policy analysis, including a practical guide that asks and seeks to answer specific questions concerning the representation of any particular \u2018problem\u2019 in the policy contextIf rehabilitation focuses on functional capability, palliative care attends to the relief of suffering. They do this through differing frameworks, guidelines and practices. Each envisages a contrasting view of the person who can benefit from their offering. Yet as the ageing population exhibits greater co-morbidity and complexity of health and social care needs, it may well be that rehabilitation and palliation are finding themselves in a shared space. We conclude by suggesting that there might be benefit to be gained in a closer alignment between the two, but we argue that both must begin to forge an understanding of shared societal circumstances, a common language and mode of practice that is most relevant to contemporary needs and policy contexts, and endeavour to be less aligned to institutional histories and the boundaries they impose. As part of that, we argue some contradictions, worries and \u2018silences\u2019 have to be addressed.To summarize, in rehabilitation and palliative care we have two fields, each with distinct longer histories of their own development. Mostly these exist in parallel and are unconnected to one another. Recently there have been some points of convergence but these are still only lightly sketched. Whilst the two fields are substantively different, they do share certain similarities in accounts of their development. Historical writing about each has increased in recent decades, but we are not aware of any work that seeks to contrast and compare the two fields and their histories in ways that lead to a critical post-structural point of view, which is our purpose here.The aims of this paper are: 1) to explicate the separate histories of rehabilitation and palliative care, paying attention to roots, definitions and core concepts; 2) to explore what is known about the policy and practice issues involved in bringing the two together; and 3) to offer a critical analysis of current arrangements and a perspective on future development.In doing so our sources derive from: 1) the historiography of palliative care and rehabilitation; 2) a literature review of the arguments for and the documentation relating to combining the two fields; and 3) inspiration from a post-structural perspective when making sense of rehabilitation and palliative care within areas of health policy.Following Bacchi (4) and Bacchi and Goodwin (5), we ask: 1) What is the \u2018problem\u2019 represented to be? 2) What presumptions or assumptions underlie it? 3) How has this representation come about? 4) What is left unproblematic within it, including silences and differing representations? 5) What effects are produced by this representation of the \u2018problem\u2019? 6) How/where is the \u2018problem\u2019 produced, disseminated and defended?The process of identifying the historical roots of the field is a feature of commentaries on both rehabilitation and palliation, and often begins with the etymology of the key terms in use. Both fields generate a discourse which seeks to establish the work as having a long history, and not simply as a specialty of the modern period. Some scholars comment on references to what is later termed palliation or rehabilitation that can be found within medical treatises published from the mediaeval period onwards.habilitatus/habilitare, meaning \u2018make fit\u2019 or \u2018enable\u2019. American medical literature has associated rehabilitation therapy with the Hebrew termrefusa, meaning healing. Here the ancient roots of the rehabilitation approach are also given prominence \u2013 early Chinese movement practices to relieve pain, 2nd century Galenic interventions to ameliorate military injury, and 5th century Greek exercises for the prevention and treatment of disease. LePlege and colleagues show how this is linked to the distinction between able-bodied and disabled people, that began to assert itself in the 14th century, after the Black Death. By the 18th century, Enlightenment assumptions about the \u2018natural order\u2019 of disability were beginning to be challenged, opening the door to new approaches with rehabilitative intent. Special institutions emerged to treat those with sensory impairments and by the 20th century, to treat disabilities of all kinds. Now medicine, social reform and educational interventions developed so called \u2018restorative practices\u2019 that combined to ameliorate impairment and disability across a broad spectrum. Then in the aftermath of World War I, social programmes to support those wounded and injured in conflict began to focus not just on treatment, but also developed a strong imperative to return disabled servicemen to the workplace, further deepening the moral dimensions associated with disability, re-enabling and rehabilitation.Rehabilitation tends to emphasise its linguistic origins as deriving from the medieval Latin term. He sees evidence of interest in these matters from the end of the Middle Ages and shows that by the end of the sixteenth century medical writings using terms likecura palliativa andeuthanasia medicinalis were well established and having an influence on practice. Stolberg points out that in the early modern period (c15-1800), physicians were significantly engaged in discussion about their human duty to support and care for those whose conditions were clearly incurable. This related in particular to cancer, tuberculosis (phthisis), and \u2018dropsy\u2019, which included heart or liver failure and kidney disease.For palliative care, Stolberg asserts that \u2018palliative care is definitely not an invention of the nineteenth or twentieth century\u2019palliare andpallium in their discussions of early origins. Here the notion is that palliation is a last resort when all curative measures have failed. Cloaking or shielding the patient from suffering is therefore the goal when death is inevitable or imminent. The desired outcome is then the \u2018good death\u2019, often described by European writers in the 19th century as \u2018euthanasia\u2019 and seen as a science that controls the oppressive features of illness, relieves pain, and \u2018renders the supreme and inescapable hour a most peaceful one\u2019. Numerous writers from within palliative care refer to words liketh century there is an emerging celebration of foundational individuals, mainly medical doctors, who began to take a more focussed interest in palliation and rehabilitation. These include the authors of specialist theses, textbooks and even key research articles in medical journals. We thus see how each field also defines itself in relation to oft-repeated biographical narratives that shape its unfolding discourse.For both fields, as the formalisation of basic principles was elaborated, associated with the relief of pain and suffering in the face of mortal illness, or with actions likely to re-enable, promote recovery and enhance physical wellbeing, so we also see the emergence of key individuals who through their writings, teaching and innovative clinical practice, start to influence wider thinking and practice. From this later period and into the 20. He claimed that only one fifth of his patients suffered physical, mental or spiritual discomforts and maintained that for most \u2018death was a sleep and a forgetting\u2019. Of more specific relevance, in so far as his treatise laid out a clear set of prescriptions for the care of the dying, including pain and symptom management, dealing with wider distress, diet, and the organisation of the sick room, is William Munk. His short text entitledEuthanasia: Or, Medical Treatment in Aid of an Easy Death, published in 1887, gained favourable reviews on both sides of the Atlantic and Munk has been described as the grandfather of modern palliative care. Alfred Worcester\u2019s short book, entitledThe Care of the Aged, the Dying, and the Dead, written for an American audience, highlighted the diminishing interest of doctors in care for the dying patient and drew on his own practical wisdom about clinical aspects of the \u2018process of dying\u2019, its associated symptoms, the role of fluids, and the problem of restlessness. He attends to the environment of the dying person\u2019s room, to the need for light and for ventilation and endorses the liberal use of opiates, considering morphine to have \u2018no rival\u2019. He deals with the role of faith and religion, with patients\u2019 visions and hallucinations, and with the question of uncertainty. His work is often seen as foundational to the field of modern geriatrics, but undoubtedly, he is influential in subsequent thinking in palliative care.For palliative care, some later Victorian writers on the care of the dying can be seen reaching a wide audience. Most famous among these is perhaps William Osler, who conducted a study of \u2018the act of dying\u2019 among 486 patients at the Johns Hopkins Hospital between the years 1900\u20131904. Trained as a nurse, social worker and physician, from 1958 she began writing about the need to improve care of the dying and in 1967 established St Christopher\u2019s, the world\u2019s first modern hospice, with a commitment to teaching and research, as well as clinical care. A hospice movement, peppered with other charismatic leaders, then began to develop in many countries, often using variants of the St Christopher\u2019s approach adapted to local conditions, and achieving growing recognition for what was to become a new field of medical and healthcare practice, and which by the mid-1970s, came to be known as palliative care.Certainly, Osler and Worcester had an influence upon Cicely Saunders from the 1950s as she began to formulate her ideas about the care of the dying. The life and legacy of Dr Cicely Saunders has been documented in detail. Modern rehabilitation has several origins, however. In Europe, Robert Jones has been described as the founding father of modern orthopaedics and rehabilitation. With a background in surgery and orthopaedics, Jones developed the treatment and training of crippled children together with the nurse Agnes Hunt at Basechurch in Shropshire, UK. In World War I he joined the conflict as a Captain in the Army Reserve. At the frontlines, he noticed that the treatment for fractures, in war as in domestic hospitals, was insufficient. His efforts to promote better rehabilitation of wounded soldiers led to the establishment of specialized military hospitals where he was assisted by American surgeons. At the end of his career Jones dedicated his time to creating wider social acceptance for people with disabilities, who were highly stigmatised, and helped facilitate the recovery of soldiers to reintegrate them back into society.One of the earliest groups to form around the rehabilitation field was the German Society of Physical Medicine and Rehabilitation, established in 1878. The term \u2018Physical Medicine\u2019, rather than Physiotherapy, was first used by the London Hospital in England in 1921. Like Jones, Warren\u2019s early career was in surgery. In 1935 it took a dramatic turn, when the nearby Poor Law Infirmary was annexed into the West Middlesex Hospital where she was Deputy Medical Superintendent. The majority of the new patients were chronically bed-ridden and many were labelled \u2018incurable\u2019 in view of their musculoskeletal or central nervous system handicap. Warren personally introduced a systematic assessment of all patients and a multidisciplinary rehabilitative approach. \u2018To be successful, rehabilitation must be undertaken by everybody coming in contact with the patient, beginning with the gate porter\u2019, she said, and added that \u2018underlying that approach is a very profound philosophy, which we should do well to adopt\u2019 . At the time this represented a deeply altered way of thinking about chronic care, and of impairment in old age - seen as something malleable. As Gilleard and Higgs show in commenting on Warren\u2019s work: \u2018By \u201cactive\u201d rehabilitation, old age was to be rescued from the margins of society and, through the agency of \u201chospital\u201d medicine, returned to a real and valued position within society\u2019.Also from the UK, Marjorie Warren has been described as the \u2018mother of geriatrics\u2019 and a pioneer in introducing rehabilitation medicine into the care of older people, hemiplegics and amputees. Nine years later, following the efforts of Krusen and colleagues and with the support of philanthropic funding, the American Society of Physical Therapy Physicians was established, later becoming the American Society of Physical Medicine and Rehabilitation. The International Federation of Physical Medicine and Rehabilitation was founded in 1950, and by 2020, renamed as the International Society of Physical and Rehabilitative Medicine, had 135,000 members from 75 countries. Parallel to the development of the Society another important organisation developed, also with its origins in the USA. What is now known as Rehabilitation International was founded in 1922 under the name The International Society for Crippled Children by the social worker Bell Greve and the orthopaedic surgeon Henry Kessler; it had the explicit aim of giving people with disabilities the chance to lead full and productive lives. These two organisations illustrate a tension in modern rehabilitation between a medical/clinical and a social/political focus which is also founded in explanatory models of disability.The origins of modern rehabilitation are also linked to the role of Frank Krusen, the American physician who, after contracting tuberculosis in 1922, became interested in the process of recovery. His focus was on physical medicine, which he established in a programme of physical therapy and inpatient rehabilitation at Temple University in 1929. Other regional associations followed with the Latin American Association of Palliative Care (2000), The Asia Pacific Palliative Care Network (2001), the African Palliative Care Association (2004). In 2003 the first \u2018summit\u2019 on international palliative care development took place in The Hague. Others followed in Seoul (2005), Nairobi (2007) and Vienna (2009) \u2013 leading that year to the creation of the Worldwide Palliative Care Alliance.In 1988, the European Association for Palliative Care (EAPC) was formed in Milan, Italy, and Vittorio Ventafridda, who had been involved in the early discussions and shaping of the World Health Organization (WHO) approach to cancer pain relief, became its first president the following yearFor both fields \u2013 rehabilitation and palliative care - the processes of specialist and policy formalisation begin with the coalescence of ideas, practices, knowledge claims and sites of innovation. This involves the formation of a habitus that will subsequently provide a platform from which to pursue technical and bureaucratic strategies that will facilitate formal accreditation for practitioners and recognition in policy arenas. In this the two fields are not co-terminous. The habitus of modern rehabilitation starts to be formed in the years after World War I, whilst the habitus of modern palliation is largely a product of the period after World War II.. People with physical differences are exposed to rehabilitative practices in order that they can then disappear from discourse and become assimilated into a world where only the able bodied have agency. In this scenario the individual body, rather than the social world is seen as the arena for change. It is a normative orientation centred on compliance.Stiker maintains that central to the rise of rehabilitation as a modern field of practice is the goal of transforming the disabled body in ways that will allow it to fully enter the realm of material production and consumption. It recognises the multi-facetted nature of pain as suffering, which can comprise physical, mental, material, social and spiritual dimensions. Once described as both a nomenclature of inscription and a nomenclature of facilitation, total pain defines the goals of the modern palliative project \u2013 to relieve suffering at the end of life using multi-disciplinary approaches and technologies. It too requires a measure of compliance, including accepting the use of powerful pain-relieving medications that are often demonised in other contexts, being willing to engage in end of life conversations, life review, the reconciliation of troubled relationships, and the acceptance of death.In palliation, the central organising concept is that of \u2018total pain\u2019, first described by Cicely Saunders in the early 1960sth century, both fields sought specialist recognition of these specific knowledge claims and practices. For rehabilitation this was first achieved in the USA in 1947, when \u2018physical medicine and rehabilitation\u2019 was formally recognised by an independent Board, established under the authority of the American Board of Medical Specialties. Forty years later, in 1987, a specialist training in palliative medicine was first authorised by the Royal Colleges in the UK. In the next two decades more than 20 other countries took a similar path; often, as in the USA in 2007, recognising palliative medicine as a sub-specialty of some other field, such as oncology, anaesthetics or cardiology.In the second half of the 20Within these processes of recognition, both rehabilitation and palliative care became the subject of WHO policy documents that in each case define the field of activity and locate it within a discourse of health system planning and resourcing.International Classification of Impairments, Disabilities and Handicaps. But as LePLege and colleagues show, this received mounting criticism from disability groups that pointed to the absence of reference to the social determinants of disability. This was acknowledged in the foreword to the fourth edition of the classification in 1993 and led to theInternational Classification of Functioning, Disability and Health (ICF) in 2001. In turn, the WHO engagement with palliative care came through the route of addressing the global problem of cancer pain relief, in a key 1986 publication. This led on to subsequent work at WHO to shape and define the emerging field of palliative care, and in time to address its particular relevance to population sub-groups, such as children and older people.As early as 1972, WHO produced its. For palliation it occurred in the resolution on palliative care in 2014.Notable for both fields, however, is that each became the subject of a World Health Assembly \u2018resolution\u2019, calling the WHO member states to action. For rehabilitation this came in the resolution on disability in 2005United Nations Convention on the Rights of Persons with Disabilities, which came into force in 2008. This document, known as the CRPD, states in article 26 that rehabilitation services must begin at the earliest possible stage, should be based on multi-disciplinary assessment and should include the provision of assistive devices and technologies. In 2011, WHO laid out a process for the production of guidelines on \u2018health-related\u2019 rehabilitation. The motivations for this included: poverty reduction; a growing population that could benefit from rehabilitation; gaps in provision, access to and quality of rehabilitation; and the need to strengthen services within existing systems. In the same year the WHO World Report on Disability defined rehabilitation as \u2018a set of measures that assist individuals who experience, or are likely to experience, disability to achieve and maintain optimal functioning in interaction with their environments\u2019. In 2017 WHO reinforced its commitment in this area, in a report with recommendations onRehabilitation in Health Systems. The document makes the case for a greater focus on rehabilitation, in the context of ageing populations around the world, and with a particular need for development in low- and middle-income countries. The key recommendation is for rehabilitation to be integrated within health systems, with the ministry of health responsible for this at country level, thereby ensuring more rational and appropriate governance. This should in turn guarantee that rehabilitation contributes to the provision of person-centred care, across the care continuum. Rehabilitation should also be seen as part of universal health coverage, and by implication not an \u2018extra\u2019. Efforts should therefore be made to provide quality, accessible and affordable services to meet needs that have been assessed for specific populations. The document makes considerable strides to match attention to the widely used indicators of mortality and morbidity, with a third dimension, that of functioning, and is the cornerstone to the WHO Rehabilitation 2030 initiative. One global estimate suggests that 2.41 billion individuals have conditions that would benefit from rehabilitation.The 2005 disability resolution drew on elements from a United Nations ruling, going back to 1993 and specifying that \u2018States should ensure the provision of rehabilitation services to people with disabilities in order for them to reach and sustain their optimum level of independence and functioning\u2019. The resolution was in turn given further support by the. A review of rehabilitation definitions, however, found no specific means or interventions which could define rehabilitation and found that rehabilitation has been variously described as \u2018a set of measures\u2019, \u2018a process\u2019 and \u2018a health strategy\u2019. A new definition is being prepared in the Cochrane Rehabilitation group, which issued a provisional wording in October 2020: Rehabilitation is a \u201cmultimodal person-centred process including functioning interventions targeting body functions, and/or activities and participation, and/ or the interaction with the environment\u201d (..) aimed at \u201coptimising functioning\u201d (..) in (1) persons with health conditions (a) experiencing disability or (b) likely to experience disability, and/or (2) persons with disability\u201d (p659). The 2011 WHO definition of rehabilitation has been criticised for being too narrow, in describing rehabilitation as \u2018a set of measures\u2019. It came in the form of a technical report that considered more broadly what could\u2014and should\u2014be done to comfort patients suffering from the distressing symptoms of advanced malignant disease and also marshalled arguments for palliative care based on the magnitude of unrelieved suffering experienced by the majority of terminally ill people. Although methods for the relief of pain continued to be emphasized, other physical, psychological, social and spiritual needs for comfort were also included in the report's recommendations. This conceptualisation of palliative care turned on its concern with quality of life and comfort before death, emphasising the family as the unit of care, dependence on teamwork, and its relationship to curative interventions. It framed palliative care as: \u2018\u2026 the active total care of patients whose disease is not responsive to curative treatment\u2019, described the goal of palliative care as achievement of the best quality of life for patients and their families, and saw many aspects of palliative care as applicable earlier in the course of the illness in conjunction with anti-cancer treatment. The first major WHO milestone, specifically oriented to palliative care, rather than the narrower field of cancer pain relief, occurred in 1990. Globally, the field was becoming better known to policy makers and practitioners, debates about its mission and scope were proliferating, and there was an increasing interest in expanding its vision beyond terminal care and oncology to make it available to patients and families where diseases of various types were not so far progressed, or where the distinction between curative and palliative approaches might not be so clear cut. Published in 2002, the second WHO definition saw some critical changes and is summarised as: \u2018Palliative care is an approach that improves the quality of life of patients and their families facing the problems associated with life-threatening illness, through the prevention and relief of suffering by means of early identification and impeccable assessment and treatment of pain and other problems, physical, psychosocial, and spiritual\u2019. Twelve years later, in 2002, a new definition of palliative care appeared from the WHODespite, this growing interest, it was not until 2014 that the World Health Assembly drew attention to the need for action on the limited availability of palliative care in most of the world, the avoidable suffering of millions of people and their families, and the need to create or strengthen health systems that include palliative care as an integral component of treatment.Lancet Commission Report on Pain and Palliative Care introducing the phrase \u2018serious health related suffering\u2019 (SHRS), declared this to be the problem on which palliative care should be focussed, and stated that over 40 million people experienced SHRS in 2015, of which 25.5 million died. By this means, considerable expansion took place in the \u2018target group\u2019 of people who might benefit from palliative care, many of whom may not be imminently dying. The following year the International Association for Hospice and Palliative Care proposed a new definition and formalised it in a publication of 2020: \u2018Palliative Care is the active holistic care of individuals across all ages with serious health-related suffering. Suffering is health related when it is associated with illness or injury of any kind. Health-related suffering is serious when it cannot be relieved without medical intervention and when it compromises physical, social, spiritual, and/or emotional functioning \u2026 because of severe illness , and/or is burdensome in symptoms, treatments, or caregiver stress \u2026 and especially of those near the end of life. It aims to improve the quality of life of patients, their families, and their caregivers\u2019.The 2002 definition of palliative care is still the WHO position today. But in 2017 aThrough these diverse vectors of clinical research and reflection, the growth of professional societies, the focus on definitional matters and the engagement of non-state international actors in policy formation, we see that each field also seems to have a wider relationship to a deeper societal reality.. His project seeks to uncover how the \u2018difference\u2019 of disability was over-valorised in earlier periods, but under-valorised in the modern \u2018age of rehabilitation\u2019, in a context where rehabilitation is co-extensive with pluralised disabilities and their extension in time.For rehabilitation, the close relationship is with disability. Stiker\u2019s classic work uses the social construction and semiotics of disability to reveal how the condition has shifted from something that is seen as innate, fixed and natural in some people, to one that can be construed as temporary and malleableFor palliation the key relationship is with death. In the earliest writings, palliation is seen as a means to relieve suffering when death is inevitable and imminent. It is essentially focussed on the deathbed and its medical management. In its more recent manifestations, palliation is seen to have a wider role, located within a context of prognostic uncertainty and where the promotion of quality of life and wellbeing in the face of serious illness is the primary goal. This means, at least in relation to claims made for the modern field, that palliation should be introduced earlier in the trajectory of disease, accompanying and not displacing curative measures, and should also be available to those with chronic and complex conditions, where there may be no singular diagnostic starting point to a disease trajectory that is subsequently extended and erratic in its course. In this context, notions of \u2018terminal care\u2019 are played down and the scope of palliative care is widened.So both fields relate to cultural understandings associated with stigma, inclusion and integration, functionality, fear, suffering and pain. They also respond to and in turn they shape and influence the changing perceptions of important clinical framings of \u2018disability\u2019 and \u2018terminal illness\u2019. As we shall see, it is in the inter-relations with these themes that each field seeks to generate new, progressive, even transformative narratives. That in turn has provided the conditions of possibility for them to come together in a new collaborative sub-field of practice.In part we can see the two fields dialectically. They appear as opposite forces, engaged in different struggles. Rehabilitation seeks to triumph over adversity, chronic illness and disability, thereby opening up new life possibilities. Palliation seeks to relieve suffering, not to cure or modify its underlying cause, and in so doing it aims to bring about acceptance and short-term quality of life improvement in the face of inevitable death. We can see, however, some resolution of this dialectic. New treatment modalities, palliative interventions that extend life, innovative health technologies and an ageing population in which significant numbers of people experience multiple or co-morbidities, are all blurring the boundaries between the two fields and the target groups to which they are orientated. What then, we might ask, are the opportunities that result from a closer and more collaborative encounter between the two specialties? Thinking in this way the dialectic starts to be resolved and actors begin to see new forms of innovative practice that flow from mutual engagement.. For these reasons, universal health coverage has become a major goal for health reform in many countries and a priority objective of WHO. In so doing it has linked rehabilitation and palliative care in a continuum of care, and by extension fostered dialogue between them.Until very recently, in the key rehabilitation policy documents from WHO, the issue of living with a life-threatening disease and the need for palliative care was not addressed. Likewise, there were initially no references to rehabilitation in the palliative care documents. The key linking point that brings the two together is around the notion of universal health coverage. WHO states that \u2019Universal health coverage is defined as ensuring that all people have access to needed health services  of sufficient quality to be effective while also ensuring that the use of these services does not expose the user to financial hardship\u2019. It asks if rehabilitation in palliative care is asking the impossible and concludes to the contrary, by affirming the value of \u2018palliative rehabilitation\u2019. Acknowledging the apparent contradiction between the two fields, Hockley draws on earlier work, mainly from the oncology setting which ventures to bring forward a connection between the two. In particular she highlights the work of Dietz, who argued that the goal of rehabilitation for people with cancer is to improve quality of life for maximum productivity with minimum dependence, regardless of life expectancy. For Dietz, there were four modes of cancer rehabilitation: preventative, restorative, supportive and palliative. Hockley takes the view that in palliative care, the goal of improving quality of life requires attention not only to physical symptoms but should also include \u2018social, emotional and vocational\u2019 dimensions: \u2018Aligning rehabilitation to palliative care helps to sharpen the focus on these other aspects, integrating the expertise of multi-disciplinary professionals\u2019 (p10).A reflective and seemingly landmark paper by Hockley from 1993 explores this issue, long before it appears to have gained policy endorsement. A significant amount of content had emerged in the 23 years since the Hockley paper. The review covered publications in English, Swedish, Norwegian and Danish, published 2003\u20132013. Sixty-two articles were included, divided into four areas of focus: recommendations (29), patient perspectives (8), structure, organization, models and referral processes (36) and interventions (12). The literature was found to be of varying quality, with an absence of control groups in the intervention studies and inadequate methodological reporting.In 2016 the Danish Knowledge Centre for Rehabilitation and Palliative Care published a narrative synthesis review of the literature concerning the arguments and the evidence for combining and coordinating rehabilitation and palliative care for people with life-threatening diseases including chronic obstructive pulmonary disease (COPD), stroke, cancer, and also for geriatric patientsHowever, a narrative synthesis asking why, when, how and for whom rehabilitation and palliative care are combined, resulted in four themes, reflecting the arguments for bringing the two fields together, but at the same time exhibiting a lack of consensus on when and how rehabilitation should be offered, the relevance of rehabilitation to non-cancer conditions, and recommendations for co-ordinating rehabilitation and palliative care.Whilst recognising the need to clarify the differences and similarities of practice in the two fields, these arguments included the recognition that patients may need both types of intervention, with appropriate coordination between them. The unmet \u2018rehabilitative\u2019 needs of people in palliative care (and vice versa) were promoted as arguments for combining the two. These unmet needs were often articulated by relatives. Mostly the needs for rehabilitation for those in palliative care comprised: the need to be normal and in control, the need for better every day functioning and mobility, and the need to alleviate the fear of being a burden. People in rehabilitation, however, might have unmet expectations that professionals should take up end-of-life questions, though some studies documented the aversion to having end-of-life conversations.Another argument for combining the two approaches was that patients, relatives and professionals may have changing, conflicting and ambivalent preferences for rehabilitation and palliative care and these should be met by both perspectives, simultaneously. There was also awareness of unpredictability in the disease course for many patients in ways that can require ad hoc and co-operative solutions. This argument was particularly prominent in the context of COPD. This in turn raised organizational issues about coherent engagement between the two sets of services. Studies showed that professionals in the field of rehabilitation was unlikely to identify palliative care needs, and conversely. Finally, there was a poorly documented argument that coordinating rehabilitation and palliative care would maximise efficiency.The review found no consensus on when in the continuum of care, rehabilitation and palliative care become relevant. With regard to the timing and coordination of rehabilitation and palliative care during the course of the disease, the review identified three models: dichotomic ; progressive/complementary ; and ad hoc/complementary .Yet the literature study showed that both the notion of a phased course of disease and the widely used idea of a sliding transition from rehabilitation to palliative care are contestable. The relationship between rehabilitation and palliative care varies across diagnosis and individual courses of disease, depending upon prognosis and the situation and preferences of the patient and his or her relatives. The transition from rehabilitation to palliative care can be challenging for patients, relatives and health care professionals. The idea of a breakpoint where the switch from \u2018cure to care\u2019 takes place is not well supported in clinical practice or in the illness experiences of people living with life threatening disease. The review did not support the notion that coordination of rehabilitation and palliative care is particularly relevant in the early course of the disease, nor is there evidence in support of the recommendations concerning when rehabilitation and palliative care should be coordinated. The literature review included patients with COPD, stroke, cancer and geriatric patients and found that coordination of rehabilitation and palliative care may be relevant for these groups of patients or conditions \u2013 as well as for other patient groups with progressive disease leading to death, though these were not included in the review.Papers arguing that rehabilitation should be available across the entire illness trajectory, including the terminal phase, were found for all included target groups. Five papers were from cancer care, four papers from COPD, two papers were from geriatrics and two were from stroke. The review showed that coordination of rehabilitation and palliative care can be organized in various ways. These include partly integrated units within which there is rehabilitation with palliative components, or palliative care with rehabilitation components, or alternatively in co-operating units where otherwise independent rehabilitation and palliative care teams work together. Rehabilitation components are predominantly represented by the integration of single professions  adopting a function focused approach. Palliative care components are predominantly addressed in terms of end-of-life conversations.The review found no evidence of fully integrated units, where rehabilitation and palliative care are organised together in equal measure.It highlighted recommendations to establish effective coordination of rehabilitation and palliative care, including: systematic assessment procedures ; individual plans and objectives which are revised frequently; an appropriate balance between autonomy, help and support; and outcome measures to establish effectiveness. Intervention studies on \u2018palliative rehabilitation\u2019 were considered as interventions that combined rehabilitation and palliative care. The limited evidence base in this area was noted, though a handful of studies did suggest that people with serious and progressing illnesses may benefit from rehabilitative interventions in the later stages of their disease. Five out six palliative rehabilitation intervention studies were about cancer.et al. is now in process, still, not published, and preliminary results indicate an increase in palliative rehabilitation intervention studies . A Danish study by Nottelmannet al. for example, describes a palliative rehabilitation intervention model, for patients newly diagnosed with advanced cancer, and presents data on how it was utilised during a randomised control trial. In Portugal, another review is underway on rehabilitation interventions in palliative care. Its protocol states that \u2018Both palliative care and rehabilitation share essential characteristics in that they are symptom-oriented approaches that focus on function and comfort within a holistic framework. The goal is to promote independence in self-care activities, better symptomatic control, and stabilization of functional decline in line with individual life preferences. Ongoing assessment of patient response indicating improvement, stabilization, or deterioration is conducted, and regimens are modified as appropriate\u2019 (p.2350). From 2016 onwards, more publications have addressed rehabilitation and palliative care as coordinated efforts. A new literature review, by Thuesen seem to support those from the 2016 review and indicate that more interventions are being developed, mostly in the form of \u2018palliative rehabilitation\u2019. They also seem to indicate that social integration and inclusion in society is not mentioned as a goal when rehabilitation encounters palliative care. Rather, functioning as a goal is described in terms of basic activities of daily living and mobility. It appears that, when rehabilitation is described within a palliative care context, the goal of rehabilitation is narrowed down.Preliminary findings from the Portuguese studyBased on the above explorations, it is possible to draw out some analytical similarities and differences between the core elements of rehabilitation and palliative care .. These state-initiated public health strategies are forged at the macro level of policy and government, organized and assessed at the meso level of health care authorities and professional bodies, but also resonate at the individual level in which they are actively internalised by citizens in society.Our account of the histories and more recent coming together of the fields of rehabilitation and palliative care raises a number of critical issues. It is helpful to situate these in their wider societal context. Both fields demonstrate some of the characteristics of healthcare delivery and governance in a changing policy context. In particular, we see them shaped by elements of public health and social care strategies that seek to regulate populations through cultural and normative inscriptions, for example concerning \u2018reablement\u2019, \u2018autonomy\u2019, \u2018a good life\u2019 and indeed \u2018the good death\u2019 and his conception of \u2018the social state\u2019 that dominated Western welfare governance in the first half of the 20th century, we can see that citizens had tobe able to work and provide for themselves and their families. For those who, for a while or good reason, were not able to do so, society was \u2013 ideally \u2013 there to help \u2018from cradle to grave\u2019.Using the perspective of Rose. Hospice care emerged in this context, growing out of charitable, third sector and non-profit organisations, often forging partnerships with state providers, and drawing attention to the social state\u2019s failure to fully attend to care at the very end of life. During the late 1960s and into the 1970s as the need for and possibilities of welfare seemed endless and the economies of western societies did not, social movements as well as economic and political challenges to the social state model of welfare governance led to new strategies of public sector management, NPM gained considerable currency. More recently, ideas about New Public Governance (NPG), with its concerns about the policy challenges involved in public welfare in late modern societies have also been influential.During the 1980s and the 1990s, New Public Management (NPM) became the leading political strategy for governing western societies in an increasingly globalized world. Based on ideas about the construction of an effective public sector through a managed market orientation, internal competition, need assessment and outcome measurement. The prevention of risk was (and still is) seen as mainly an individual responsibility, albeit supplemented by specialist guidance, laws, and regulations. From the 1980s health is therefore defined as the overall concept and the core resource to ensure a life of quality.A further neologism is that of the New Public Health (NPH), which emerged from the 1970s; focusing on the environment, health promotion, disease prevention \u2013 and riskWhile NPG was from the beginning inspired by the claims of social movements concerning citizens\u2019 rights, user involvement and the ownerships of one\u2019s own body, it may now be more concerned with involving citizens and \u2018users\u2019 in securing quality of life and health and social care, across public, private and voluntary sectors and stakeholders.As we have seen, the target groups of both rehabilitation and palliative care have changed and expanded over time and in these changing contexts. The two fields are now crossing over each other in a broader focus on lifestyle diseases, quality of life and the inheritance of a form of welfare thinking that endures across the life course, including health promotion, disease prevention, treatment, rehabilitation and palliative care for all ages and diseases. Crucially from the perspective of global health, this must be in the context of universal health coverage.premature death from and lives spent with prolonged chronic diseases \u2013 and the lack of wellbeing and quality of life related to both, not least in the later stages of life. These challenges are in turn framed as life-style symptoms and diseases.While the \u2018problem\u2019 underlying the introduction of rehabilitation was represented as disablement from war or disaster, the \u2018problem\u2019 through which palliative care was represented was that of unrelieved suffering and \u2018total pain\u2019, when dying from cancer. What then is the \u2018problem\u2019 today and can it be combined for the two fields? In our view the problem represented can be seen asThe main causes of death in western societies are chronic diseases  and many people live and die with associated co-morbidity. At the same time, through better life circumstances and improved treatment, average life expectancy is growing among western populations. Some diseased people live for longer, albeit with severe symptoms of an \u2018un-healthy life-style\u2019, several diseases, and late side effects from treatment. In this context, the population of older people with diminished functionality and in need of care will continue to grow for some time.This understanding of the \u2018problem\u2019 within the two fields explored here leads to expanding target groups and expanded timespans for professional intervention. Rehabilitation may concern reablement after disasters, diseases, traumas, treatment, alcohol abuse and many more causes. The relief of \u2018total pain\u2019 may be relevant to people suffering from a severe disease or situation - from the time of diagnosis or the causative event, and right up to the time of death, and indeed beyond for the relatives concerned. The literature review discussed here confirms that the combination of rehabilitation and palliative care is relevant for all the groups included , and probably several more. It also points to the changing needs and preferences that occur during unpredictable and often prolonged disease trajectories, which in turn support arguments for combining the two approaches.If health and quality of life are defined as the overall wished for outcomes of health interventions throughout the lifespan - where does this leave, or limit, rehabilitation and palliative care, and the combination of the two? As seen in the review, there is no evidence of when and where in the disease course a coordinated effort of rehabilitation and palliative care will be most efficient or effective, even though experienced as relevant.While rehabilitation of soldiers after war, as well as relief of pain among dying cancer patients, was mainly a hospital effort, nowadays rehabilitation and palliative care are taking place in many different settings and contexts, such as in peoples\u2019 private homes, in nursing homes, in hospices, in special institutions, in day care \u2013 and in hospitals. The target groups and contexts for rehabilitation and palliative care, and the two in combination, therefore seem to be expanding and may include many more people in many more types of organisational settings. As seen in the review, CRPC works within and across different organisational contexts.. At the same time this comes with a heavy emphasis on individual responsibility in creating for oneself, not only a healthy life, but also a good death. Within the governance context we have outlined here, the \u2018conduct of conduct\u2019 becomes an important dimension, often realised through exhortatory instruments, an emphasis on compliance and the internalisation of personal responsibilities. These dimensions are of course at work within our two fields of concern \u2013 rehabilitation and palliative care.WHO strategies for public health emphasize the empowerment and enablement of all citizens - defining health as the totality of physical, mental and social wellbeing. Such ideas seem not too distant from the aim of \u2018normalization\u2019 for disabled soldiers returning from World War I. Research in English hospices has documented how \u2018the good death\u2019 is socially constructed and negotiated within a special kind of setting for death and dying, that is excluded from everyday life and for certain groups of dying people. Recent research in Danish hospices has documented how hospice care is continuously negotiated between the values of hospice philosophy and the demands of the public health care system in the context of its requirements for documentation, referral and discharge.One example is a study showing how Danish courses of rehabilitation among cancer patients contribute to the construction of a narrative of \u2018being-as-if-well-again\u2019. For example, when needs assessment was first introduced in the United Kingdom context in the 1980s, health authorities were instructed to refrain from assessing needs that could not be met. We now see rulings governing the number of hours of rehabilitation that should be made available to older people. Likewise, entry to hospice in Denmark is mainly for those imminently dying.Another role for the relevant professionals in making the system work is in aligning rehabilitation, palliative care and now CRPC, with the dominant system logic of NPM. This promotes a focus on competition through quality assurance and the measurement of outcome. Here some of the instruments include documentation procedures, evidence-based practice, quality indicators, clinical guidelines, education, clinical trials and scientific publications. All of these can be seen co-shaping the inclusion and exclusion of patients, the changing roles of professionals, and the structures of organisational practice. Examples include the measurement of needs, goal-setting and quality of life using screening instruments and structured questionnaires, that in turn determine the right to admission or not, the professional effort to be deployed, and the success, or not, of this effort. When rehabilitation and palliative care expand to more extensive target groups and their scope becomes wider, control of admission and access then becomes critical. As described in the review, needs assessment becomes an issue in providing access to rehabilitation and palliative care. Needs is, however, a dynamic concept shaped by policy and resource contextsWhen rehabilitation and palliative care are distributed according to needs, then needs must be considered as negotiated in a social context. The relation between societal strategies, the logic of the health care system and the roles of professional groups and service users can thus be seen to frame the distribution, development, organisation and practice of rehabilitation, palliative care and CRCP. Yet these overall circumstances and their consequences often appear to be silenced within professional discourse, or obscured by the language of prioritisation, cost-effectiveness and efficiency.th century. Weisz sees this as an intellectual strategy that divides problems and people into smaller and more manageable groups, a form of \u2018divide and conquer\u2019. Part of this is the formation of specialties that are actively multi-professional in character, of which rehabilitation and palliative care are good examples. It is well recognised that despite moves towards disciplinary collaboration, medicine remains the dominant health care profession in most systems and jurisdictions. Whilst, as we have seen, medicine and prominent individual doctors play an important role in the history of rehabilitation and palliative care, today the narrative tends to focus on interdisciplinarity and to downplay, or even silence, the dominant role of the medical profession in both fields. As Weisz and others have noted, medicine divided into a large number of specialties and sub-specialities during the 20. Such an over-determined bio-medical distinction is a good example of the cost that results from trying to integrate specialties without addressing their ontological and epistemological foundations.The specialties we describe here and their partial coalescence therefore represent clearly this form of sub-division and also a particular characterization of \u2018problems to be solved\u2019. At the same time, however, merging specialties together is likely to expose fundamental differences in the assumptions, knowledge claims and goals of each field. A recent illustration of this can be found in the Lancet Oncology Commission report on the integration of oncology and palliative care, which sees the former as \u2018tumour directed\u2019 and the latter as \u2018host directed\u2019. But there are also internal worries within each specialty, to the effect that the aim and focus of the other will under-mine and might do harm to their respective patient groups. One example is the worry that the normative focus on recovery, restoration and reablement found in rehabilitation may put pressure on people suffering from severe illness who are being cared for within the palliative care frame of reference . Conversely, there is concern that the lack of attention to recovery and enablement within palliative care, may lead to loss of function and quality of life . Moreover, when rehabilitation and palliative care are combined, both may be changed. As described in the review, when rehabilitation is integrated in palliative care, it may be transformed in favour of function-focused and one-dimensional needs, while the complexity and interrelation of needs may be more difficult to articulate and are therefore silenced.Both rehabilitation and palliative care are specialties that presume they are already including or are able to include the otherThe literature review suggests that actual coordination and combination in clinical practice may be rather sparse, for example, palliative care integrates some functionality focus or rehabilitation incorporates some end-of-life conversations. It seems obvious in this context that CRPC would not replace either of the two specialties, but at the same time those specialties will continue to face challenges concerning their knowledge claims, effectiveness and value to service users and to society.. One of the theoretical issues discussed in rehabilitation is the conceptualisation of disability. As we have seen, the ICF was promoted by WHO in 2001. It was seen as \u2018the answer\u2019 to the previous one-dimensional ICIDH model by expanding to include the biological, psychological and the social, and to practice by operationalizing the bio-psycho-social model within a classification that reflects the person as a body, a self and a social being. The model has had a great impact on thinking and practice in rehabilitation. It has, however, also been criticised for a lack of clarity in its theoretical foundations and empirical testing. Moreover, ICF has been criticized for not addressing temporality and dimensions of quality of life. In particular subjective quality of life is a key missing component of the ICF model. In addition, rehabilitation for older people (reablement) has been criticised for a lack of theoretical understanding of ageing.Rehabilitation and palliative care, like many specialties, appear to lack explicit theoretical definitions of their core concepts and models of practice. The need for theory and theoretical understandings in rehabilitation has been addressed more recently\u2019. Saunders recognized that \u2018Much of our total pain experience is composed of our mental reaction\u2019. Commentators have shown, however, that addressing total pain requires both the prevention of physical pain and attention to patients\u2019 illness experiences. But research about total pain in clinical practice and policy is limited and total pain, despite its central place in the history of palliative care, remains remarkably under-researched, conceptually and empirically. Consequently, engaging with total pain in clinical practice lacks a secure theoretical and analytic foundation.Whilst the definitions of palliative care do not mention the concept of \u2018total pain\u2019 directly, it is at the core of all of them, as the co-thinking of \u2018pain and other problems, physical, psychosocial and spiritualWith such conceptual deficits apparent in both fields, it is evident that considerable work will be required to bring them together in a holistic and effective manner.Rehabilitation and palliative care have different narratives and patterns of development, but are seemingly getting closer to one another. Our assessment of the diverse histories of the two fields and the limited literature on how they are coalescing suggests the logic of collaboration is articulated mainly at the common-sense level. Careful studies of the effects of collaboration and deeper consideration of the conceptual aspects of the two fields working together are largely absent. Yet combining rehabilitation and palliative care approaches appears to make prima facie sense for patients and service users, as well as for professionals involved in trajectories of life-threatening diseases and vulnerabilities. It also resonates with current approaches to health care management and governance. We are enthusiastic about the benefits that might result from closer connections, but take the view that both could profitably attend more to their shared societal circumstances, if a common language and mode of practice is to emerge that is aligned to contemporary needs and robust enough to deal with the current policy landscape, including its opportunities and threats.Analysing these developments from a critical theoretical position \u2013 seeing rehabilitation, palliative care and CRPC as parts of a public health strategy, and asking questions, concerning the \u2018problems\u2019, silences and struggles that are uncovered, brings new perspectives to the narrative. We conclude with two key points, First, rehabilitation, palliative care and CRPC are all located within a health care system which is highly segmented and specialized. This has limitations but has also created opportunities for formal recognition and for collaboration and merger. Second, rehabilitation, palliative care and CRPC are themselves elements in and co-constructors of the current dominant strategies of governance and self-governance in health care. They are both expanding fields, yet increasingly constrained by methods of prioritisation that constrain citizens\u2019 access to them.The interest in combining rehabilitation and palliative care is growing. We look forward to further studies which can throw light on the consequences and implications. At the same time, such work should not proceed without some deep reflection on the conceptualisation of rehabilitation and palliative care in the context of broadly neo-liberal health care systems. Such work will benefit each field individually, as well as in their efforts to collaborate and coalesce.All data underlying the results are available as part of the article and no additional source data are required. This is a very interesting and thoughtful paper. It contains relevant information on the historical development of both specialties, synthesizes their philosophical underpinnings and beliefs and draws these together in a credible way. The paper sets out to examine the histories and possible future of two specialties \u2013 rehabilitation and palliative care. This is both a strength and a weakness. It is a strength as this makes the topic manageable. It is a weakness as it fails to consider other specialties with similar histories and philosophies. The two that specifically came to mind are geriatric medicine and pain medicine, both of which are well developed specialties in their own right. If there is a case to combine rehabilitation and palliative care, why not also include geriatric medicine and chronic pain management? The paper is more an exploration of ideas rather than a research article. Yet some of the ideas could be better defined. The paper suggests the concept of \u2018combined rehabilitation and palliative care\u2019 without spelling out what a \u2018combined\u2019 service would look like. Other terms  tend to be used as though they are interchangeable with the idea of \u2018combined\u2019 service. \u00a0 The key scoping issue in this paper is that it is, by design, theoretical rather than practical. This necessarily raises what I would call the \u2018so-what?\u2019 question. So what if palliative care and rehabilitation share some commonalities? Has the case been made that efforts should be made to bring them together as a combined service? If so how? These questions are not explored in this paper but suggest critical areas for further research and analysis. Australia (like the United States and elsewhere) has come to this issue from a practical perspective that forms a useful contrast to the theoretical framework in this paper. Australia brought palliative care and rehabilitation at a practical level in 1992. It did so by differentiating three types of hospital-based care \u2013 acute, subacute and non-acute care. Palliative care and rehabilitation were brought together as subacute care and this remains the case 30 years later. Subacute care has different data collections and funding arrangements to acute care. The separate introduction of a subacute care stream in Australia was driven by a decision to introduce the Diagnosis-Related Group (DRG) classification  and to use it as the basis of hospital funding. This required that the scope of the DRG system be defined. The term \u2018subacute care\u2019 was coined for use in Australia in 1992 to describe care that should not be classified and funded by DRG. This was the defining characteristic that brought rehabilitation and palliative care together. They were not \u2018acute care\u2019 and should not be funded by DRG. Subacute care in Australia is defined as care provided for a person who requires health services but whose principal medical diagnosis (modified for factors such as age and procedures) is not adequate in explaining the need for, or the cost of, the services that s/he receives. In subacute (function-related) care the predominant goal is the enhancement of a patient's quality of life and/or improvement in his or her functional status. All rehabilitation, all palliative care, some mental health and some aged care are included in the definition of Australian subacute care. The reason I mention the Australian experience is to suggest areas for further research that arise from this paper. While the authors rightly caution on the need to reflect on the conceptualization of rehabilitation and palliative care in the context of neo-liberal health care systems, they essentially conclude that combining rehabilitation and palliative care is fundamentally a good idea. If so, the issue that now needs to be explored is how this is best achieved. This includes the levers for achieving better integration, how potential risks are managed, workforce implications and models of care. Our Australian experience is that the funding model has driven the two together without any serious detriments to either of them. Yet, even 30 years later, it is not clear whether other potential benefits have been realized. These issues remain ripe areas for further research.Is the work clearly and accurately presented and does it cite the current literature?YesIf applicable, is the statistical analysis and its interpretation appropriate?Not applicableAre all the source data underlying the results available to ensure full reproducibility?No source data requiredIs the study design appropriate and is the work technically sound?YesAre the conclusions drawn adequately supported by the results?YesAre sufficient details of methods and analysis provided to allow replication by others?YesReviewer Expertise:Health services research, palliative care, rehabilitationI confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard. This is a clear and important article for current and future practice. The origins and development of both fields provide an interesting background to the arguments raised. The evidence at present is emerging and hopefully, this article will spur collaboration for further research.\u00a0 The article covers many of the problems facing modern healthcare and provides a much-needed discourse into future service delivery models. More collaboration is required between rehabilitation and palliative care if we are to optimally serve the needs of our combined populations. I thought there would be some evidence from the MND literature about \"neuro-palliative rehabilitation\" - a term I have heard from our local MND specialist centre in Melbourne. I also wonder if Alex Jadad's article on the WHO definition of Health might be pertinent in the argument:\u00a0\u201cHealth is the ability that we have as individuals or communities to adapt and to manage the inevitable challenges that we face through life - physical, mental, or social.\u201d - just a thought.Is the work clearly and accurately presented and does it cite the current literature?YesIf applicable, is the statistical analysis and its interpretation appropriate?Not applicableAre all the source data underlying the results available to ensure full reproducibility?YesIs the study design appropriate and is the work technically sound?YesAre the conclusions drawn adequately supported by the results?YesAre sufficient details of methods and analysis provided to allow replication by others?YesReviewer Expertise:Palliative care. Cancer Pain. End-of-Life-Care. Public Health Palliative Care.I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard. effective CRPC must adapt to the highly segmented and specialized systems in which it is required to operate, recognizing that rehabilitation and palliative care are themselves co-constructors of such segmentation and specialization, but also potential agents for change\u201d are echoed in the work of Candy et al.\u00a0and Hortonet al.\u00a0and by as they sought to characterize ways to research and effectively embed complex health interventions into practice.I think this is a very timely and well-constructed paper and would encourage all those working within both specialities to read it. The authors helpfully describe the historical and epistemological founding of both specialities before focusing on how and why they have become entwined. The assertions and sentiments of the authors that \u201c Rehabilitation delivered by palliative care services should not just be about physical function and needs to consider the broader concepts of enablement, choice and autonomy. Likewise, palliative care delivered by rehabilitation services needs to look beyond care choices as death approaches, supporting people to live as well as they can, whilst coping constructively with losses or symptoms arising from their deteriorating health. There is a need to recognise the social, context-sensitive, and theoretical underpinnings of combined rehabilitation and palliative care services. Not to do so will ultimately diminish the potential benefits to patients and families of this approach. I look forward to further studies and reflections on the conceptualisation of both rehabilitation within palliative care, and of palliative care within rehabilitation as these fields continue to evolve. The full reference for reference 46 in the paper is Dietz, J.H. 1981.\u00a0Rehabilitation Oncology.\u00a0UK: John Wiley and sons.Is the work clearly and accurately presented and does it cite the current literature?YesIf applicable, is the statistical analysis and its interpretation appropriate?Not applicableAre all the source data underlying the results available to ensure full reproducibility?YesIs the study design appropriate and is the work technically sound?YesAre the conclusions drawn adequately supported by the results?YesAre sufficient details of methods and analysis provided to allow replication by others?YesReviewer Expertise:Working within specialist palliative care as a dietitian and an allied health professional\u00a0lecturer and researcher since 2004. My PhD focused on rehabilitation in palliative care.I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard."}
+{"text": "Pulmonary alveolar microlithiasis is a rare autosomal recessive condition that is characterized by the formation of excessive calcium phosphate microliths in the alveoli. Most patients are diagnosed in adulthood due to the slow progression of the disease. Children with this disease are asymptomatic, and changes in the lung parenchyma are usually discovered incidentally. The diagnosis is made by the combination of a positive chest imaging and histological examination. Rheumatoid arthritis (RA) is a chronic systemic autoimmune disease characterized by chronic seropositive symmetrical inflammatory polyarthritis with numerous extra-articular manifestations. It targets the lining of the synovial membranes, frequently affects females more than males, and is treated with the disease-modifying antirheumatic drugs (DMARDs). If left untreated, it leads to increased morbidity, mortality, and socioeconomic burdens. In this case, we report a 19-year-old young man who presented with clinical and radiographic features of PAM associated with RA. Pulmonary alveolar microlithiasis (PAM) is a fairly rare lung disease which is characterized by the deposition of alveolar calcium and phosphate in the lung tissues bilaterally and predominantly affecting the lower and middle lung zones. The etiology and pathogenesis are not completely understood , 2.PAM is a genetic disease caused by a mutation in the SLC34A2 gene. This gene is a sodium-dependent phosphate transporter expressed in the lungs which is essential in the removal of phospholipids from the alveolar spaces. Dysfunctional SLC34A2 leads to the accumulation of calcium and phosphate causing microlithiasis [PAM is considered an autosomal recessive disorder and is associated with consanguinity. A majority of patients have at least one affected sibling. Over 1200 cases were reported in the literature, and its prevalence is unknown. 35.8% of PAM patients are diagnosed before the age of 20 and 88.2% before the age of 55 .In early stages, patients are asymptomatic with a usual slow progression. Over time, they develop exertional dyspnea, dry cough, weakness, chest pain, cyanosis, hemoptysis, and pneumothorax. Physical examination may show rales and bilateral clubbing of the finger. Additionally, it has been shown that smoking and infection may hasten the disease progress .PAM is mainly diagnosed by imaging with the support of clinical manifestations. The typical findings of PAM on a plain radiograph are a fine scattered micronodular pattern forming a characteristic \u201csandstorm\u201d sign. The borders of the heart, diaphragm, and vascular markings are obscured due to the dense calcification. Findings on the high-resolution computed tomography (HRCT) scan of the chest are classified into several stages based on the severity of imaging abnormalities. These range from the precalcific phase and calcific micronodules to ground-glass opacity, thickening of the intralobular septa, and the classic white lung sign .Lung biopsy shows classical histopathologic features of PAM which are diffuse filling of alveolar air spaces by calcospherites, lamellated calcifications, fibrosis, and opacification in the late stages .Unfortunately, the role of medical therapies such as systemic corticosteroids, calcium chelating agents, and bronchopulmonary lavage has been ineffective in slowing the disease progression. Diphosphonate therapy usage is still controversial and needs further evidence . The onlPrognosis of PAM remains unclear, but according to a Japanese study which included 53 patients showed that 34.1\u201342.9 % died within 10\u201349 years after the diagnosis, most commonly due to respiratory failure [Rheumatoid arthritis (RA) is a systemic autoimmune inflammatory disease that most commonly affects the joints causing progressive, symmetric, and erosive destruction of the cartilage and bone. It is usually associated with autoantibody production such as rheumatoid factor (RF) and antibodies against citrullinated peptides (anti-CCP). RA affects \u223c1% of the population in developed countries. Although joint disease is the main presentation, there are a number of extra-articular manifestations including lung disease, cardiovascular disease, subcutaneous nodule formation, vasculitis, and inflammatory eye disease. Of these manifestations, lung disease is a major contributor to morbidity. In some cases, respiratory symptoms may precede articular symptoms .In this report, we present a case of PAM with associated preclinical RA.A 19-year-old young man, not known to have any medical illness, presented to the pulmonology clinic for the first time in November 2019 with progressive difficulty in breathing and dry cough for 8 months. He also noted pain in multiple metacarpophalangeal (MCP) and proximal interphalangeal (PIP) joints bilaterally as well as the right wrist and both knees. This pain was not associated with swelling; however, he admitted daily morning stiffness that lasts for up to 30 minutes and improves with activity. The family history was notable for a sister with confirmed diagnosis of PAM. On his initial presentation, the patient was vitally stable with an oxygen saturation of 93% on room air. Physical examination revealed digital clubbing, symmetrical reduction in the chest expansion, bilateral dull percussion note, especially over the lower part of the chest, and vesicular breath sounds with bilateral basal inspiratory crepitation. He had multiple tender MCP and PIP joints, right wrist, and both knees with no evidence of joint swelling, effusion, or deformity. Cardiac, abdominal, neurologic, and dermatologic examinations were unremarkable.Laboratory investigations revealed the following results: WBC 7.92\u2009\u00d7\u200910^9/L , hemoglobin 120.00\u2009g/L , MCV 82.5 FI , platelets 431\u2009\u00d7\u200910^9/L , CRP 7.28\u2009mg/dl , ESR 105\u2009mm/h , rheumatoid factor 8060\u2009IU/mL , positive anti-CCP and antinuclear antibodies, negative anti-ds DNA, P-ANCA, and C-ANCA, and normal complements.The patient was admitted under the pulmonary medicine team for further assessment. Sputum culture tested negative for microorganisms, including acid-fast bacilli. The high-resolution computed tomography (HRCT) scan of the chest showed bilateral macronodular shadows with basal calcification, sand-like calcification, and black pleura sign. These findings were consistent with advanced PAM .Although histopathology is usually required to confirm the diagnosis, the high clinical suspicion along with strong family history and classical radiologic findings were all thought to be sufficient to diagnose this patient with PAM. He was started on systemic steroids and oxygen therapy and was referred to thoracic surgery to initiate lung transplant workup.The patient was also seen by the rheumatology team for assessment of his polyarthralgia. In the light of absent symptoms and signs of inflammatory arthritis but strongly positive serology, he was diagnosed with preclinical RA and was started on hydroxychloroquine therapy to halt/delay the progression to clinical RA.A few months later, while waiting for lung transplant, he presented to the rheumatology clinic for follow-up. He admitted noncompliance to hydroxychloroquine and prednisolone. He had flare of his joint symptoms with multiple tender and swollen joints . BedsideA variety of lung manifestations are known to develop in patients with RA. These include pulmonary parenchymal disease, pleurisy, airway disease, and pulmonary vascular disease . The patOther than the aforementioned lung manifestations, the literature review supports possible association between PAM and RA . Three cUp to our knowledge, our case is the fourth to be reported in the literature. Unlike the previous reported cases, it does not only confirm the association between PAM and RA but also shows that PAM may predate the onset of RA.PAM is a rare lung disease in patients with RA. The etiopathologic link between PAM and RA is interesting yet unclear. We support the theory suggested by the authors who reported the previous cases that PAM can induce an autoimmune process that may initiate the pathogenesis of RA. We think that more cases of PAM-associated RA are needed to further explore this link."}
+{"text": "Persistent pain is sustained by maladaptive changes in gene transcription resulting in altered function of the relevant circuits; therapies are still unsatisfactory. The epigenetic mechanisms and affected genes linking nociceptive activity to transcriptional changes and pathological sensitivity are unclear. Here, we found that, among several histone deacetylases (HDACs), synaptic activity specifically affects HDAC4 in murine spinal cord dorsal horn neurons. Noxious stimuli that induce long-lasting inflammatory hypersensitivity cause nuclear export and inactivation of HDAC4. The development of inflammation-associated mechanical hypersensitivity, but neither acute nor basal sensitivity, is impaired by the expression of a constitutively nuclear localized HDAC4 mutant. Next generation RNA-sequencing revealed an HDAC4-regulated gene program comprising mediators of sensitization including the organic anion transporter OAT1, known for its renal transport function. Using pharmacological and molecular tools to modulate OAT1 activity or expression, we causally link OAT1 to persistent inflammatory hypersensitivity in mice. Thus, HDAC4 is a key epigenetic regulator that translates nociceptive activity into sensitization by regulating OAT1, which is a potential target for pain-relieving therapies. Chronic pain is sustained by alterations in gene transcription. Here, the authors show that increased expression of Organic Anionic Transporter 1 in the spinal cord is epigenetically controlled and key to hypersensitivity in pathological pain. Treatments are still, however, far from satisfactory for many patients. Thus, there is an unmet need for the uncovering of novel molecular and cellular players serving as fresh targets for future, more adequate, therapeutic interventions.Chronic pain is a debilitating condition that affects a conspicuous percentage of the worldwide population, resulting in tremendous yearly socio-economic costs3 and shares common molecular mechanisms with other adaptive processes of the central nervous system (CNS) such as memory5. Signal-regulated gene transcription has been identified as one of the key events enabling the maladaptive plasticity typically associated with chronic pain6. In recent years, mounting evidence linked epigenetic\u2013mediated regulation of transcription to different forms of chronic pain11, but a precise picture is still missing.Steps have been made in understanding the signals mediating central sensitization, which is characterized by aberrant pain sensation to both noxious and non-noxious stimuli, but a clear understanding of this complex adaptive process is still lacking. The transition from acute to chronic pain is considered a pathological manifestation of neuronal plasticity in nociceptive pathways12. The several existing HDACs can be categorized into four major classes according to their structural and functional properties. There are three classes of canonical zinc-dependent HDACs where the second class can be further divided into two subclasses based on the functional properties of its members , and one class comprised of the nicotinamide adenine dinucleotide-dependent sirtuins (class III)13. The members of class IIa can shuttle between the cytoplasm and the nucleus in a signal-dependent manner, and their subcellular localization is pivotal in physiological and pathological adaptations17. In particular, in pyramidal neurons, synaptic activity promotes the nuclear export of class IIa HDACs and negatively impacts their nuclear deacetylase activity19. In other types of neuronal cells, this phenomenon is less characterized.Within the heterogeneous group of epigenetic players, DNA methyltransferases and histone deacetylases (HDACs) are prominent mediators of adaptive processes in the CNS owing to their capacity to translate incoming synaptic activity into long-lasting transcriptional responses. HDACs catalyze the removal of acetyl groups from the lysine residues of histone and non-histone proteins and regulate gene transcription by changing chromatin structure and modifying the activity of interacting partners23. Due to a lack of precise pharmacological tools, however, it is not yet possible to draw definitive conclusions regarding the roles of distinct HDACs, and redundancy is a major caveat. Further, there is no information on the regulation in spinal cord circuits of a fundamental aspect modulating the activity of class IIa HDACs, namely their subcellular localization, and how its potential alteration may functionally contribute to chronic pain. Most importantly from a mechanistic viewpoint, the identity of genes underpinning central sensitization and controlled by class IIa HDACs in spinal cord neurons in pathological pain is still elusive.Recent evidence points towards the importance of histone acetylation and HDAC activity in different pathological pain states24, to be an HDAC4 target in the spinal dorsal horn. Further, functional experiments validated spinally-expressed OAT1 as a mediator of nociceptive hypersensitivity and confirmed its targetability for treating inflammatory pain. Thus, OAT1 provides a molecular link between nociceptive activity, epigenetic-regulated transcription, and spinal sensitization, and represents a potential therapeutic target for the pharmacological treatment of persistent inflammatory pain.In this study, we examined a number of HDACs and found the class IIa member HDAC4 to be specifically regulated by synaptic activity and persistent inflammatory pain in neurons of the spinal dorsal horn. We further discovered that activity-dependent changes in the subcellular localization of HDAC4 contribute to mechanical hypersensitivity in mouse models of nociceptive inflammation without affecting either basal or acute sensitivity. Transcriptome analysis revealed organic anion transporter 1 (OAT1), a multi-specific antiporter of anionic substrates and drugs predominantly known for its role in kidney functionsgamma-aminobutyric acid-A-receptor antagonist bicuculline (Bic) to induce bursts of synaptic activity, and consistent with our observations in hippocampal neurons18, we found that members of class I (HDAC1 and 3), class IIb (HDAC10), and class IV (HDAC11) did not change their subcellular distribution  levels whereas total histone 3 levels remained constant , which is frequently used to mimic the time course of chronic inflammatory pain conditions such as rheumatoid arthritis or tendonitis19. Via changes in its localization, HDAC4 influences activity-dependent gene transcription in neurons, thereby playing an essential role in information processing and in different forms of plasticity30. We hypothesized that nociceptive activity-mediated nuclear exclusion of HDAC4 might be instrumental to central sensitization via the modulation of gene transcription in spinal cord neurons. To test this hypothesis, we employed a constitutively nuclear localized dominant-active mutant of HDAC4 (HDAC4 3SA), which features three mutations in phosphorylation sites key for the nuclear export of HDAC4 and is therefore always confined to the nuclear compartment31. Overexpression of \u03b2-galactosidase (LacZ) as a control, wild-type HDAC4 (HDAC4 wt), or HDAC 3SA transgenes in cultured spinal cord neurons had no effect on neuronal viability , a secreted glycoprotein involved in inflammatory responses;40H19, a long non-coding RNA recently linked to the development of neuropathic pain;41 myelin protein zero (Mpz), which has been linked to neuropathy;42 and periaxin (Prx), which is associated with neuropathic pain43. Changes in the expression levels of selected genes were confirmed in an extended cohort of samples using quantitative RT-PCR (QRT-PCR) analysis  of dorsal spinal tissue from mice spinally expressing LacZ, HDAC4 wt, or HDAC4 3SA. 24\u2009h following CFA injection, 104 genes were upregulated and nine genes downregulated in the spinal dorsal horn of LacZ-expressing mice, 47 genes were upregulated and five genes downregulated in HDAC4 wt-expressing mice, and 252 genes were upregulated and six genes downregulated in mice expressing HDAC4 3SA Fig.\u00a0. Comparisis Fig.\u00a0. These dSlc22a6, which codes for the organic anion transporter 1 (OAT1), also known as solute carrier family 22 member 6. OAT1, which is primarily expressed in the kidney, has also been found in the CNS and is known to mediate the transport of diverse low molecular weight endogenous and exogenous substrates, including steroids, hormones, neurotransmitters, numerous drugs, and xenobiotics44. However, a detailed understanding of its function in the nervous system is still lacking. In agreement with our data showing increased mRNA levels of Slc22a6 after intraplantar CFA injection  to achieve knockdown of OAT1 in the dorsal spinal cord . We verified their efficiency for reducing Slc22a6/OAT1 mRNA and protein levels in cultured neurons  designed to target two different regions of the To understand whether, conversely, elevated levels of OAT1 in the spinal dorsal horn could be sufficient to increase sensitivity and exacerbate inflammatory nociceptive sensitivity, we overexpressed OAT1 in the dorsal horn of adult mice via rAAVs (rAAV-OAT1). As a control, we injected rAAV-LacZ. We verified in cultured neurons and in the dorsal horns of injected mice the correct expression of transgenes . PBN is largely used in clinical practice for the treatment of gout and hyperuricemia46. In terms of specificity, the IC50 of PBN for OAT1 is reported to be at least one order of magnitude smaller than for\u00a0other potential targets48. We delivered PBN or vehicle intrathecally to preferentially target OAT1 in the nervous system of inflamed mice. Compared to vehicle-treated inflamed mice, mice treated with PBN displayed significantly reduced CFA-triggered mechanical allodynia up to 24\u2009h after a single intrathecal dose  is, at present, the sole OAT1 inhibitor suitable for clinical use  have shown analgesic properties in models of chronic pain60. Some studies, however, have come to contradictory conclusions based on results in which pain sensitivity was either enhanced following HDACs inhibition or ameliorated by the inhibition of histone acetyl transferases, enzymes that mediate histone acetylation63. Thus, it is not yet clear whether treatments aimed at reducing or increasing histone acetylation are expected to ameliorate or exacerbate chronic pain23. An additional complicating factor in the potential use of HDACi for pain treatment is the fact that the employed HDACi are often far from being specific for one particular HDAC64. Their use thus causes\u2014at best\u2014a broad and non-specific inhibition of all HDACs belonging to one particular class or\u2014most commonly\u2014the inhibition of HDACs belonging to both classes I and II64. In such a scenario, it is virtually impossible to define the contribution of any single HDAC to pain chronicity and/or sensitivity. We took a different approach in this study. Driven by the input-dependent nature of some HDACs, we first characterized the impact of nociceptive activity on HDACs expression and subcellular localization, and identified HDAC4 as distinctively responsive in that it translocates from the nucleus to the cytoplasm in response to synaptic activity in primary spinal cord neurons or subsequent to the induction of peripheral inflammation in vivo. We then proceeded to specifically manipulate the relevant aspect\u2014the subcellular localization\u2014of this particular HDAC and thus verify its functional relevance for persistent nociceptive inflammatory hypersensitivity. Thus, although we cannot exclude the possibility that the activity of other HDACs might contribute to pathological signaling in chronic pain, we believe that nucleocytoplasmic translocation of HDAC4 plays a unique role in persistent inflammatory pain and that the specific modulation of its activity might be beneficial in pain therapy.Reduced acetylation of histone 3 has been previously linked to the nuclear accumulation of HDAC420. Therefore, a multi-faceted scenario involving HDAC4 in both the peripheral and central pain pathways is emerging. In peripheral sensory neurons, HDAC4 seems to promote the aspects of sensitization that are associated with thermal hypersensitivity, while centrally, in spinal cord neurons, its activity prevents development of mechanical hypersensitivity. The reasons for this dualistic behavior are most likely to be found in the genes controlled by HDAC4 that could differ between these compartments. Indeed, future comparative studies are needed to clarify this aspect of HDAC4\u2019s involvement in chronic pain. Notably, the differential effect of a given molecular signal such as HDAC4 on thermal or mechanical hypersensitivity is not an uncommon observation. Indeed, the literature offers plenty of examples showing a differential influence of specific signaling pathways on thermal and mechanical sensitivity in inflammatory pain67, consistent with the idea that thermal and mechanical sensitization are modulated via the recruitment of distinct circuits or molecular signaling cascades.By preventing the nociceptive activity-dependent nuclear exclusion of HDAC4 specifically in spinal cord dorsal horn neurons, we were able to interfere with the development of the mechanical hypersensitivity that accompanies long-lasting inflammation. The development of thermal sensitivity was, however, not affected by this manipulation. Another study showed, by contrast, that the conditional knockout of HDAC4 in primary sensory neurons of mice affects peripheral sensitization and thermal hypersensitivity in the CFA model68. Gene Ontology (GO) analysis of the differentially expressed genes (DEGs) after CFA delivery identified \u201cglycine transport\u201d in addition to the expected terms indicative of inflammatory responses. Surprisingly, the expression of Glyt1 (Slc6a9) or Glyt2 (Slc6a5), which are considered the main effectors in glycine transport, was left unchanged after CFA delivery. However, CFA significantly induced the expression of Slc6a20a, Slc36a2, and Slc38a5, which are listed under the GO term \u201cglycine transport\u201d but whose transport activity of glycine or other amino acids  is not fully characterized23.In our study, we identified an HDAC4-regulated gene transcription program in inflammatory hypersensitivity that included several upregulated genes. This finding might seem surprising, as HDACs have traditionally been associated with the repression of transcription. HDAC4, however, has an extremely heterogeneous network of interacting proteins in both the cytosol and the nucleus, and appears to both induce and repress gene transcriptionSlc22a6). CFA triggered the upregulation of OAT1 in dorsal horn neurons. The identity of the non-neuronal cells expressing OAT1 also at basal levels remains an open question. The mechanisms governing Slc22a6 transcription are largely unknown. Of note, putative binding sites for myocyte enhancer factor-2 (MEF2) have been identified in the promoter region of human SLC22A669 and might explain the role of HDAC4 in the modulation of its transcription. MEF2 negatively regulates gene transcription by recruiting HDAC4 and associated co-repressor complexes while nuclear export of HDAC4 promotes MEF2 interaction with p300, stimulating transcription71.Among the identified genes regulated by HDAC4 we found OAT1 , but also tryptophan metabolites such as kynurenic acid\u00a0(KYNA), and the serotonin metabolite 5-hydroxyindol acetate (5-HIAA)82. One substrate of OAT1 that has particular relevance for CNS function is KYNA, an endogenous antagonist of ionotropic glutamate receptors83. We hypothesize that OAT1 increases pain hypersensitivity via the modulation of inflammatory mediators like PGE2 and/or by promoting the clearance of KYNA from the spinal cord. In line with this thought, PBN, which inhibits the function of OAT1, amongst other targets, induces accumulation of KYNA within the CNS84 and has analgesic properties in inflammatory pain models85. Our data, however, additionally indicate that the peripheral inflammation-triggered increase in OAT1 expression in the dorsal horn activates NF-kB, thus supporting the idea that OAT1 participates in the clearance of inflammatory signaling molecules. This possibility remains, however, an open question and a dynamic field of study to pursue in the future.OAT1 can transport prostaglandins and tryptophan metabolitesTaken together, we show that HDAC4 and OAT1 are key mediators of activity-dependent plasticity in the spinal cord and represent exciting targets for the treatment of inflammatory pain. Given that drugs specifically modulating HDAC4 are under development and especially in light of the fact that OAT1 can be pharmacologically modulated by drugs that are already clinically available, our findings open the way for innovative therapeutic interventions in chronic pain states. One possible delivery strategy for existing OAT1 modulators such as PBN could rely on an intrathecal pain pump to minimize doses and systemic side effects. On the other hand, the development of specific activators of HDAC4 that might prevent the establishment of epigenetically-regulated transcriptional changes in inflammatory pain states and, as such, interfere with the multiple diverse consequences of altered HDAC4 signaling, could possibly provide a more comprehensive and effective therapy.All animal procedures in this study were carried out in accordance with the ARRIVE guidelines and following approval by the local animal welfare committee . Adult male or female C57BL/6N wild-type mice between 8 and 14 weeks of age (Charles River) were used throughout all in vivo experiments. For primary neuronal cultures, P0 C57BL/6N mice were used. Mice were housed in groups of maximally three animals in standard cages (15\u2009cm\u2009\u00d7\u200921\u2009cm\u2009\u00d7\u200913.5\u2009cm) within approved animal facilities at Heidelberg University on a 12:12\u2009h light:dark cycle and maintained at 45\u201365% humidity and 20\u201324\u2009\u00b0C, and with ad libitum access to food and water. Housing conditions were constantly monitored.10. In brief, HEK293 cells  were grown in high-glucose-containing (4.5\u2009g/liter) Dulbecco\u2019s Modified Eagle Medium  supplemented with 10% fetal bovine serum and antibiotics. Before transfection by standard calcium phosphate precipitation, medium was replaced with modified Dulbecco medium  containing 5% fetal bovine serum and no antibiotics. After transfection cells were returned to DMEM. Cells were collected at low speed centrifugation, resuspended in 150\u2009mM NaCl-20\u2009mM Tris-HCl and lysed by 0.5% sodium deoxycholate. rAAVs were purified on heparin affinity columns  and concentrated via Amicon Ultra-4 centrifugal filter devices (Millipore).Recombinant adeno-associated viruses (rAAVs) serotype 1/2 were packaged and purified as described18. Vectors used for shRNA-mediated knockdown contain a U6 promoter driving shRNA expression and an additional expression cassette encoding for GFP under the control of the CBA promoter . The following sequences, obtained from commercial siRNA targeting OAT1 (Qiagen) were used: CCCAGACAATTAAATAAATTT (shOat1-1), ATCCTAGTGAATGGCATAATA (shOat1-2). As a control, a non-targeting shRNA sequence was used (shUNC)10. cDNA encoding for OAT1 was subcloned into a pAAV expression construct under the CMV/CBA promoter followed by an HA-tag at the C-terminal end to achieve overexpression of OAT1 (pAAV-OAT1). The genetically encoded calcium indicator jRGECO1a was subcloned in a pAAV expression construct under the CaMKII\u03b1 promoter and followed by a nuclear localization signal to generate pAAV-jRGECO1a-NLS.pAAV-LacZ, -HDAC4 wt, and -HDAC4 3SA plasmids and their respective rAAVs contain a Flag-tag at their C-terminal end and have been extensively characterized in vitro and in vivo for their specificity and efficacySlc22a6) or negative control siRNA (Qiagen) were intrathecally delivered using the in vivo-jetPEI\u00ae transfection reagent (Polyplus transfection) according to the manufacturer\u2019s instructions and established protocols86. siRNAs were intrathecally delivered once per day for three consecutive days. The following sequences were used: Slc22a6: AAGGAACTGACTCTAAACAAA (SI01420783), TCGGAAGGTGCTGATCTTGAA (SI01420790), ATCCTAGTGAATGGCATAATA (SI01420797), CCCAGACAATTAAATAAATTT (SI01420804), and, as a negative control, AATTCTCCGAACGTGTCACGT (1022076).A cocktail of four small interfering RNAs (siRNAs) targeting OAT1 (87. Mice were anesthetized with a mixture of fentanyl (50\u2009\u00b5g/kg), midazolam (5\u2009mg/kg), and medetomidine (0.5\u2009mg/kg), and laminectomy was performed. Using a microprocessor-controlled minipump and a 35 gauge bevelled \u201cNanoFil\u201d needle (World Precision Instruments), 500\u2009nl of a 2:1 mixture of rAAV stock with 20% mannitol were injected into the parenchyma of the spinal cord dorsal horn of the L3-L5 segments on each side  at a flow rate of 100\u2009nl/min87. Anesthesia was reversed using a mixture of atipamezole (0.75\u2009mg/kg), flumazenil (0.5\u2009mg/kg), and naloxon (1.2\u2009mg/kg), and the animals placed overnight on a heating plate set to 39\u2009\u00b0C.\u00a0Mice were allowed to recover for a minimum of 3 weeks after surgery before further analysis to allow for transgene expression. After experiments, mice were sacrificed, and correct delivery and expression or functionality of the injected rAAVs was assessed.Bilateral injection of rAAVs into the spinal parenchyma of adult mice allows for spatially- and temporally-restricted, stable gene transduction without causing persistent inflammation, tissue injury, or glial scar formation86 and were used to deliver siRNAs (Qiagen) or probenecid (p-(di-n-propylsulfamyl) benzoic acid) (Sigma Aldrich). 50\u2009mg probenecid were dissolved in a total volume of 5\u2009ml solvent consisting of 3790\u2009\u00b5l saline (0.9%), 850\u2009\u00b5l Tris , 150\u2009\u00b5l NaOH (2\u2009M), and 210\u2009\u00b5l HCl (2\u2009M) to yield a 10\u2009\u00b5g/\u00b5l stock solution. 16\u2009\u00b5g of probenecid was delivered to each mouse intrathecally. The amount of probenecid injected\u2014adjusted for the mouse cerebrospinal fluid volume\u2014was determined based on the literature88.Intrathecal injections were performed as previously described10. Acute inflammatory pain was induced by injecting either 20\u2009\u00b5l of 0.03% capsaicin (Tocris) or 1% formalin (Sigma Aldrich) solution in saline into the plantar surface of one hindpaw10. An equivalent volume of 0.9% saline was injected as control. All intraplantar injections were performed under inhalation anesthesia. Paw edema was verified via measurements of the paw width and thickness, obtained using a Micrometer Caliper (Fisher Scientific).For the induction of long-lasting inflammatory pain, 20\u2009\u00b5l of Complete Freund\u2019s Adjuvant (CFA) (Sigma Aldrich) was subcutaneously injected into the plantar surface of one hindpaw89. Briefly, animals were anesthetized with a mixture of fentanyl (0.5\u2009mg/kg), medetomidine (0.5\u2009mg/kg), and midazolam (5.0\u2009mg/kg). Once immobile, an ~1\u2009cm incision was made in the skin overlying the biceps femoris muscle. The muscle was then bluntly dissected to reveal the sural, tibial, and common peroneal branches of the sciatic nerve. The tibial and common peroneal branches were ligated using 5/0 suture, and a 1\u20132\u2009mm section of both nerves peripheral to the ligature removed. The wound was subsequently closed with sutures, and xylocain spray applied as a local anesthetic. Anesthesia was reversed using a mixture of atipamezole (0.75\u2009mg/kg), flumazenil (0.5\u2009mg/kg), and naloxon (1.2\u2009mg/kg), and the animals placed overnight on a heating plate set to 39\u2009\u00b0C. Animals were sacrificed 21\u201326 days following SNI surgery.The spared nerve injury (SNI) model for neuropathic pain involving ligation of the tibial and common peroneal branches of the sciatic nerve has been described in detail previouslyAll behavioral tests were conducted by the same experimenter in awake and unrestrained mice during the light phase. Mice were acclimatized to the testing environment prior to behavioral measurements and on each day of testing. Baseline measures were obtained in separate sessions within one week prior to treatment. Mechanical sensitivity was assessed by measuring responses to paw pressure as previously described by applying a series of von Frey filaments  with ascending forces (0.008\u20132\u2009g) to the plantar surface of the hindpaws. Each filament was applied five times to each paw in increasing order, starting with the filament producing the lowest force. To avoid repeated stimulus-induced sensitization, testing of both paws alternated between mice, and single paws were not re-stimulated before all other mice had been tested. Withdrawal frequency was calculated as a percentage of withdrawals out of the total number of applications for each von Frey filament. Sum of mechanical responses comprehensively represents data on response frequencies to graded 0.008\u20131.0\u2009g von Frey hairs over the entire course of the behavioral experiment. Each filament was presented 5 times and in the case of 5 times withdrawal the percentage of responses to that filament would be 100%. The sum of percentage responses to 0.008\u20131.0\u2009g von Frey forces was then calculated. The response frequency graphs are individual snapshots of sensitivity on the indicated day and depict the distinct percentage responses to the different filaments. Threshold was determined as the minimum von Frey force eliciting 40% of responses.10. Following intraplantar capsaicin or formalin injections, mice were placed in an empty plexiglass chamber and the duration of all nocifensive behaviors, such as licking, shaking or flicking of the injected paw, were determined over a time period of 5\u2009min after injection of capsaicin and over a time period of 60\u2009min, divided into 5\u2009min intervals, after formalin injection37.Thermal sensitivity was assessed according to the Hargreaves method using a plantar test analgesia meter (IITC Life Science)290. Cultured neurons were transfected with 0.3\u2009\u00b5g/ml plasmid encoding for pAAV-LacZ, -HDAC4 wt or HDAC4 3SA on day in vitro (DIV) 8 using Lipofectamine 2000 (Invitrogen) as described91. Cell death rates of cultured spinal cord neurons transfected LacZ, HDAC4 wt or HDAC4 3SA were determined by manually counting the number of collapsed or disintegrated nuclei of transfected neurons and dividing by the total number of transfected cells91. Primary cultured neurons were infected with rAAVs on DIV3 and maintained at 37\u2009\u00b0C and 5% CO2 until analyses on DIV13/14. Cultured spinal cord neurons were treated with 50\u2009\u00b5M bicuculline (Bic) to induce synaptic activity on DIV13/14 for 0.5\u2009h, 2\u2009h, or 8\u2009h. Control cells were treated with vehicle only.Primary spinal cord and hippocampal cultures were plated and cultivated according to established protocols at 37\u2009\u00b0C and 5% CO2, 2.0 CaCl2, 10 HEPES, 1.0 glycine, 35.6 D-glucose, and 0.5 Na-pyruvate. Fluorescence images were acquired from cells bathed in room-temperature HBS at 2.0\u2009Hz with a cooled CCD camera  through a \u00d720 water-immersion objective  on an upright microscope . Fluorescence excitation  was provided by an LED light source (CoolLED pE-4000). jRGECO1a fluorescence was filtered using a 620\u2009\u00b1\u200930\u2009nm emission filter . Data were collected using proprietary software , and analyzed using Fiji and IgorPro (WaveMetrics). Calcium signals in regions of interest drawn around individual nuclei were quantified as the baseline intensity-normalized change in fluorescence over time, \u0394F/F\u2009=\u2009(F\u2212Fbaseline)/Fbaseline. Responses to the GABAA receptor antagonist Bic (50\u2009\u03bcM) were quantified in terms of the peak amplitude of the first transient and the mean inter-peak interval during the first five minutes following the start of Bic stimulation.Nuclear calcium imaging was performed on DIV10 for primary spinal cord neurons infected with rAAV-jRGECO1a-NLS in a HEPES-buffered saline solution (HBS) containing, in mM: 140 NaCl, 2.5 KCl, 1.0 MgCl92. Extracted RNA was reverse transcribed into first strand cDNA and quantitative reverse transcription PCR (QRT-PCR) was performed on a StepOne Plus real-time PCR system using TaqMan\u00ae gene expression assays (Applied Biosystems) for the following genes: Gusb (Mm00446953_m1), Gapdh (Mm99999915_g1), Ptgs2 (Mm00478374_m1), Slc22a6 (Mm00456258_m1), Slc26a7 (Mm00524162_m1), Prx (Mm00479826_m1), H19 (Mm01156721_g1), Msln (Mm00450770_m1), Nov (Mm00456855_m1) and C1qc (Mm00776126_m1). Expression of target genes was normalized against the expression of endogenous Gusb and Gapdh using StepOne software.Total RNA was extracted from the dorsal part of the spinal cord or cultured neurons using an RNeasy Mini Kit (Qiagen), including optional DNase I treatment at room temperature for 15\u2009min according to manufacturer\u2019s instructions95. Mus musculus, mm10/GRCm38, Ensembl reference genome was used for transcript alignment. Genes were annotated using the v85 Ensembl database as reference. The cut-off for determining significantly differentially expressed genes (DEGs) was set to padjusted\u2009<\u20090.05 . Genes significantly regulated by CFA inflammation over the na\u00efve state in saline injected mice, expressing either LacZ, HDAC4 wt or HDAC4 3SA, were considered upregulated with a log2 fold change\u2009>\u20090.5 and downregulated with a log2 fold change\u2009<\u2009\u22120.5. Gene ontology analysis of up- or downregulated DEGs in LacZ-expressing mice subjected to CFA or saline injection was performed using the PANTHER overrepresentation test (released 2020-04-07) and database (released 2020-02-21)96. Fisher\u2019s exact test with false discovery rate (FDR) correction was chosen and the background database was restricted to the pool of genes annotated in our RNA sequencing analysis. Data are available on GEO: GSE159895.Total RNA was isolated from the dorsal spinal cord tissue (L3\u2013L5) of three individual mice per experimental condition. 0.3\u2009\u03bcg of total RNA from each sample was used for RNAseq analysis. Integrity tests of RNA samples and differential gene expression analysis were performed by GATC Biotech . Briefly, a strand-specific cDNA library was generated by purification and fragmentation of poly-A containing mRNA molecules, synthesis of strand-specific cDNA with random primers, adapter ligating on the cDNA and adapter-specific PCR amplification. Sequencing was performed with a Genome Sequencer Illumina HiSeq 4000 using 50\u2009bp single end reads in FastQ format with at least 30 million reads (\u00b13%). Gene expression was analyzed using the Bowtie, TopHat, Cufflinks, Cuffmerge, Cuffdiff software suite2HPO4, 400\u2009mM NaCl). Nuclei were visualized using Hoechst 33258 (2\u00a0\u00b5g/ml\u00a0in 1x PBS). For analysis, up to eight random pictures from each condition were acquired using a Nikon Ni-E upright fluorescence microscope (NIS-Elements Software) or a Leica DM IRBE inverted fluorescence microscope with a \u00d740 objective. Intensity of nuclear fluorescence was measured in ImageJ using the Hoechst channel to define the nuclei as a region of interest in NeuN-positive cells. Measured intensity values were normalized to those obtained from vehicle-treated controls.Cells were fixed for 20\u2009min at room temperature (RT) with 4% paraformaldehyde and 4% sucrose in PBS (pH 7.4). Cells were washed three times with PBS and subsequently incubated for 3\u2009h with primary antibodies , mouse anti-Flag 1:200 (Sigma Aldrich Cat# F3165 clone M2), rabbit anti-HA 1:200 (Santa Cruz Cat# sc-805), rabbit anti-HDAC1 1:200 (Thermo Scientific Cat# PA1-860), rabbit anti-HDAC3 1:200 , rabbit anti-HDAC4 1:200 , rabbit anti-HDAC5 1:200 , rabbit anti-HDAC7 1:200 (Sigma Aldrich Cat# H2662), rabbit anti-HDAC9 1:200 (Abcam Cat# ab18970), rabbit anti-HDAC10 1:200 (Abcam Cat# ab53096), rabbit anti-HDAC11 1:200 (Abcam Cat# ab18973), mouse anti-NeuN 1:500 (Merck Cat# MAB377 clone 147714), rabbit anti-OAT1 1:200 (Abcam Cat# ab135924)) and for 1\u2009h with appropriate secondary antibodies  Alexa Fluor 488 (Life Technologies Cat# A11008); goat-anti-mouse IgG (H\u2009+\u2009L) Alexa Fluor 594 (Life Technologies Cat# A11005)). All antibodies were diluted in GDB ) diluted in blocking solution overnight at 4\u2009\u00b0C, and for 1.5\u2009h at RT with secondary antibodies diluted 1:1000 in blocking solution. Slides were washed and incubated with 0.1% Sudan Black B (Sigma Aldrich) in 70% ethanol for 10\u2009min at RT to reduce autofluorescence. For analysis, up to ten images of the dorsal horn area were acquired per condition using a Leica DM IRBE inverted fluorescence microscope with a \u00d720 objective. Intensity of nuclear fluorescence within neurons of the dorsal horn was measured in ImageJ using the Hoechst and NeuN channel to define the nuclei of neuronal cells as regions of interest. Intensity values were normalized to those from saline-injected controls. High-resolution images were acquired using a confocal laser microscope (Leica).Mice were transcardially perfused with PBS followed by 10% formalin (Sigma Aldrich). Spinal cords and dorsal root ganglia (DRG) were isolated and post-fixed for 2\u2009h in 10% formalin. Cryosections (20\u2009\u00b5m) of the lumbar spinal cord segments L3\u2013L5 and respective DRGs were mounted on Superfrost Plus Adhesion Microscope Slides\u2122 (Thermo Scientific). Sections were blocked and permeabilized in blocking solution containing 10% normal goat serum or normal donkey serum in 0.2% gelatin, 0.6% Triton X-100, 30\u2009mM NaSpinal cords were quickly isolated and fresh tissue was rinsed in cold PBS before the dorsal part of lumbar spinal cord segments L3\u20135 was dissected and homogenized in RIPA buffer ). Primary spinal cord cultures were lysed directly in sample buffer containing 30% glycerol, 4% SDS, 160\u2009mM Tris-HCl, pH 6.8, and 0.02% bromophenol blue. For SDS-PAGE, 25\u2009\u00b5g of spinal cord protein samples or 20\u2009\u00b5l of primary culture lysates were used. Standard protocols for western blot were used. Relative protein content was normalized to loading controls. Images were generated using a Chemidoc Imaging system (Biorad). Antibodies used were: rabbit anti-AcH3 1:2000, rabbit anti-H3 1:4000 (Sigma Aldrich Cat# 06-755), mouse anti-Flag 1:500, rabbit anti-HA 1:1000, rabbit anti-HDAC1 1:2000, rabbit anti-HDAC4 1:6000, rabbit anti-HDAC6 1:1000 (Abcam Cat# ab1440), rabbit anti-HDAC7 1:2000, rabbit anti-HDAC9 1:4000, rabbit anti-OAT1 1:2000, rabbit anti-cFos 1:2000 (Santa Cruz Cat# sc-52), mouse anti-\u03b2-Actin 1:2000 (Santa Cruz Cat# sc-47778 clone C4), mouse anti-tubulin 1:400,000 (Merck Millipore Cat# T9026 clone DM1A), Goat anti-mouse IgG (H\u2009+\u2009L) Peroxidase 1:5000 (AffiniPure Jackson Immuno Research Cat# 115-035-003), Goat anti-rabbit IgG (H\u2009+\u2009L) 1:5000 Peroxidase (AffiniPure Jackson Immuno Research Cat# 111-035-144).Data are presented as mean\u2009\u00b1\u2009SEM and were analyzed using Microsoft Excel, Fiji, ImageJ. Statistical analyses were performed using GraphPad Prism 7 . All data were tested for normality distribution. Details on the statistical tests used are indicated in the respective figure legends. All mice or dishes of cultured neurons were randomly assigned to the different experimental groups. All analyses were performed by a person blind to the condition used.Further information on research design is available in the\u00a0Supplementary InformationReporting summaryPeer Review File"}
+{"text": "In areas where malaria is endemic and where trained microscopists are not available, rapid diagnostic tests (RDTs) are needed not only to allow prompt treatment without delay but also to prevent overdiagnosis and overtreatment based on clinical judgements that may lead to drug resistance. This study aimed to compare the performances of the CareStart Pf/Pan Combo test to field microscopy, which is considered to be the gold standard for malaria diagnosis.Any person with a fever or a history of fever within 48\u00a0h who came to the health centre was recruited for the study and tested both by the CareStart Pf/Pan test and by field microscopy. Sensitivity, specificity, positive predictive value, negative predictive value, and accuracy were analysed with both methods.Two-hundred study participants were enrolled: 96 (48%) were found to be positive through microscopy, while 100 (50%) participants were found to be positive through RDT. The RDT produced four false-positive results. High sensitivity and specificity were observed for the CareStart Pf/Pan test . The CareStart Pf/Pan test also showed excellent agreement with the field microscopy results.The Carestart Pf/Pan could be used as an alternative diagnostic test in malaria-endemic areas where facility for performing microscopy is not available. Malaria remains a leading cause of morbidity and mortality worldwide, with an estimated 229\u00a0million new cases and 409,000 deaths annually . As in oPlasmodium falciparum than for Plasmodium vivax compared to microscopy [However, microscopy is time consuming and requires trained personnel, careful preparations and a constant supply of power for operating equipment, which is limited in most areas where malaria is endemic . RDTs cocroscopy , 13.P. vivax, pan-lactate dehydrogenase (pLDH) tests showed sensitivities ranging from 62.0 to 95.0% [For to 95.0% \u201314. Polyto 95.0% .P. vivax compared to field microscopy, which is the gold standard, in the Tanjung Leidong subdistrict, North Sumatra, Indonesia. This strategy is in line with the recent Malaria National Strategic Plan for 2020\u20132024 for Indonesia that recommends regular evaluations with RDTs within the country [P. vivax.This study was conducted to evaluate the performance of the CareStart RDT Pf/Pan combo test for the detection of  country . The hyp, 83 blood smear-positive and 83 blood smear-negative samples were required. The detection of malaria-carrying species was limited to P. vivax.To detect In inclusion criteria, any person with a fever or a history of fever within 48\u00a0h who visited the health centre was selected as a study participant. As for exclusion criteria, individuals who took anti-malarial drugs within the four weeks prior to the study or refused to participate were excluded. Both CareStart Pf/Pan combo tests and thick and thin blood smear microscopic examinations were simultaneously performed on samples obtained from each patient with acute febrile illness.The diagnostic performance of the CareStart Pf/Pan combo test targeting the pLDH antigen was evaluated against the performance of thick blood-smear microscopy. Microscopy is still the gold standard for diagnosing malaria, thus under field condition it may miss some symptomatic cases, however it is still suitable for clinical diagnosis . The heaThe CareStart tests used in this study were donated by the Ministry of Health. Test kits were stored at room temperature and the expiration date was checked before use. The tests were performed according to the directions of the manufacturer. The CareStart tests were labelled with a patient identification number and date, and the results were recorded after 20\u00a0min. The presence of a unique pan-pLDH line is found when an individual is infected with one or more non-falciparum species. A test result without a control line was considered invalid, and the sample was retested. All the data were double-entered by study staff to maintain accuracy.Data were entered and analysed using STATA version 15.1  after checking for completeness. Data were analysed for descriptive statistics, while results of both methods were compared using Cohen\u2019s kappa co-efficient. The sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) were calculated. All statistical analyses were done at 95% level of significance.P. vivax infection, while 100 (50%) tested positive with the CareStart Pf/Pan test. The only species detected in the positive blood smears and RDTs was P. vivax. The baseline characteristics are shown in Table During the study period, 207 patients with fever were screened and tested for malaria with the CareStart Pf/Pan test and field microscopy, among which 200 patients agreed to participate. The other seven patients refused to participate. More than one-half of the study participants were male (55.5%). By field microscopy, 96 (48%) study participants were found to be positive for P. vivax infection, the sensitivity and specificity of the CareStart Pf/Pan combo test were 100% (95% CI 96.23\u2013100) and 96.15% (95% CI 90.18\u201398.43), respectively. There were four (2%) discordant results obtained between field microscopy and the CareStart Pf/Pan test. All the discordant results were negative in the field microscopy test and positive in the CareStart Pf/Pan test. Unfortunately, the discordant samples were not tested to PCR as a confirmatory test that would help improve the reliability of the test.Using microscopy as the gold standard test for detecting malaria by The PPV and the NPV of the CareStart Pf/Pan test were determined to be 98% (95% CI 94.96\u201399.45) and 100%, respectively. The agreement of the results between the CareStart Pf/Pan tests and the field microscopy tests were excellent, with a kappa value of 0.96 Table . The resThe CareStart Pf/Pan test revealed an increasing number of positive results with increasing levels of parasitaemia, with parasitaemia \u2009>\u20091000/\u00b5l, accounting for 91.67% of all positive RDT results Fig.\u00a0.Fig. 2NuP. vivax. This finding was similar to the study performed at the same site in 2013 [Plasmodium falciparum was the only observed parasite [RDTs for malaria diagnosis are vital to make prompt treatment decisions, especially at healthcare facilities that do not have trained microscopists , 19. Eve in 2013  but diffparasite . The majparasite , 24.A study performed in Korea found that one patient tested negative by RDT (SD BIOLINE), while the microscopy result was positive, which was a result similar to that of a study performed in Ethiopia with the CareStart combo test , 26. TheInterestingly, the high sensitivity of the CareStart Pf/Pan test in this study was different to that in a study from Sumba, Indonesia, where the sensitivity of the test was only 13.6% . The higLimitations of this study included the cross-sectional design that limited the detection of sub-microscopic parasites that may have been found in a follow-up test. In the present study, only the CareStart Pf/Pan test results were compared with field microscopy results; however, the true burden of sub-microscopic infection would have been better determined by polymerase chain reaction (PCR) and new diagnostic tests should have the ability to detect sub-microscopic infections to achieve malaria elimination .Nevertheless, the comparison of RDT and microscopy results for malaria diagnosis is seldom conducted in Indonesia, and the study results are important because they are in line with the current Malaria National Strategic Plan for 2020\u20132024.The CareStart Pf/Pan test showed very good sensitivity and specificity with excellent agreement with the results obtained with use of the gold standard, field microscopy. This result provides important information to the Ministry of Health."}
+{"text": "Alzheimer's disease (AD) is considered to be the most common neurodegenerative disease and also one of the major fatal diseases affecting the elderly, thus bringing a huge burden to society. Therefore, identifying AD-related hub genes is extremely important for developing novel strategies against AD.  Here, we extracted the gene expression profile GSE63061 from the National Center for Biotechnology Information (NCBI) GEO database. Once the unverified gene chip was removed, we standardized the microarray data after quality control. We utilized the Limma software package to screen the differentially expressed genes (DEGs). We conducted Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses of DEGs. Subsequently, we constructed a protein-protein interaction (PPI) network using the STRING database.  We screened 2169 DEGs, comprising 1313 DEGs with upregulation and 856 DEGs with downregulation. Functional enrichment analysis showed that the response of immune, the degranulation of neutrophils, lysosome, and the differentiation of osteoclast were greatly enriched in DEGs with upregulation; peptide biosynthetic process, translation, ribosome, and oxidative phosphorylation were dramatically enriched in DEGs with downregulation. 379 nodes and 1149 PPI edges were demonstrated in the PPI network constructed by upregulated DEGs; 202 nodes and 1963 PPI edges were shown in the PPI network constructed by downregulated DEGs. Four hub genes, including GAPDH, RHOA, RPS29, and RPS27A, were identified to be the newly produced candidates involved in AD pathology.  GAPDH, RHOA, RPS29, and RPS27A are expected to be key candidates for AD progression. The results of this study can provide comprehensive insight into understanding AD's pathogenesis and potential new therapeutic targets. Alzheimer's disease (AD) is typical hippocampal amnesia and cognitive disorder . It is cAs known to all, a variety of complex pathogenic factors, such as genetic and environmental factors, lead to the occurrence of AD , 9. Up tGene chip technology is a toolset that arranges a large number of nucleic acid molecules in a large-scale array on a small carrier and detects the strength of the hybridization signal by hybridizing with a labeled sample and then determines the number of detected molecules in the sample . It has This article is dedicated to screening and identifying the differentially expressed genes (DEGs) in the gene expression profile GSE63061 and DEGs related to AD. We performed function and pathway enrichment analysis of the DEG and subsequently constructed the protein-protein interaction (PPI) network. Finally, we obtained several genes related to AD: GAPDH, RHOA, RPS29, and RPS27A.http://www.ncbi.nlm.nih.gov/gds/) [The gene expression profile GSE63061 on Illumina HumanHT-12 V4.0 expression beadchip was acquired from the Gene Expression Omnibus (GEO) of NCBI (ov/gds/) . A totalP value <0.01. For analyzing the DEGs in-depth, we constructed a heat map utilizing the Pheatmap package (https://cran.r-project.org/package=pheatmap) in R.We utilized the Limma package in R to identify the DEGs, with an adjusted https://david.ncifcrf.gov/tools.jsp) [P < 0.05, the result is considered statistically significant.We performed the enrichment analysis of AD-associated genes by Gene Ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses utilizing the Database for Annotation, Visualization and Integrated Discovery  . GO termhttp://string-db.org) [We employed the STRING database (-db.org) , which wP values are RPL36AL, LOC100132795, NDUFA2, and so on. Then, we utilized a heat map of identified DEG in the GSE63061 database to conduct cluster analysis  was used for our research. We firstly analyzed the DEGs between AD samples and age-matched normal samples. We obtained 2169 genes. The genes with the most significant analysis . Genes wTo obtain the function and pathway of these DEGs, we used the online tool DAVID to analyze 1313 upregulation DEGs and 856 downregulation DEGs. Functional enrichment analysis showed that these DEGs with upregulation exhibited a significant association with the immune response of activated neutrophil involvement, the degranulation of neutrophils, and the immunity mediated by neutrophils . LysosomWe carried out a PPI network analysis to explore the interaction and hub genes of DEGs. Except for disconnected nodes in the network, 379 nodes (proteins) and 1149 PPI edges (interactions) were demonstrated in the PPI network constructed by upregulated DEGs . LikewisAs an intricate and refractory neurodegenerative disease, AD seriously affects people's life and living quality, particularly for the elderly , 20. ItsWe used public databases to identify and screen DEGs and related pathways in AD through various bioinformatics methods. We identified GAPDH, RHOA, RPS29, and RPS27A as the hub proteins and key genes of AD. Previous studies have shown that GAPDH is a regulator of cell death, and GAPDH is involved in tumor progression and has become a new therapeutic target. Research by Hwang et al. showed that GAPDH-mediated mitosis eliminated defective mitochondria and led to apoptosis, which can provide a potential treatment method for the treatment of Huntington's disease and other neurodegenerative diseases . ResearcFunctional enrichment analysis revealed that DEGs with upregulation primarily took part in and were enriched in the immune response, neutrophil degranulation, lysosome, osteoclast differentiation, and so on. Mishra and Brinton's research has pointed out that the inflammatory immune response is the unifying factor that connects each risk factor of AD . The DEGThis study has some limitations. First, the key DEGs need to be verified by RT-qPCR. In future studies, we will collect clinical samples to verify the expression levels of key DEGs through RT-qPCR. Secondly, we plan to further explore the mechanism of key genes in AD in the animal model.In summary, we identified 2169 DEGs between AD patients and healthy controls. Functional enrichment analysis demonstrated that DEGs with upregulation displayed a significant association with immune response, neutrophil degranulation, lysosome, and osteoclast differentiation; the DEGs with downregulation exhibited a significant association with peptide biosynthetic process, translation, ribosome, and oxidative phosphorylation. Subsequently, we identified GAPDH, RHOA, RPS29, and RPS27A as key genes in AD by analysis of the PPI network. The purpose of this research is to improve our understanding of the molecular mechanism of AD through comprehensive bioinformatics analysis and may give a hint of developing the treatment of AD patients."}
+{"text": "OsROS1a, OsROS1a gene editing mutants were generated using the CRISPR/Cas9 system. The osros1a mutants had shrink spikelets, smaller anthers and pollen grains, and were not stained by iodine staining showing a significant reduction in total soluble sugar and starch contents as compared to wildtype (WT), which caused complete male sterility. Similarly, the expression of genes involved in pollen and anther development was decreased in osros1a mutants as compared to WT. Furthermore, bisulfite sequencing showed that the CG and CHG methylation of the OsPKS2 gene promoter was significantly increased in the osros1a mutant, which caused a reduced expression of OsPKS2 in osros1a mutants. DNA methylation of the TDR gene promoter was similar between WT and osros1a mutants, indicating that the DNA methylation effect by OsROS1a was gene specific. The expression of OsROS1a in the mutants was not changed, but it produced a frame-shift mutation to truncate the Pem-CXXC and RRMF domains. Combined with previous studies, our findings suggested that the RRMF domain in OsROS1a is the functional domain and loss of RRMF for OsROS1a causes sterility in rice.For crop seed production, the development of anthers and male fertility are the main agronomic traits and key biological processes for flowering plants. Active DNA demethylation regulates many plant developmental processes and is ensured by 5-meC DNA glycosylase enzymes. To find out the role of  Arabidopsis. DNA demethylation mediated by DME is important for the reproduction of plants, and the inheritance of loss-of-function paternal or maternal mutant dme alleles resulting in a reduced sperm transmission or seed abortion, respectively [ROS1 causes DNA hypermethylation of CG and CHH and decreases the related gene expression in Arabidopsis [ROS1 paralog genes  that mediate DNA demethylation [OsROS1a is highly expressed in rice anthers and pistils [OsROS1a mutants exhibited severe defects in both male and female gametogenesis to produce a sterile phenotype [OsROS1a and is vital for viable seed production, and sperm non-CG methylation is indirectly promoted by DNA methylation in the vegetative cell, which suggests that dynamic DNA methylation reprogramming occurs during plant embryogenesis [OsROS1 generates a new transcript mOsROS1 with a 21-nt insertion, which can lead to DNA hypermethylation and suppresses the expression of two transcription factors, RISBZ1 and RPBF, increasing the number of aleurone cell layers [OsROS1b causes DNA hypermethylation of Tos17 retrotransposons in rice, and overexpression of OsROS1b extensively reduces DNA methylation of the rice genome [ROS1 is involved in the seed development of rice, wheat and barley by its epigenetic influence on the accumulation of seed storage proteins (SSPs) [DNA methylation plays a crucial role in plant growth, development, and stress responses . In planectively ,7. preseectively . The losbidopsis ,4. The rhylation . OsROS1a pistils . The dishenotype ,11. The ogenesis . The alel layers ,14. Knoce genome . Furthers (SSPs) ,16.OsPKS2 is involved in the normal development of pollen wall formation and mutation in the OsPKS2 caused male sterility in rice [TDR) has an essential role in regulating the transcriptional network for the development and degradation of tapetum, and mutation in this gene caused male sterility [MEL1) gene has a crucial role in microsporogenesis, abnormal accumulation of MEL1 can make a semi-sterile phenotype in rice [CYP704B2 is specifically expressed in the tapetum and microspores, which is required for the synthesis of cutin monomers, and its mutant cyp704B2 shows a male sterile phenotype with a swollen sporophytic tapetal layer and aborted pollen grains [Pollen fertility is important for successful seed production in flowering plants. However, defective development of the anthers can lead to either an absence or formation of non-functional pollen grains . Success in rice . The ricterility ,25. MEIO in rice . Cytochrn grains . Pollen OsROS1a regulates pollen fertility. The CRISPR/Cas9 system was used to generate OsROS1a gene editing mutants, and the osros1a S6, S7, and S16 mutants exhibited complete male sterile, which resulted from the disrupted Pem-CXXC and RNA recognition motif fold (RRMF) domains. The qRT-PCR results showed that the expression of genes involved in pollen and anther development, and starch biosynthesis decreased in these mutants as compared to WT. Furthermore, bisulfite sequencing showed that the CG and CHG methylation of the OsPKS2 gene promoter was significantly increased in the osros1a mutants. These findings could provide new insights to reveal the DNA demethylation of OsROS1a on sterility in rice.This study aimed to reveal the underlying mechanism of how OsROS1a contains 17 exons that encode a protein having 1952 amino acids (aa), and 3\u2032-and 5\u2032-UTRs of 607 and 73 bp, respectively  of 159 genes was then used for constructing the phylogenetic tree with the neighbor-joining (NJ) method (ROS1 (mROS1), dicots DME (dDME), dicots ROS1 (dROS1) and dicots DML2 (dDML2)  method . The phy (dDML2) . AlignmeThe sequence logos of the identified DNA glycosylase domain in all 52 species were generated using the WebLogo program to further verify the conservation of aa residues A. A totaOsROS1a exhibited severe defects in both male and female gametogenesis to produce a sterile phenotype [OsROS1a in active DNA demethylation on pollen and seed development, the stable OsROS1a gene editing mutants were created using the CRISPR/Cas9 system, and three stable mutants  were obtained. The S6 and S16 mutants are homozygous mutations on both target sides, while S16 had a homozygous mutation in the first target and a biallelic mutation in the second target  decreased in both mutants, respectively, compared to WT controls  can block the catalytic activity of DNMT1 specifically on non-methylated DNA [OsROS1a knock-in mutant disrupted the whole gene affecting both male and female gametophytes [OsROS1a knock-out mutants using CRISPR/Cas9 induced pollen and embryo sac defects in rice [OsROS1a gene affects the sterility phenotype in rice through its DNA demethylation function.The expression of the controls , and 180controls . The whoated DNA . Furthertophytes , while t in rice . Recentl in rice . All theOsROS1a was obtained from Phytozome (https://phytozome-next.jgi.doe.gov/) (accessed on 19 October 2018). The conserved domain sequence position of the OsROS1a protein was searched using the Inter Pro software. The protein sequences of OsROS1a were used to identify the homologous gene copies in 52 plant species by BLAST search. The protein sequences of all identified OsROS1a homologous genes were aligned via Muscle by the default settings with manual alterations. The phylogenetic analysis was done by the neighbor-joining (NJ) method using MAGA-X with the default parameters, and the bootstrap test with 1000 replicates was made to determine the confidence of the evolutionary tree [http://weblogo.berkeley.edu/) (accessed on 15 January 2019).The gene structure (Exon-intron distribution) and protein (CDS) sequence of the DNA demethylase gene ary tree . The conosros1a mutants [OsROS1a gene by using the web tool (http://cbi.hzau.edu.cn/crispr/) (accessed on 19 December 2016), which were ligated with rice OsU6a and OsU6b small nuclear RNA promoters, respectively. The sgRNAs were cloned into a binary vector that contained the sgRNA and Cas9 expression cassettes. Then the binary vector was transferred into rice (Nipponbare) calluses by following the method of Agrobacterium-mediated transformation of Japonica Rice.The CRISPR/Cas9 system was used for generating  mutants . Two sinhttp://skl.scau.edu.cn/dsdecode/) (accessed on 12 September 2017), a web-based tool, was used to further analyze the genotypes of the targeted mutations.Genomic DNA was isolated from the leaves of the rice transgenic plants to detect the mutation types. The Cas9-specific primers were used to identify the successful transgenic plants by PCR and osros1a mutants were sampled from the spikelets just before flowering and a 1% potassium iodide (I2-KI) solution was used to stain the pollen grains. A Nikon eclipse Ni fluorescence microscope was used to visualize and photograph the stained pollen grains. For transverse section analysis, spikelets of various anther development stages according to the previous study [Floral organs were photographed with a dissecting microscope. The anthers from the wild type (WT) and us study  were colosros1a mutants were weighed and ground in 5 mL of 80% ethanol according to the Anthrone method [The 0.05 g anthers from WT and e method . The mixosros1a mutants using TRIzol\u00ae Reagent . RT reagent Kit PrimeScriptTM having gDNA Eraser  was used to eliminate genomic DNA residues, then cDNA was synthesized. For RT-PCR reactions, the first strand of cDNA was used with gene-specific primers and the OsACTIN gene was used as an internal control  (http://katahdin.mssm.edu/kismeth/revpage.pl) (accessed on 21 October 2019) was used for DNA methylation analysis.For bisulfite sequencing of osystem) . DNA sam"}
+{"text": "The pressure relief door (PRD) is a vital structure to ensure the safety and reliability of the engine. This paper established a zero-dimensional transient simulation mathematical model to study the plenum compartment pressure threshold and maximum opening angle effects on the nacelle pressure relief process under different opening modes. Then, a computational fluid dynamics model verified by experimental literature data was used to simulate the nacelle pressure relief process and to determine the influence of two different opening modes on the force and discharge characteristics of the PRD. The results of this study show that different opening modes strongly impact the nacelle pressure relief process. Reducing the nacelle compartment pressure threshold of the PRD opening can reduce the time required for the pressure relief process to reach the equilibrium stage. Reducing the maximum opening angle may increase the nacelle compartment pressure during the equilibrium stage. In addition, under the same nacelle compartment pressure thresholds and maximum opening angles, the pressure relief process under the vertical opening mode can reach a lower nacelle compartment pressure during the pressure relief equilibrium stage compared to that under the horizontal opening mode. Therefore, the vertical opening mode is better than the horizontal opening mode. This paper provides two lower calculation costs and high accuracy research models for studying the nacelle pressure relief process. Therefore, a pressure relief door (PRD) must be installed on a nacelle that opens after the internal pressure of the nacelle increases to a certain threshold, which will maintain the internal pressure in the core casing of a high-bypass turbofan engine under a safe level to avoid structural damage or failure of the nacelle structure. In recent years, civil aviation engines have developed in the direction of a high bypass ratio and overall pressure ratio, and the fan pressure ratio has gradually decreased3. Compared with aircraft from the 1950s, the operating pressure and temperature of modern aircraft engines are higher, which has led to a significant increase in emissions from the nacelle pressure relief system: after the engine bleed air duct is ruptured, the nacelle structure will bear a higher pressure load, which in turn, will result in higher requirements for the discharge performance of the PRD5. Therefore, it is essential to design a reasonable structure for a pressure relief valve and understand its operating performance and control method.The nacelle of an aircraft contains engine accessories, such as the engine, casing, booster pump, and various sensors. The pressure change in the engine nacelle will significantly impact the working conditions of susceptible electronic components, fuel lines, circuits, etc. During the flight of an aircraft, if the engine's air intake pipe is damaged so that a large amount of high temperature and high-pressure gas leaks and the leaked gas cannot be discharged from the nacelle in time, the internal pressure of the nacelle will rapidly increase. Excessive force can damage the nacelle structure and even the engine. According to the US Federal Aviation Regulation (FAR) Part 25.1103 (d)The pressure relief characteristics of a nacelle PRD should be understood to determine the maximum pressure in the nacelle of an engine following high-pressure gas leaks. During the operation of an aircraft nacelle, the outflow Mach number and pressure ratio affect the pressure difference. The discharge characteristics between the PRD and the plenum, the shape, opening method, opening angle, and aspect ratio of a PRD influence the gas flow state and force characteristics of the PRD during the nacelle pressure relief process. Therefore, the influence of the above factors needs to be clarified when studying the pressure relief characteristics of the PRD.7 experimentally measured the discharge characteristics of an auxiliary exhaust port with a particular inclination. The results show that when the discharge flow rate is low, the inclined outlet has a higher emission coefficient than the outlet perpendicular to the airflow. In addition, when the discharge flow coefficient is constant, the outflow Mach number has little effect on the flow coefficient of an outlet with a sloped or curved pipe. Vick et al.8conducted an experimental study on the discharge and force characteristics of the auxiliary air outlet of a curved duct with flap discharge in transonic airflow. However, Vick's work mainly focused on the additional ventilation flow, so it was limited to pressure ratios below 1. Pratt et al.10, and Yu et al.11 used the CFD method to numerically study the discharge and force characteristics of a PRD during pressure release from a nacelle and studied the nacelle pressure relief process under different Mach numbers, pressure ratios, and opening angles. This research found that there is an optimal opening angle value and that the discharge coefficient for the PRD reaches a maximum under this opening angle value. Benard et al.12 experimentally investigated the pressure relief process for a PRD with a pressure ratio greater than 1. The results showed that the discharge flow ratio decreases with an increasing Mach number at a given pressure ratio. Vedeshkin et al.13 performed an experimental study of the discharge and force characteristics of a nacelle PRD and compared the experimental results with numerical simulation results. The results showed a good agreement between the numerical simulation results and the experiment. However, the PRD opening mode in this study is unlike that found in previous research, in which the PRD hinge is oriented parallel to the freestream direction. Schott et al.14 studied the effects of the PRD aspect ratio and conducted comprehensive numerical simulations with a series of pressure ratios, Mach numbers, internal temperatures, and other conditions. The results show that the angle of the equilibrium torque increases with increasing pressure in the high-pressure chamber and decreases with an increasing Mach number. The discharge coefficient rises with a rising opening angle and does not increase after reaching a maximum value. In addition, many engineers have proposed various structures for a nacelle PRD and given some suggestions for reference20.In recent decades, there have been few public research reports of experimental and numerical simulations of the pressure relief process of a nacelle. Dewey et al.22. Therefore, a new calculation method is proposed in this paper. First, steady-state CFD analysis is used at a fixed opening angle to obtain the relationship between the discharge coefficient CD and the moment M under the plenum compartment pressure at a certain freestream Mach number and aspect ratio. Then, the opening angle is changed to obtain the relationship between the discharge coefficient CD and the moment M with the opening angle of the PRD. On this basis, a zero-dimensional transient simulation mathematical model of the nacelle pressure relief process is established. The CD and M values obtained are substituted into the model for calculation. This method dramatically reduces the calculation cost while ensuring accuracy, so one can better meet actual engineering needs. In addition, there is still a lack of relatively simple simulation models for practical engineering application prediction.The nacelle pressure relief process is well known to involve complex transient flow behavior. However, the pressure relief door in many of the above studies is stationary, which cannot reflect the relationship between the plenum compartment pressure and the opening angle of the PRD overtime during the actual pressure relief processMost existing studies are based on the PRD opening mode being a vertical opening mode in which the PRD hinge is oriented perpendicular to the freestream direction, as shown in Fig.\u00a0The pressure in the high-pressure bleed air duct and the external ambient pressure and temperature of the nacelle remain unchanged during the entire pressure relief process.Air is considered an ideal gas, and heat transfer is not considered during the pressure relief process. Therefore, it is an adiabatic process.It is considered that the thermal parameters, such as temperature, pressure, and density, in the control volume of the nacelle are uniform.The internal gas discharge during the pressure relief process is discharged through the PRD, i.e., there are no other gas leaks.The PRD is sufficiently rigid, so there is no need to consider the effect of its deformation on the pressure relief process.When the engine bleed air duct is broken and has a severe leak, the leaked gas will flow into the nacelle, increasing the internal pressure of the nacelle above the threshold. Then, the PRD will open, and the pressure relief process will occur under the collective effect of the plenum compartment discharge gas and external free stream. In addition, the pressure relief process will also be affected by factors such as the PRD opening mode, aspect ratio, and opening angle. Therefore, the pressure relief process is a complex flow process with a multi-factor coupling, and the flow diagram of the pressure relief process is shown in Fig.\u00a0p1/p0\u2009>\u2009\u03c0p,cr, the flow is in a noncritical state, and the leakage mass flow rate for the high-pressure bleed air duct is given byWhen p1/p0\u2009\u2264\u2009\u03c0p,cr, the flow reaches a critical state, and the leakage mass flow rate for the high-pressure bleed air duct is given by\u03bc is the flow coefficient of the high-pressure bleed air duct leakage. A0 is the leakage area. R is the specific gas constant ). p0 is the pressure in the high-pressure bleed air duct. T0 is the temperature of the high-pressure bleed air duct. And p1 is the nacelle interior pressure.When p2/p1\u2009>\u2009\u03c0p,cr, the flow is in a noncritical state, and the discharge mass flow rate for the PRD is given byWhen p2/p1\u2009\u2264\u2009\u03c0p,cr, the flow reaches a critical state, and the discharge mass flow rate for the PRD is given byCD is the discharge coefficient of the PRD under different plenum compartment pressures and opening angles, which is obtained from CFD steady-state simulation calculation. Adoor is the area of the PRD. \u03c6 is the opening angle of the PRD. T1 is the internal nacelle temperature. And p2 is the freestream static pressure of the external environment.When m1 is the gas mass inside the nacelle. m1, in is the gas mass flowing in the nacelle. And m1, out is the gas mass flowing out of the nacelle.The mass of the gas inside the nacelle varies over time as follows:p/\u03c1\u03b3\u2009=\u2009C and ideal gas equation pV\u2009=\u2009mRT, the derivative of time can obtainV is the internal volume of the nacelle.From the simultaneous adiabatic flow equation The nacelle's internal temperature varies over time as\u03b1 is\u03c9 is the rotational angular velocity. M is the total moment of the PRD, which is obtained from CFD steady-state simulation calculation. J is the moment of inertia of the PRD rotating around the hinge, and Mf is the resistance moment when the PRD is turned, i.e.,c is the rotation resistance coefficient of the PRD. \u03c11 is the density of the discharge gas. l is the PRD chord length. And b is the PRD width.When the PRD rotates, the rotation angle acceleration 2 and aspect ratio of 1. The maximum opening angle of the PRD is set to 50\u00b0, and the nacelle compartment pressure threshold of the PRD opening is 0.16\u00a0MPa and 0.14\u00a0MPa. The calculated variations in the plenum compartment pressure and PRD opening angle with time under the different opening modes are shown in the Figs.\u00a0The calculation conditions are based on a Mach number of 0.7, pressure in the high-pressure bleed air duct of 0.4\u00a0MPa, initial pressure in the nacelle of 0.1\u00a0MPa, PRD area of 0.0625 mCD of the PRD under the vertical opening mode is higher than that under the horizontal opening mode. However, due to the reciprocating swing in the vertical opening mode, the nacelle compartment pressure will slightly fluctuate during the pressure relief equilibrium stage. In the horizontal opening mode, the nacelle compartment pressure is stable at a specific value because the PRD is stabilized at the maximum opening angle.As seen in Fig.\u00a0In addition, as can be seen from Fig.\u00a02, and aspect ratio of 1. The nacelle compartment pressure threshold of the PRD opening was set to 0.14\u00a0MPa, and the maximum opening angle of the PRD was 50\u00b0, 40\u00b0 or 30\u00b0. The calculated variations of the plenum compartment pressure and the opening angle of the PRD with time under the different opening modes are shown in the Figs.\u00a0The calculation conditions are based on a Mach number of 0.7, pressure in the high-pressure bleed air duct of 0.4\u00a0MPa, initial pressure in the nacelle of 0.1\u00a0MPa, PRD area of 0.0625 mAs seen from Figs.\u00a0In the horizontal opening mode, since the moment acting on the PRD is always positive, no matter what the maximum opening angle is set to, the PRD will be fixed at the maximum opening angle during the pressure relief equilibrium stage. However, in the vertical opening mode, as the maximum opening angle decreases, the reciprocating swing angle of the PRD during the pressure relief equilibrium stage decreases.The physical model established in this paper is shown in Fig.\u00a07, 5.0\u2009\u00d7\u2009107, and 14.3\u2009\u00d7\u2009107 were calculated under the vertical opening mode to validate the independence of the mesh, and the calculation results are shown in Table 7, the error in the simulation results compared with a larger mesh number is minimal. Therefore, the mesh number and the simulation results can be considered to have reached independence.Since the computational physical model is a symmetrical structure for the vertical opening mode, the computational domain is symmetric. Only one-half was computed, significantly reducing the computational cost while satisfying the calculation. A high-quality structured mesh was obtained for the calculation by ICEM CFD due to the consideration that the boundary layer mesh of the PRD surface needs to be encrypted and that the structure of the flow outlet region is complicated, with the overall mesh shown in Fig. 2 and an aspect ratio of 1 to obtain the relationship between the discharge coefficient CD and the moment M with the opening angle of the PRD. The PRD aspect ratio is defined as the ratio of the door span to the door chord length. There are two ways to open the PRD: the vertical opening mode and the horizontal opening mode. The external freestream Mach number is 0.7, and the plenum compartment pressures p1 are 0.1\u00a0MPa, 0.12\u00a0MPa, 0.14\u00a0MPa, and 0.16\u00a0MPa. Then, cases for PRD opening angles of 10\u00b0, 20\u00b0, 30\u00b0, 40\u00b0, and 50\u00b0 are numerically calculated.A computational fluid dynamics calculation is carried out based on a PRD area of 0.0625 m23, and a second-order upwind discrete scheme was adopted. The realizable k-\u03b5 turbulence model was used because it has a known accuracy when dealing with flows involving jets, separations, and secondary flows. Standard wall functions were used to simulate the influence of the wall, and solid walls were assumed to have no speed slip and mass penetration24. The commercial ANSYS software package Fluent 19.2 was to solve the calculations.A pressure-based coupled solver was used to solve the compressible Reynolds-averaged Navier\u2013Stokes equations (RANS)The plenum compartment and the freestream flow had a uniform total temperature and total pressure distribution. It was assumed that the plenum compartment flow remained static, so the total pressure and static pressure were equal, similar to that found for the high-pressure region after a burst duct event. The outflow boundary condition was set as a uniform static pressure. The freestream's static pressure at the basin's inlet was adjusted to provide the required Mach number, and the total pressure of the plenum compartment was adjusted to study the force and discharge characteristics under different plenum compartments pressures.CD: The discharge coefficient was calculated based on isentropic relationships assuming an ideal gas. This is an important parameter used to measure the effectiveness of the PRD discharge characteristic at a particular opening angle.Discharge coefficient p1 is the plenum compartment pressure, and p2 is the freestream static pressure.The pressure ratio was calculated using\u03b3 is the ratio of the specific heat. For air, \u03b3\u2009=\u20091.4.The critical pressure ratio is calculated using\u03c0p\u2009>\u2009\u03c0p,cr, the discharge coefficient is calculated usingFor a pressure ratio \u03c0p\u2009\u2264\u2009\u03c0p,cr, the discharge coefficient is calculated as:m is the actual mass flow rate through the outlet, Adoor is the area of the PRD, \u03c6 is the opening angle of the PRD, R is the specific gas constant ), and T1 is the plenum compartment temperature.When the pressure ratio 8 was obtained by numerical simulation using the above method. The mesh is shown in Fig.\u00a0DFR) was extracted from the numerical calculations for a single angle and plotted in Fig.\u00a010. DFR is defined as:m is the mass flow rate through the plenum compartment outlet, U2 is the freestream velocity, and \u03c12 is the freestream density. A is the effective outlet area, defined as the minimum cross-sectional area between the outlet wall and the door.The existing experimental data were compared with the numerical simulation results obtained in this paper to verify the accuracy and correctness of the mathematical models and numerical methods. The experimental work described in NACA TN4007PR) is defined as:p2, t is the freestream total pressure. The results show that the numerical simulation results match the curve trend observed from the experimental data. Furthermore, the simulation results obtained in this paper are closer to the experimental data than those of Pratt et al.10. The simulation results have an error of less than 4.9% and a maximum error of no more than 19.8%, which indicates the established model has high accuracy. The total average error is about 10%.The pressure ratio (CD, and hinge moment M with the PRD opening angle and plenum compartment pressure are shown in Fig. Under different plenum compartment pressures and opening angles, the simulation results obtained for the relationship among the PRD discharge mass flow rate, discharge coefficient CD shows a downward trend with the opening angle increases. At different opening angles and plenum compartment pressures, the PRD discharge mass flow rate and the discharge coefficient CD in the pressure relief process under the vertical opening mode are higher than those under the horizontal opening mode.It can be seen from the Fig.\u00a0As shown in Fig.\u00a0CD of a PRD under the vertical opening mode is higher than that under the horizontal opening mode. The pressure relief process reaches a lower nacelle compartment pressure during the pressure relief equilibrium stage under the vertical opening mode compared with that under the horizontal opening mode. In conclusion, the vertical opening mode is better than the horizontal opening mode.Different opening modes will significantly impact the discharge and force characteristics of a nacelle PRD. The discharge coefficient Reducing the nacelle compartment pressure threshold of a PRD opening does not affect the nacelle compartment pressure and opening angle changes at the equilibrium stage. Still, it can reduce the time required for the pressure relief process to reach equilibrium. So decreasing the nacelle compartment pressure threshold is recommended.In the vertical opening mode, appropriately reducing the maximum opening angle can effectively mitigate the PRD reciprocating swing angle in the equilibrium stage and does not affect the nacelle compartment pressure in the equilibrium stage. Therefore, the pressure relief efficiency can be improved by appropriately reducing the maximum opening angle.However, the mathematical model established in this study is relatively limited. For example, the numerical model used is made of simple geometric constructions, which is different from the complex shape of the actual engine compartment. Therefore, it is necessary to conduct a more detailed model to make it more practical and explore the stress change and pressure relief flow mechanism of the PRD in future work."}
+{"text": "Psidium guajava) is a fruit infested in nature by the tephritids Anastrepha striata and A. fraterculus. In contrast, the extremely polyphagous A. ludens infests guava only under artificial conditions, but unlike A. striata and the Mexican A. fraterculus, it infests bitter oranges (Citrus x aurantium). We used these models to analyze whether the gut microbiota could explain the differences in host use observed in these flies. We compared the gut microbiota of the larvae of the three species when they developed in guava and the microbiota of the fruit pulp larvae fed on. We also compared the gut microbiota of A. ludens developing in C. x aurantium with the pulp microbiota of this widely used host. The three flies modified the composition of the host pulp microbiota . We observed a depletion of Acetic Acid Bacteria (AAB) associated with a deleterious phenotype in A. ludens when infesting P. guajava. In contrast, the ability of A. striata and A. fraterculus to infest this fruit is likely associated to a symbiotic interaction with species of the Komagataeibacter genus, which are known to degrade a wide spectrum of tannins and polyphenols. The three flies establish genera specific symbiotic associations with AABs. In the case of A. ludens, the association is with Gluconobacter and Acetobacter, but importantly, it cannot be colonized by Komagataeibacter, a factor likely inhibiting its development in guava.The gut microbiota is key for the homeostasis of many phytophagous insects, but there are few studies comparing its role on host use by stenophagous or polyphagous frugivores. Guava ( Bactrocera oleae (Gmelin) (Diptera: Tephritidae) and its obligate endosymbiont \u201cCandidatus Erwinia dacicola\u201d. This bacteria is essential for B. oleae larvae, playing a key role for its development in the immature olive  larvae and adults stemming from six natural hosts including the native ancestral Casimiroa edulis La Llave and C. greggii (S. Watson) F. Chiang (both Rutaceae), the exotics Mangifera indica L. cv. Ataulfo (Anacardiaceae), Prunus persica (L.) Batsch (Rosaceae), and Citrus x aurantium L. (Rutaceae), and the occasional native host Capsicum pubescens Ruiz and Pav. cv. Manzano (Solanaceae), observed a decrement in the relative abundance of representatives within the Acetobacteraceae, followed by an increment of representatives within the Enterobacteriaceae in larvae developing in certain hosts. This shift in the gut microbiota, with an increment in the ratio Enterobacteriaceae/Acetobacteraceae was most apparent in the marginal host C. pubescens, most likely leading to the poor larval development and high fitness costs observed by The gut of most living organisms harbors complex microbial communities, which are involved in multiple processes of the host\u2019s biology . In insere olive , as unrieuropein . \u201cCandidelopment . Recentlvia direct damage to the fruit (larvae feeding in them render the fruit unmarketable) or indirectly via severe trade restrictions  and mango (M. indica), as well as many other commercial fruit and vegetables  (A. ludens is represented by guava (Psidium guajava), a chemically defended fruit widely attacked by A. striata and the Mexican form of A. fraterculus , C. x aurantium or M. indica  . Importaterculus . In the nificant . Under t. indica . In contk citrus  [it doesk citrus ], but in persica . Althougs common , infestas common . While A tannins , A. fratyphenols .A. striata and A. fraterculus larvae developing in P. guajava, plus A. ludens larvae artificially developing in this fruit. The gut microbiota of A. ludens larvae developing in the natural host C. x aurantium was also analyzed to contrast the composition of the gut microbiota in a natural and a conditional/artificial host  interactions\u201d by st sensu. In bothby Birke  could beA. striata, A. fraterculus and A. ludens larvae were collected from mature fruit with signs of infestation in the orchard \u201cFinca Costa Rica\u201d . Additionally, we collected A. ludens from Teocelo, Veracruz . Guavas were harvested from various trees and were used to obtain A. striata and A. fraterculus. Meanwhile, bitter oranges (C. x aurantium) were harvested to obtain A. ludens. In both cases, the fruit were transported to the laboratory and immediately dissected to search for third instar larvae.Wild A. striata and A. fraterculus simultaneously infesting the same guava fruit  and determine the likely microbial exchange between the insect and the fruit, we also collected fruit pulp surrounding the larvae. Both larvae and pulp samples were frozen in liquid nitrogen and stored at \u221280\u00b0C until processing.A. ludens adults originating from infested bitter oranges (C. x aurantium) collected in neighboring localities. Procedures for fruit handling and harvesting of pupae/adults are described in detail in A. ludens adults emerged, they were kept in 30 \u00d7 30 \u00d7 30 cm3 plexiglass cages, covered with Teflon mosquito netting. They were supplied with water and an artificial diet ad libitum. The diet consisted of hydrolyzed protein  and refined cane sugar in a 1:3 ratio. The flies were kept for 15 days at 27 \u00b1 1\u00b0C, 63 \u00b1 5% RH and 12:12 h light: dark photoperiod until they reached sexual maturity. Females were not supplied with an oviposition substrate to promote the accumulation of eggs in their ovaries and thus facilitate oviposition into the non-natural host guava , with st guava .A. striata or A. fraterculus). We inspected all fruit to make sure that they did not show signs of infestation by Conotrachelus (Coleoptera: Curculionidae: Molytinae), a beetle commonly infesting guavas in the study region . These flies were released into the fruit-bearing branches we had previously covered to preclude wild flies and beetles from laying eggs into them. The sex ratio of flies in each bag was 1:1, and the amount of fruit in each bag was between two and four. At the moment of fly release, the guavas were at the \u2018yellow in transition\u2019 maturity stage  until processing within 24 h of collection.Larvae from guavas and oranges were superficially sanitized by the following series of washes: 1-min wash in 500 \u03bcl of washing solution , followed by a 1-min wash in 500 \u03bcl of 1% commercial sodium hypochlorite, and a 1-min wash in 500 \u03bcl of 70% ethanol. Finally, two 1-min washes were performed with 500 \u03bcl of sterile distilled water. After sanitization, the digestive tracts were dissected from the proventriculus to the terminal region of the hindgut. The dissection of larvae was performed with sterile forceps using a stereoscope . The sample unit was defined as a pool of five gut tracts. In addition, five mg of the pulp surrounding each larva were collected. Then, pools of 25 mg were obtained for each pulp sample. In turn, each sample unit had five replicates. Samples of larvae guts were kept in 450 \u03bcl of RNAlater\u2122 Stabilization Solution . The gDNA obtained was used for the amplification of the 16S rRNA gene. Primers for 16S rRNA gene v3-v4 amplification were selected from 1 : 16S Amplicon PCR Forward 5\u2032 TCGTCGGCAGCGTCAGATGTGTA TAAGAGACAGCCTA CGGGNGGCWGCAG and 16S Amplicon PCR Reverse 5\u2032 GTCTCGTGGGCTCGGAGATGTGTATAAGAGACAGGACT ACHVGGGTATCTAATCC. PCR amplification was performed using an enrichment strategy. The enrichment consisted of an initial amplification of 15 cycles, starting from 300 ng of gDNA. Then, 2 \u03bcl of PCR1 were used as a template to carry out a PCR2 of 25 cycles. The composition of PCR1 (25 \u03bcL) and PCR2 (50 \u03bcL) consisted of Qiagen buffer 1X, dNTPs 0.2 mM, MgCl2 0.1 \u03bcM, 16s Amp F and R 0.2 \u03bcM each one, and Taq Polymerase 0.05 U, with an amplification program of 94 \u00b0C/2 min, 15 (PCR1) or 25 (PCR2) cycles of 94 \u00b0C/15 seg, 55 \u00b0C/30 seg y 72 \u00b0C/1 min, and finally 72 \u00b0C/5 min. Amplicons were purified with the Promega Wizard\u00ae SV Gel and PCR Clean-Up System kit and their concentration was determined by a Qubit 2.0 Fluorometer . Purified PCR products were indexed with Ilumina\u00a9 sequencing adapters using the Nextera XT Index Kit from Illumina\u00a9 and were purified and quantified as previously described. The quality of the library was determined using the Agilent Bioanalyzer 2100\u00ae system. The indexed amplicons were sequenced in a paired-end format (2 \u00d7 300 bp) using a MiSeq Reagent Kit V3 (600 cycles), on a MiSeq platform from Illumina\u00a9. Sequencing was carried out by the Sequencing Unit of the BioMimic\u2122 Scientific and Technological Cluster at the Instituto de Ecologia, A.C.\u2014INECOL.Once the RNAlater\u2122 was removed, all samples were deep frozen in liquid nitrogen and pulverized with a sterile pistil. gDNA extraction was performed with the QIAamp DNA Mini Kit from QIAGENThe raw reads in paired-end layout (2 \u00d7 300) were processed in the QIIME2 (v. 2020.6) platform . We usedWolbachia) which were analyzed separately. To gain insight into specific species of ASVs classified within Wolbachia, we performed a BLASTn search against the SILVA database (v132) with a cutoff of e-value < 0.0003 and \u201c-max_target_seqs\u201d 3. To correct the bias related to the different size in the sample counts, both data sets were normalized with the cumulative sum scaling (CSS) method using the metagenomeSeq package  method. A Principal Coordinate Analysis was plotted to visualize the ordination of samples based on UniFrac distance metrics (beta diversity). With the aim of detecting differentially abundant genera in the gut microbiota between species, we also performed a linear discriminant analysis Effect Size (LEfSe) , we used the phyloseq  package  package . The com package . The alp package  in one w (LEfSe) . StatistWolbachia) was composed of 3,398,023 reads, with a sample mean of 84,951 reads. Considering all samples, we detected a total of 1,884 ASVs, distributed among 19 phyla, and 29 classes. Within the wide taxonomic spectrum, 11 classes exhibited more than 1% of relative abundance in at least one sample , but in the pulp, counts were very low, with a sample mean of 58. That is, the few Wolbachia\u2019s detected in the pulp of guava most likely stemmed from the feces of the larvae, as opposed to be naturally living there. In this data set, we detected 40 ASVs. The most abundant ASVs where closely related or belonging to Wolbachia pipientis wUni , likely playing a key role in metabolizing deleterious secondary metabolites in A. striata, a tephritid fly that attacks guava when still very unripe and rife with tannins; (3) the less abundant presence of the same bacteria in A. fraterculus, another tephritid species naturally developing in guava, albeit in a much more developed ripening stage when compared to A. striata; (4) the lack of Komagataeibacter in the guts of A. ludens forcibly infesting ripe P. guajava fruit and the dysbiotic state of the larval microbiota of this species when developing in guavas; (5) the apparent critical role of the Enterobacteriaceae/Acetobacteraceae ratio in the fitness of the three tephritid flies studied; 6) The endosymbiont W. pipientis wUni was present in large numbers in A. striata and A. fraterculus, but almost inexistent in A. ludens and the pulp of guava and C. x aurantium.Several findings stand out that we believe merit discussion: (1) Overall (considering the three fly species and two fruits studied), Alphaproteobacteria (49.8%), Gammaproteobacteria (34.1%), and Bacilli (5.1%) where the dominant groups of bacteria identified in guts of larva and fruit pulp; (2) the ubiquitous presence of Anastrepha species  to their surroundings by defecating, regurgitating, and sucking. This process could ameliorate the tannin-rich pulp condition via the microbiota metabolism, enriching its surroundings with beneficial taxa that potentially break down toxic compounds, as we infer from our results. We fully recognize, that ideally, we should have compared the two types of pulp, totally clean, uninfested pulp, and pulp in the vicinity of the feeding larvae. However, based on literature reports . We were interested in learning how larvae construct their niche in this \u201ctoxic\u201d environment and transfer their gut microbiota . It is known that when a fruit fly larva develops in guavas, particularly in unripe fruit, it is exposed to high concentrations of secondary metabolites .We were surprised/intrigued at the similarity of the gut microbiota of abolites , especiaabolites . In thispe fruit  that hasectively . Thus, iKomagataeibacter, Weissellla, and Gluconobacter, were differential between A. ludens and A. striata. Of them, Komagataeibacter, an Acetic Acid Bacteria (AAB) was represented in large numbers in A. striata, in lower ones in A. fraterculus and absent in A. ludens when forcibly infesting guava and naturally infesting bitter oranges. This observation suggests a trend from up to low abundance of this bacterial genus related with the maturation stage. The ASVs of Komagataeibacter that we found have high sequence similarity with Komagataeibacter intermedius and K. hansenii. K. intermedius is used for industrial production of bacterial cellulose , as guava is only infested by relatively few fruit fly species worldwide  when compared to the ancestral host, C. edulis (23.02 \u00b1 0.2) and grapefruit (18.2 \u00b1 0.2) (both Rutaceae). Based on the results obtained here on the microbiota of A. ludens larvae developing in P. guajava, as well, as those obtained by A. ludens developing on the poor host C. pubescens, a plausible explanation is the depletion of AABs symbionts in these two hosts.This shift could be related to the differential pupal weight of larvae developing in C. edulis, one of the purported ancestral hosts of A. ludens, can be considered a \u201csugar bomb\u201d with high carbohydrate content and low protein levels [carbohydrate/protein ratio of 18.4 (16.6/0.9%)], contrasting to what happens in P. guajava [8.7 (15.8/1.8%)] and C. pubescens [6.3 (6.32/1%)  ]. That ied fruit . Phytophed fruit . On the via their gut microbiota , where nitrogen-fixing bacteria, using the nitrogenase complex can transform the atmospheric dinitrogen into ammonia, which can be later assimilated as the non-essential amino acids glutamine and glutamate  delays the development of its larvae . Processed data to replicate the analysis are in the The raw data presented in this study are available in the NCBI repository (BioProject MA, AM, and MO-S: conceptualization. MO-S and DC-G: data curation and formal analysis. MA: funding acquisition. MO-S, AA-M, DC-G, and MA: investigation and methodology. MA and AA-M: project administration and resources. DC-G: software and visualization. MA, AA-M, EI-L, DD, and AM: supervision. MO-S, MA, and DC-G: writing\u2014original draft. MA, DC-G, MO-S, AM, AA-M, DD, and EI-L: writing\u2014review and editing. All authors contributed to the article and approved the submitted version."}
+{"text": "Arg1, Mrc1, and Igf1, and downregulated Tnf\u03b1 in M1-like macrophages compared to other treatment conditions. These licensing microgels are a potent immunomodulatory approach that shows substantial promise in elevating the efficacy of current MSC therapies and may find utility in treating chronic inflammatory conditions.Mesenchymal stromal cells (MSC) are sensors of inflammation, and they exert immunomodulatory properties through the secretion of cytokines and exosomes and direct cell-cell interactions. MSC are routinely used in clinical trials and effectively resolve inflammatory conditions. Nevertheless, inconsistent clinical outcomes necessitate the need for more robust therapeutic phenotypes. The immunomodulatory properties of MSC can be enhanced and protracted by priming (aka licensing) them with IFN\u03b3 and TNF\u03b1. Yet these enhanced properties rapidly diminish, and prolonged stimulation could tolerize their response. Hence a balanced approach is needed to enhance the therapeutic potential of the MSC for consistent clinical performance. Here, we investigated the concentration-dependent effects of IFN\u03b3 and TNF\u03b1 and developed gelatin-based microgels to sustain a licensed MSC phenotype. We show that IFN\u03b3 treatment is more beneficial than TNF\u03b1 in promoting an immunomodulatory MSC phenotype. We also show that the microgels possess integrin-binding sites to support adipose tissue-derived MSC (AD-MSC) attachment and a net positive charge to sequester the licensing cytokines electrostatically. Microgels are enzymatically degradable, and the rate is dependent on the enzyme concentration and matrix density. Our studies show that one milligram of microgels by dry mass can sequester up to 641 \u00b1 81 ng of IFN\u03b3. Upon enzymatic degradation, microgels exhibited a sustained release of IFN\u03b3 that linearly correlated with their degradation rate. The AD-MSC cultured on the IFN\u03b3 sequestered microgels displayed efficient licensing potential comparable to or exceeding the effects of bolus IFN\u03b3 treatment. When cultured with proinflammatory M1-like macrophages, the AD-MSC-seeded on licensing microgel showed an enhanced immunomodulatory potential compared to untreated AD-MSC and AD-MSC treated with bolus IFN\u03b3 treatment. Specifically, the AD-MSC seeded on licensing microgels significantly upregulated  Mesenchymal stromal cell (MSC) therapy has attracted clinicians and researchers as a viable treatment option for many inflammatory and immune diseases. MSC are a potent immunomodulatory tool that can regulate innate and adaptive immune responses through direct cell-cell and paracrine signaling . Their pH1 and TH17 proliferation  and HLA-G expression that are vital for sustaining Tferation , 6. But henotype . It shouThe efficacy of MSC therapies remains highly variable and is greatly affected by factors such as treatment timing, administration route, and hemocompatibility. In the clinic, MSC are administered locally or systemically, dependent on the clinical indication and pathology. But there is no clear consensus on the optimal mode of delivery . When adin vivo. Hence there is a critical need for strategies that can prolong the immunomodulatory phenotypes of MSC in vivo and prevent their rapid clearance from the site of injection. We propose a material-based approach that can anchor the MSC to the site of injection and prolong their immunomodulatory phenotype through a localized and sustained release of licensing factors.Like immune cells, MSC can keep a \u2018memory\u2019 of stimulus for a shorter period after moving to new environments . TherefoMaterial-based systems are used widely in applications that require a sustained release of biochemical factors and deliver therapeutic cells. For instance, bone tissue engineering approaches that release osteoinductive factors have already seen clinical application . MateriaHere, we characterized the physical and biochemical properties of the microgels and demonstrated their ability to ameliorate inflammatory conditions. We show that the microgels efficiently sequester IFN\u03b3 and release it upon degradation while readily supporting MSC attachment. We also show that the localized release of IFN\u03b3 promotes an immunomodulatory MSC phenotype efficiently. Further, we confirm their immunomodulatory phenotype through macrophage co-culture studies. In addition, we evaluated the osteogenic potential of the microgels to show that they naturally direct MSC toward an osteogenic line over a prolonged culture period. Overall, our microgels show substantial promise in elevating the efficacy of current MSC therapies and demonstrate an improved option to address persistent chronic inflammatory diseases. software  to determine the size and swelling rates.A stock solution was prepared by adding a precise amount of dehydrated and lyophilized microgels in 1X DPBS (stock = 0.67 mg/mL). 200 \u03bcL of the sock solution (~130 \u03bcg/well) was dispensed in a 96 well plate, and an equal volume of collagenase solution  was added to the wells at different concentrations and sealed with a clear plate cover to prevent evaporation. The fluorescence of the supernatant was then measured at Ex/Em 590/620 every 5\u00a0min for 48 hours to assess microgel degradation rates.For loading, 10 \u00b5L of IFN\u03b3 stock solution (100 \u00b5g/mL of DPBS) was added per 1 mg of lyophilized microgels and were placed in a humidified incubator for 24 hours. Then they were washed with 0.1 wt% bovine serum albumin (Sigma) in DPBS to remove unbound IFN\u03b3. The maximum loading capacity was calculated by performing an ELISA assay on fully degraded microgels and measuring unbound IFN\u03b3 in the remaining solution after loading. To fully degrade the microgels and release the bound IFN\u03b3, the loaded microgels were suspended in 1 mL of 100 U/mL collagenase solution and kept at 37\u00b0C until completely degraded. The supernatant was collected periodically and analyzed using an IFN\u03b3 ELISA kit to measure the release of IFN\u03b3 from the degrading microgel. The corresponding degradation rates of the microgels were also measured fluorometrically at Ex/Em 590/620.th passage and used for experiments.All studies involving animal cells or tissues are conducted according to approved protocols by the Institutional Biosafety Committee (IBC) protocol and Institutional Animal Care and Use Committee  at the University of Kentucky, respectively. Inguinal fat pads were isolated from adult mice (wildtype C57BL/6J - JAX stock #000664 and ACTb-EGFP expressing C57BL/6-Tg(CAG-EGFP)1Osb/J, JAX stock #003291), washed in serial dilutions of betadine and sterile 1X DPBS. The washed tissues were then finely minced with sterile scissors, and suspended in 0.1 wt% collagenase solution, and incubated at 37\u00b0C with constant agitation for 30\u00a0min or until most of the tissue was degraded. An equal amount of Dulbecco\u2019s modified eagle\u2019s medium  with 10% fetal bovine serum  was then added to neutralize the collagenase and filtered through a 100 \u03bcm cell strainer. The single-cell suspension was then centrifuged at 250\u00a0g for 5\u00a0min, supernatant aspirated, and the cell pellet was resuspended in DMEM+10% FBS and cultured in a 6 well plate. After 24 hours, the wells were washed twice with warm 1x DPBS to remove non-adherent cells and debris leaving the adherent adipose tissue-derived mesenchymal stromal cells (AD-MSC). The cells were culture-expanded until the 52 incubator. To license AD-MSC, they were cultured until ~80 confluent in growth media and replaced with licensing media, i.e., growth media supplemented with the cytokines , for 24 hours. AD-MSC were seeded on microgels by incubating IFN\u03b3 loaded/unloaded microgels in growth media for 1 hour prior to seeding. This growth media was aspirated and replaced by 100 \u00b5L of MSC suspension (~0.3 million cells per mg of microgel) in growth media. The suspension was kept in the incubator for 1 hour to allow cell attachment to the surface of the microgels. During this period the suspension was gently mixed every 10\u00a0min to prevent aggregation and allow uniform seeding. The microgels were then washed with fresh growth media to remove unbound cells. For co-culture studies, macrophages were cultured in corresponding growth media until ~80% confluent and then polarized for 24 hours using cytokines (50 ng/mL LPS and 25 ng/mL IFN\u03b3) suspended in growth media.Primary AD-MSC were cultured in growth media containing DMEM+10% FBS+ antibiotics and antimycotics (Gibco). An IC-21 murine macrophage cell line (ATCC) was acquired, cultured in expansion media (RPMI (Gibco) + 10% FBS + antibiotics and antimycotic), and used for co-culture studies. All cells were maintained at 37\u00b0C in a COFor gene expression analysis, the samples were collected in TRIzol\u2122 reagent (Invitrogen), and the whole RNA was isolated according to the manufacture\u2019s protocol. A SuperScript\u2122 III Platinum\u2122 One-Step qRT-PCR Kit (ThermoFisher) and TaqMan probes (ThermoFisher) were used to analyze gene expression profiles using a QuantStudio 3 real-time PCR system (ThermoFisher). The following TaqMan probes were used for our studies: Gapdh (Mm99999915_g1) as a housekeeping control, Alp (Mm00475834_m1), Bglap (Mm03413826_mH), Col1a1 (Mm00801666_g1), Ibsp (Mm00492555_m1), Sp7 (Mm04209856_m1), Spp1 (Mm00436767_m1), Runx2 (Mm00501584_m1) for AD-MSC osteogenic analysis. Il-1\u03b2 (Mm00434228_m1), Nos2 (Mm00440502_m1), Ccl2 (Mm00441242_m1), Tnf\u03b1 (Mm00443260_g1), Lgals9 (Mm00495295_m1), Tgf\u03b2 (Mm01178820_m1), Il-6 (Mm00446190_m1), Ptgs2 (Mm00478374_m1), Ppar\u03b3 (Mm01184322_m1), Arg1 (Mm00475988_m1), Cxcl2 (Mm00436450_m1), Mrc-1 (Mm00485148_m1), Il-12\u03b2 (Mm00434174_m1), Igf1 (Mm00439560_m1) and, Chil3 (Mm00657889_mH). Gene expression data were presented as heatmaps showing foldchange (different treatment) or Z-scores (concentration and time-dependent change).Cell proliferation was analyzed by quantifying the total DNA content of samples using Quant-iT\u2122 PicoGreen\u2122 dsDNA Assay Kit (ThermoFisher). Lambda DNA standard was used for generating a calibration curve. Briefly, samples were collected at the specified time points and homogenized through sonication in 1X Tris-HCl buffer and centrifuged at 10,000 g for 10\u00a0min. The supernatant was then mixed with PicoGreen, incubated in the dark for 5\u00a0min, and the fluorescence was read at Ex/Em wavelength of 485/528 nm.2 (Sigma) dissolved in DI water at different concentrations served as standards.The o-cresolphthalein complex one (OCPC) method was used to quantify calcium deposition by the AD-MSC on microgels as previously described . BrieflyThe collagen deposition by the AD-MSC was quantified using a Sircol soluble collagen assay kit (Biocolor). Briefly, the samples were homogenized through sonication in 1X Tris-HCl buffer and centrifuged at 10,000 g for 10\u00a0min. The supernatant was collected, and 20 \u03bcL of each sample was brought up to 100 \u03bcL with Tris-HCL buffer and then mixed with 0.5 mL of Sircol dye reagent. Samples were mixed for 30\u00a0min, centrifuged at 12,000 RPM for 10\u00a0min, the supernatant was decanted, and 750 \u03bcL of acid-salt wash reagent was added to the pellet. The suspension was mixed and centrifuged again at 12,000 RPM for 10\u00a0min, the supernatant was decanted, and 250 \u03bcL of alkali reagent was added to the pellet. This suspension was vortexed and incubated for 5\u00a0min at room temperature. The absorbance of the final solution was read at 555 nm. Rat-tail collagen was used to create a standard curve.A commercially available ALP activity fluorometric assay kit (BioVision) was used according to the manufacturer\u2019s protocol. Briefly, the samples were homogenized through sonication in Tris-HCl buffer and centrifuged at 10,000 g for 10\u00a0min. Then 100 \u03bcL of the supernatant was mixed with 20 \u03bcL of 0.5 mM 4-methylumbelliferyl phosphate (MUP) substrate and incubated at 25\u00b0C in the dark for 30\u00a0min. Finally, 20 \u03bcL of stop solution was added to the mixture, and fluorescence was measured at Ex/Em of 360/440 nm. Samples of 0.1-0.5 nmol of MUP substrate with purified ALP enzyme were used to create a standard curve.Samples were fixed in a 10% aqueous buffered zinc formalin  overnight, washed with 1X DPBS, and briefly incubated in a 0.1% Triton X-100 (Sigma) in 1X DPBS solution for 5\u00a0min before staining. Phalloidin conjugate  was used to stain the F-actin of the cell cytoskeleton and DAPI (Invitrogen) for staining the nucleus. Samples were stained for 30 mins in the dark and washed with 1X DPBS. Z-stacks of the stained samples were acquired using a Nikon A1R confocal microscope and processed using Fiji (ImageJ) software (NIH).Samples were fixed with a 3% glutaraldehyde in sodium cacodylate buffer overnight, washed with DI water, and incubated in 2% Osmium Tetroxide solution for 16 hours. Samples were washed with DI water and dehydrated using ethanol series. Samples were then flash-frozen in liquid nitrogen and lyophilized overnight. The dried samples were sputter-coated with a 5 nm thick layer of platinum using Leica Ace 500 sputter coater and imaged using an FEI Quanta 250 environmental scanning electron microscope (SEM).All measurements were performed at least in triplicate. Data are plotted as means with error bars representing the standard deviation. The Pearson correlation coefficient (r) was used to evaluate the linear correlation between two variables. One- and Two-Way ANOVA were used to identify significance within groups, and Holms-Sidak pairwise comparisons followed to identify which groups were significant.) and stored at room temperature for extended periods without any deterioration to their properties and can be rehydrated within seconds for cell delivery. When hydrated microgels were suspended in collagenase solution, they exhibited a concentration-dependent degradation (pI) of ~8.7  was seen in the TNF\u03b1 treatment, but it also exhibited the highest upregulation of inflammatory cytokines (Il-6 and Ccl2). IFN\u03b3 treatment, on the other hand, upregulated immunomodulatory factors while having modest effects on inflammatory factors. Hence, IFN\u03b3 is deemed more suitable for sustained licensing of AD-MSC.We first investigated the concentration-dependent effects of two potent cytokines, IFN\u03b3 and TNF\u03b1, to sustain the immunomodulatory phenotype of adipose tissue-derived MSC (AD-MSC). A low (0.1 ng/mL) and high (100 ng/mL) concentration of the IFN\u03b3 and TNF\u03b1 and a mixed treatment condition (10 ng/mL of each) were studied. AD-MSC that express green fluorescence protein conjugated to the actin cytoskeleton (ACTb-EGFP) were used to monitor changes in morphology and viability longitudinally  showed a concentration-depended increase in the expression levels of the key immunomodulatory markers Table\u00a02.Nos2, one of the critical factors of the licensed phenotype, was downregulated after 24 hours. In contrast, Ptgs2 expression remained consistent until day 7, when it was significantly downregulated, indicating a sustained phenotype. Ccl2 expression dropped within 24 hours and continued to drop until day 7. Tnf\u03b1 expression, on the other hand, showed downregulation after 24 hours but bounced back to licensed levels. Il-6 expression remained unchanged until day 7, while Lgals9 and Tgfb did not show significant changes. Overall, the licensed phenotype of the AD-MSC were partially retained for a week after licensing signals were removed.Then we investigated the retention of the licensed AD-MSC phenotype over 7 days after the initial treatment. The AD-MSC were treated with 10 ng/mL of IFN\u03b3 for 24 hours, washed thoroughly to remove all cytokines, and monitored for 7 days Table\u00a03.Nos2 over time  and evaluated their potential in modulating AD-MSC phenotype and function using co-culture studies. We compared the effects of the licensing microgels to an equivalent bolus media (60 ng/mL of IFN\u03b3) treatment that is renewed every other day. Untreated AD-MSC served as controls. In all co-culture conditions, the AD-MSC were seeded on the microgels, as shown  macrophages and treats inflammatory colitis , they promoted a robust immunomodulatory AD-MSC phenotype where Ptgs2 is upregulated, and Tnf\u03b1 is downregulated. Over time, some licensed phenotypes, notably, Nos2 and Ccl2 expressions, diminish if the media is not replenished with IFN\u03b3. Then we tested the ability of the licensing microgels to prolong a licensed-MSC phenotype.MSC are the sensors of the body\u2019s immune response, can be derived from various adult tissue sources, and possess immune modulatory functions. They can promote inflammation when the immune response is underactive and restrain the inflammatory response when it is overactive. The immunomodulatory effects of MSC vary depending on their tissue source and activation state. The major immunomodulatory factors secreted by MSC include NO, PGE2, TGF\u03b2, Galactin 9, and TSG6 . In our feration . Ccl2, aresponse . But, st colitis . Notablyreg axis . Furtherervation . They caervation . This isThe unique physical and chemical properties of gelatin microgels make them versatile carriers for cell and growth factor delivery. They can form complexes with different drugs, cytokines, and growth factors and are therefore widely used in drug delivery and tissue engineering applications . The tunArg1, Mrc1, and Igf1 in macrophages more robustly compared to monolayer cultures. These enhancements can be attributed to the tissue-like microenvironment provided by the microgels. It is known that substrate mechanics influence MSC phenotype, including their immunomodulatory capabillites  protocol and Institutional Animal Care and Use Committee (IACUC) at the University of Kentucky.MP and RT contributed to the conception and design of the study, performed the statistical analysis, and wrote the manuscript. MP performed all experiments and data acquisition. All authors contributed to manuscript revision, read, and approved the submitted version.Research reported in this publication was supported in part by the National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS) Award Number R21AR078447, National Institute of General Medical Sciences (NIGMS) of the National Institutes of Health under Award Numbers P20GM130456 and P20GM103436-20 , National Center for Research Resources and the National Center for Advancing Translational Sciences of the National Institutes of Health under Award Number UL1TR001998, and Orthopedic Trauma Association . The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health or other grant funding agencies.The authors would also like to thank Dr. Eric Blalock in the University of Kentucky Department of Pharmacology and Nutritional Sciences for consultation and advice on statistical analysis of the gene expression data.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher."}
+{"text": "All pandemic outbreaks because of their rapid spread and high mortality rate cause to everyone considerable stress and anxiety.The aim of the present study is to investigate how news media exposure moderates the relationship between stress, anxiety, depression and self-efficacy, social support, knowledge of the coronavirus and coronavirus perception.223 healthcare providers, men 46 (20.6%) and women 177 (79.4%), working in hospitals in Greece participated in the study. independent t-test, one-way ANOVAs, Pearson\u2019s correlation, multiple-linear regression and moderator\u2019s analysis were analyzed with SPSS23.Organization support, friends support, covid-19 knowledge and covid-19 perception are most significant predictors to stress, F  = 11.47, p < .001 and Adjusted R2- .159. Friends support, covid-19 knowledge and self-efficacy, working with covid19 patients and gender are most significant predictors to anxiety, F  = 11.16, p < .001 Adjusted R2- .186. Friends support, covid-19 knowledge and self-efficacy and organization support are most significant predictors to depression, F  = 16.37, p < .001 Adjusted R2-squared: .217. News media exposure did moderate the predictive power of almost all predictors for stress, anxiety and depression, at p<.05.Therefore, the study verifies previous findings arguing that stress, anxiety and depression are strongly associated with numerous factors. These associations seem to be moderated by news media exposure. It is recommended to further explore the impact news media exposure has during crucial periods, such as covid-19 outbreak.No significant relationships."}
+{"text": "The results from HOMO/LUMO and density of states plots indicate a metallic transition from the typical semiconducting phase, near Fermi energy level (FE) as a function of bending, which can mainly occur due to bending curvatures inducing a stretching and contracting of sulfur-sulfur chemical bonds located mostly over basal (001)-plane; furthermore, molybdenum ions play a major role in such transitions due to reallocation of their metallic d-character orbitals and the creation of \u201cfree electrons\u201d, possibly having an overlap between Mo-dx2-y2 and Modz2 orbitals. This research on the metallic transition of 2H-MoS2 allows us to understand the high catalytic activity for MoS2 nanostructures as extensively reported in the literature. We present a systematic density functional theory study to determine the electronic structure of bending 2H-MoS We aim to determine the electronic structure in 2H-MoS2 and how it is influenced by mechanical bending. With this, we achieve a theoretical understanding of localized \u201cfree-electrons\u201d in terms of curved nanostructures and the characteristics of mechanical resilience as observed by in-situ TEM [2 layer is bent, as well as its electronic structure by the density of states near energy Fermi level (FE). Finally, we correlated the electrical properties of the bent 2H-MoS2 slab as a viable material for catalytic and energy storage based on our results.Molybdenum disulfide  . MoS2 carvations . Beyond rvations ,19. Prev \u00b1 2 GPa , similaro 90 MPa . Here, wsitu TEM . We pres2 molecular layer containing 117 atoms and having dimensions of 22 \u00c5 \u00d7 10 \u00c5 in length and width; no periodic conditions are considered. This layer was cut in the 001 direction from a unit cell of a hexagonal 2H-MoS2 (space group P63/mmc) with lattice parameters of a = b = 3.1 \u00c5, and c = 18.4 \u00c5. Then, by using the open access Avogadro Chemistry\u00ae package [2 model was performed, from which frames of the molecular model while bending were exported to the Materials Studio\u00a9 package, focusing only on 10\u00b0, 22\u00b0, 44\u00b0, and 75\u00b0 bending angles. These bending angles match observations made on in-situ TEM, as schematically described in Our computational study was completed using a conventional 2H-MoS package , a serie3 implementing the Generalized Gradient Approximation (GGA) and Perdew\u2013Burke\u2013Herzenhof (PBE) as exchange-correlation functional; for single point energy calculations a self-consistent field (SCF) convergence criteria of 1 \u00d7 10\u22126 eV/atom under double numerical polarization (DNP) basis set was used; all species were treated with effective core potentials and unrestricted spin with thermal smearing value of 0.027 eV (0.001 Ha) together with a long-range dispersion correction DFT-D2 [HOMO and LUMO energy levels localization was calculated by density functional theory (DFT) code Dmoln DFT-D2 .2 preparation can be found in previous works by the author [Transmission electron microscopy was performed using a JEOL 2010  equipped with a field emission electron beam at 200 kV. Detailed descriptions of MoSe author . AFM-TEMe author ,22.2 molecular layer. The mean Mo-S bond length is reduced from 2.4 \u00c5 to 1.2 \u00c5 when the slabs go from 0\u00b0 to 75\u00b0, which may result in strong interaction between s, p, and d orbitals, causing direct influence on the electronic structure of 2H-MoS2, as suggested previously by Bhattacharyya et al. [We verify that Mo-S bond length is reduced by 25% at the bending angle of 75\u00b0and that curving effects between energy levels occur in intensity by comparing the resulting electronic structure. As a result, we can compare the resulting geometry of the layer by estimating the cord length, radius, and arc length of the layer . The resa et al. ; this bo2 [In the HOMO-LUMO plots, energy levels localize as density profiles as presented in 2 .2 layer at angles of 10\u00b0 and 22\u00b0, and also mixed molybdenum and sulfur contribution was encountered. For bending angle values of 10\u00b0, the LUMO distributes around molybdenum ions all over the layer, providing theoretical proof that the majority MoS2 conduction band consists of molybdenum d-character orbitals, as described by Yazyev & Kis [FE) level [Now, when inspecting LUMO energy density plots, it is possible to detect that the distribution of the orbital remains mainly at the center site of the 2H-MoSev & Kis . HoweverF) level .F, which is higher at 0\u00b0 and at 44\u00b0 compared to the other bending angles. This is explained by recalling the previous results on \u0394E, which may indicate that if the two levels are more separated, electron wave functions are less prone to overlap and thus, populate more states near FE. At 0\u00b0 of bending (unstrained molecular layer) we have an estimation of 30 electrons in the vicinity of FE  reflect metallic or semi-metallic behavior for angles as low as 10\u00b0 of bending curvature  was found using Equation (1).Furthermore, to provide more information about metallic transition when the 2H-MoSslab + hydrogen corresponding to the energy of slab with adsorbed (and dissociated) hydrogen molecules, Eslab is the total energy of slab before reaction, and Ehydrogen is the energy of diatomic hydrogen. Hence, a negative value of Eads indicates an endothermic process, while a positive value indicates an endothermic reaction.With E2 towards hydrogen adsorption, showing that in all sites a reduction in adsorption energy is about 50% for a layer bent at 44\u00b0 compared to adsorption for a layer bent at 10\u00b0, and a dramatic decrease in energy adsorption compared to the unstrained molecular layer ; the reaction considers that H2 and H2S are released as byproducts after sulfur substitution with carbon atoms.For sulfur substitution with carbon atoms, which is considered a carburization process as discussed by others ,32, the (slab+C) is the energy of a 2H-MoS2 curved layer with n carbon atoms substituting n-sulfur atoms, E(H2S) is the energy of H2S, E(H2) is the energy of diatomic hydrogen, and Eslab is the total energy of layer before the reaction. E(CHy) is the energy of the carbon source, and y corresponds to the number of hydrogen atoms in the carbon source. Here again, a negative value of Eads indicates an endothermic process, and thus, a spontaneous one, while a positive value indicates an endothermic process. Results indicate that the energy demand for sulfur substitution by carbon is higher for 2H-MoS2 when it is bent at 44\u00b0 compared to when the layer is curved at 44\u00b0, which is in contrast to the previous results on hydrogen adsorption -basal plane. This transition is easily translated into an increase in chemical reactivity mainly for hydrogen adsorption and carbon substitution; at sulfur sites, 50% less energy is required for bending angles of 44\u00b0 as compared to bending angles of 10\u00b0. Lastly, the resilience character of 2H-MoS2 as described in the in-situ HRTEM experiment was also found in the molecular dynamics simulations using the Avogadro\u00a9 1.2.0 package displayed in the video provided in Based on information from an in-situ HRTEM study related to mechanical deformation of 2H-MoSs et al. , we are"}
+{"text": "The aim of this was to analyze the effect of different treatment options on radial neck fractures in children and to explore the factors affecting the prognosis of fractures.The clinical data of 131 children with radial neck fractures admitted to our hospital from 2010 to 2018 were retrospectively analyzed, and the patients were divided into 6 groups according to treatment methods . Postoperative elbow function and complications were analyzed.Among the 131 patients with fractures, the median age was 8\u00a0years, the median preoperative angulation was 52\u00b0, the follow-up rate was 86.3% (113/131), the average follow-up time was 58.3\u00a0months, and the postoperative complication rate was 17.7% (20/113). The comparison among the different treatment groups showed that group B had the best recovery of elbow function, postoperatively, and the lowest postoperative complication rate. Age, duration of hospitalization, and preoperative angulation were independent factors affecting postoperative complications. Older age, longer duration of hospitalization, and higher angulation increase the postoperative complications.Different treatment options have different efficacies for radial neck fractures in children, of which manipulative reduction with internal fixation using ESINs can achieve good efficacy and a low postoperative complication rate. Age, duration of hospitalization, and preoperative angulation are independent factors for postoperative complications. Radial neck fractures in children are an uncommon type of fracture, accounting for 1% of all body fractures and 5\u201310% of elbow fractures . BecauseAll patient records in our hospital from January 2010 to December 2018 were retrospectively reviewed. The exclusion criteria were as follows: (1) multiple fractures, (2) conservative treatment, (3) nontraumatic radial neck fractures, and (4) accompanying medical diseases requiring special treatment, such as leukemia or malignant tumors. Preoperative fracture angulation and displacement were measured according to the Judet classification . PostopeManipulative reduction was performed mainly with the Patterson technique, Israeli technique, or Neher and Torch technique with pressure over the radial head and rotation of the forearm in various positions of elbow flexion and extension \u201311. Perc\u03b1 = 0.05, and P < 0.05 was considered to indicate statistical significance.All 14 surgeons who participated in this study received standardized training in pediatric orthopedics, were certified, and had at least 5 years of working experience in pediatric orthopedics. All data were statistically analyzed using SPSS 20.0 software . The clinical data of the patients are described by statistical indicators such as the mean, standard deviation, median, interquartile range (IQR), and percentage. The Kruskal\u2013Wallis, Chi-square, Mann\u2013Whitney, Fisher\u2019s exact, and Student\u2019s t tests were used to statistically analyze the differences among different treatment groups. A logistic model was used to identify the prognostic factors affecting radial neck fracture results. All analyses were performed with two-sided tests and a test level A total of 131 patients were included in this study, of whom 18 were lost to follow-up, with a median follow-up rate of 86.3% and a mean follow-up time of 58.3 months . The median age of the patients was 8 years (IQR 6\u201310 years), and 45% were male. The median preoperative angulation in the 131 patients was 52\u00b0 (IQR 39\u201365) . Among these factors, group B had the most \u201cexcellent\u201d elbow scores (84.6%) without any \u201cpoor\u201d scores, and group F had the minimal \u201cexcellent\u201d elbow scores (43.8%) with most \u201cpoor\u201d scores (18.8%). However, there was no significant difference in gender, weight, time of receive surgery, and postoperative complications among groups Tables  and 3.Tan = 14, 14.4%), open reduction group: ] and relatively good Tibone function evaluation results . However, the patients in the open reduction group had relatively large preoperative angulation degrees . The clinical data were compared between the K-wire group and the ESIN group, and significant differences were found in age , weight , time of implant removal , and hospitalization cost  (P < 0.05). Under the same conditions, ESINs were preferred for older patients.In addition, the patients in the closed reduction group had relatively few postoperative complications , 1 case of heterotopic ossification, and 2 cases of joint stiffness Table . The logAt present, there are many treatment options for radial neck fracture in children, and the reported conclusions differ \u201315. A smThrough statistical comparisons of the clinical data of patients who received different treatment regimens, we found that there was a statistically significant difference in age. Moreover, despite further statistical analysis, we failed to find a significant correlation between age and postoperative function, which is different from previous studies , 19, 20.P = 0.464), and there was also no significant difference in the Tibone elbow outcome score and postoperative complications ; we did not find the time of receive surgery to be an independent predictor of efficacy, which was different from the results of previous studies [As the blood supply to the radial head mainly passes through the reverse supply of the radial neck in children , it is c studies , 21. We At present, there is no consensus on the application ranges of different surgical treatment options for preoperative angulation , 22\u201324. P = 0.047). Overall, group B had the most excellent and good scores , because the effective manual reduction method together with the hook structure of the ESINs assisted in the reduction of the fracture, and the fracture site was firmly fixed with little damage to the surrounding soft tissues. These factors were conducive to repairing the fracture site, and good functional recovery was achieved. However, it should be noted that ESINs are not applicable to younger patients due to size limitations, and indwelling in the body for a longer time, and cost is higher. After comparing different treatment groups, we found no significant difference in postoperative complications (P = 0.164). The main evaluation indexes of the Tibone elbow outcome scoring system include patient symptoms (pain) and range of motion and carrying angle [In this study, the Tibone elbow outcome scoring system was used to evaluate and compare the postoperative function of patients receiving different treatment regimens, and the results were statistically significant (ng angle , 25, whiAge, duration of hospitalization, and preoperative angulation were independent risk factors for postoperative complications. The probability of postoperative complications was higher for patients with an older age, longer duration of hospitalization, and greater preoperative angulation. We inferred that these factors were associated with postoperative nosocomial infection. Previous studies have shown an association between the duration of hospitalization and postoperative nosocomial infection . We founThis study is a single-center retrospective study and the research level is relatively low. However, based on the large number of samples from this clinical retrospective study, we can learn about the relevant clinical characteristics of radial neck fractures in children and also evaluate the efficacy of different treatment options, but further studies are required to confirm it.To obtain good functional recovery and a low incidence of postoperative complications, manual reduction combined with ESIN internal fixation may be the preferred treatment for radial neck fractures in children. However, for patients with large preoperative angulation degrees and severe injury, open reduction is still an indispensable approach. Performing reasonable and effective preoperative planning, shortening the duration of hospitalization, and applying other measures can reduce the occurrence of postoperative complications."}
+{"text": "In smart home environments, the interaction between a remote user and devices commonly occurs through a gateway, necessitating the need for robust user authentication. Despite numerous state-of-the-art user-authentication schemes proposed over the years, these schemes still suffer from security vulnerabilities exploited by the attackers. One severe physical attack is the node capture attack, which allows adversaries to compromise the security of the entire scheme. This research paper advances the state of the art by conducting a security analysis of user-authentication approaches regarding their vulnerability to node capture attacks resulting in revelations of several security weaknesses. To this end, we propose a secure user-authentication scheme to counter node capture attacks in smart home environments. To validate the effectiveness of our proposed scheme, we employ the BAN logic and ProVerif tool for verification. Lastly, we conduct performance analysis to validate the lightweight nature of our user-authentication scheme, making it suitable for IoT-based smart home environments. The Internet of Things (IoT) has rapidly expanded, with many interconnected physical nodes exchanging data and information . The groAs the number of devices within IoT networks continues to increase, it becomes crucial to address management and security concerns for remote users. Security, in particular, poses a significant challenge for IoT networks, necessitating secure information exchange with attributes such as confidentiality, integrity, and availability to resist potential security attacks ,6. AmongTo overcome these challenges and achieve secure user authentication, numerous user-authentication schemes for IoT-based smart home environments have been proposed in the literature. However, these authentication schemes often focus on general security attributes and neglect the threat of node capture attacks, particularly severe in resource-constrained devices within smart home environments. Without protection against node capture attacks, an adversary can compromise an authentication scheme, underscoring the need to design user-authentication schemes that provide mutual authentication and resist such attacks\u2019 consequences.The proposed scheme in this work is a lightweight authentication solution designed to address the security challenges faced in the smart home environment. We comprehensively analyze prevailing authentication mechanisms vulnerable to node capture attacks in IoT-based smart home environments. Our assessment identifies the shortcomings and security gaps present in these mechanisms.We introduce a novel user-authentication scheme designed to counter node capture attacks and fortify the security posture of IoT-based smart homes. This scheme is a pioneering response to the evolving threats in this domain.Our proposed scheme undergoes rigorous formal and informal analyses to validate its security strength. This ensures that our solution meets the stringent security requirements expected in smart home environments.We demonstrate a marked improvement in computation and communication costs compared to existing approaches through meticulous performance analysis. This efficiency enhancement is a significant advancement in IoT-based smart home security.The contributions of this paper are as follows, emphasizing the innovation brought forth by our study:Paper Organization: The rest of the paper is organized as follows: Many user-authentication schemes have surfaced in the literature in IoT-based smart home environments. These schemes share the goal of providing robust authentication mechanisms, although they exhibit varying effectiveness and security vulnerabilities. In this section, we explore the notable contributions in the field and present a meticulous comparative analysis of their respective schemes.Vaidya et al.  proposedLi  proposedSimilarly, Kumar et al.  also proThe comparative analysis shows that the existing user-authentication schemes for IoT-based smart home systems have various vulnerabilities and lack essential security features. Therefore, there is a need for an improved user authentication protocol that addresses these flaws and provides a higher level of security. In the following sections, we propose a novel and improved user-authentication scheme for IoT-based smart home systems, which mitigates identified vulnerabilities and enhances overall security. We also perform a detailed security analysis of our proposed scheme.The threat model is an essential component of the security analysis, providing a clear understanding of the attacker\u2019s assumptions and capabilities within the protocol\u2019s context. In this section, we define the threat model based on the Dolev\u2013Yao model  and outlCommunication Interception: Eve can intercept, inject, remove, or send new messages when two participants communicate over the public channel. This means that any information exchanged over the public channel is susceptible to manipulation or eavesdropping by Eve.Parameter Understanding: Eve can understand all the parameters exchanged over the public channel. This implies that Eve can analyze and comprehend the content of the messages transmitted between participants.Attacker Identity: Eve can be an outsider or a dishonest participant within the system. This encompasses the possibility of external attackers attempting to compromise the system\u2019s security and internal attackers with insider knowledge or unauthorized access.Gateway Security: The gateway, which plays a crucial role in the protocol, is assumed to be a secure entity. This means Eve cannot compromise the gateway or gain unauthorized access to its resources or sensitive information.Secret Parameter Protection: Eve cannot access the secret parameters used in the protocol. These secret parameters are assumed to be securely transmitted between the relevant parties and are not accessible or known to Eve.By outlining these assumptions, the threat model provides a clear understanding of the capabilities and limitations of the attacker within the proposed protocol. It helps identify potential vulnerabilities and design appropriate security measures to mitigate them.The proposed protocol follows a general network model used in smart home environments, as depicted in During the pre-deployment phase of smart devices in the network, it is assumed that the gateway has shared its identity credential and the hash of the shared key Each smart device has a unique identity and a shared key The identity of the gateway .The gateway generates two random numbers  and the shared key  along with a new random number S-5: By applying S-4 and the Session Key rule, the following is obtained:S-6: By applying the Nonce Verification rule, the following is obtained:Message 2:M-2:\u00a0S-7: By applying the Seeing rule, the following is obtained:S-8: By applying the Message Meaning rule and S-7, the following is obtained:S-9: By applying the Freshness Concatenation rule and S-8, the following is obtained:S-10: By applying the Jurisdiction rule and S-9, the following is obtained:S-11: By applying S-10 and the Session Key rule, the following is obtained:S-12: By applying the Nonce Verification rule and S-11, the following is obtained:Message 3:M-3:\u00a0S-13: By applying the Seeing rule, the following is obtained:S-14: By applying the Message Meaning rule and S-13, the following is obtained:S-15: By using S-14 and the Freshness Concatenation rule, the following is obtained:S-16: By applying the assumption S-15 and the Jurisdiction rule, the following is obtained:S-17: By applying S-16 and the Session Key rule, the following is obtained:S-18: By applying the Nonce Verification rule, the following is obtained:Message 4:M-4:\u00a0S-19: By applying the Seeing rule, the following is obtained:S-20: By applying the Message Meaning rule and S-19, the following is obtained:S-21: By applying S-20 and the Freshness Concatenation rule, the following is obtained:S-22: By applying the Jurisdiction rule and S-21, the following is obtained:S-23: By applying the Session Key rule, the following is obtained:S-24: By applying the Nonce Verification rule, the following is obtained:After analyzing the scheme using BAN logic, it can be concluded that the proposed protocol achieves mutual authentication and securely establishes session key agreement.ProVerif is an automatic tool used for analyzing the security of cryptographic protocols . It veriThe following is the interpretation of the query-wise result of the ProVerif analysis.Query 1: The query \u201cnot attacker(TIDUinew)\u201d returns true, indicating that the new identity (TIDUinew) is secure from attacks.Query 2: The query \u201cinj-event(end_U(IDUi)) ==>inj-event(start_U(IDUi))\u201d returns true, indicating that the connection functions securely for starting and closing on the user mobile.Query 3: The query \u201cinj-event(end_GWN(IDGW)) ==>inj-event(start_GWN(IDGW ))\u201d returns true, indicating that the connection on the gateway node is securely opened and closed.Query 4: The query \u201cinj-event(end_SD(SIDj)) ==>inj-event(start_SD(SIDj))\u201d returns true, indicating that the connection on the smart devices is securely opened and closed.The ProVerif analysis confirms that the proposed protocol is secure and achieves the intended security properties of secrecy and authentication.This section presents a security requirements analysis for user authentication protocols, focusing on the resistance to node capture attacks. Both general and specific functional and security requirements have been utilized to achieve the intended security properties of the schemes. Our proposed approach achieves all the security requirements, especially resistance to known attacks and node capture attacks, by comparing with the existing approaches ,22,23,24To evaluate the proposed user authentication protocol\u2019s resilience against node capture attacks, we adopt the approach presented by Wang et al. . The detExploited Vulnerabilities \u2192 Attack ConsequencesInsecure Identity Transmission In the proposed protocol, the mobile user does not use its original identity but instead employs a temporary identity updated by the gateway in each session.Insecure Transmission of Secret Key The mobile user does not directly transmit its shared secret key Improper Distribution of Secret Key Each smart device possesses a unique shared secret key with the gateway. If a node capture attack compromises a smart device (Exposure of User\u2019s Secret Parameter During the authentication phase, the mobile user\u2019s secret parameters are not forwarded in exchanged messages. These secret parameters encrypt the parameters exchanged over the public channel and a random number. If a compromised smart device attempts to compute the user\u2019s secret parameters, it will fail to extract any relevant information. Hence, an adversary cannot impersonate the mobile user in the proposed protocol.Mobile User Fails to Identify Smart Devices During the authentication phase, the mobile user selects the smart device to authenticate mutually. The mobile user possesses knowledge of the identities of all the smart devices connected to the network. Suppose the user fails to identify the smart device correctly based on its identity. In that case, it indicates that an adversary has either changed the identity of the smart device or the smart device is unresponsive when receiving authentication messages from the gateway. However, impersonating a compromised smart device does not lead to the impersonation of all smart devices within the system. This is due to each smart device\u2019s unique shared secret keys.kInsecure Transmission of Secret Key k but uses it only for session key The gateway, considered a secure entity in the proposed scheme, does not transmit its secret key Forward Secrecy Issue The proposed scheme achieves forward secrecy, as discussed in the security requirements above. An adversary cannot derive the session key computation from a previous session since only the trusted entity, the gateway, can compute the session key.Improper Distribution of Smart Device Secret Keys With its unique identity, each smart device must be registered with the gateway before joining the environment. The gateway distributes a unique secret key corresponding to each smart device\u2019s identity. Additionally, the session key is updated during each session. Consequently, even if an adversary manages to capture a node and obtain the session key, it does not compromise the security of the entire system.Insecure Transmission of Updated Session Key The gateway entity updates the session key using its secret key. The new session key (This section compares the proposed protocol with previously proposed security protocols ,22,23,24The comparison of communication cost is shown in The proposed protocol exhibits lower communication costs than the mentioned protocols, except for the Fakroon et al.  scheme. The comparison of computation cost is shown in The computation time experiment by Kilinc and Yanik  is used The execution/running time of the proposed protocol is 0.0299 ms. The comparison of the computational cost of the proposed approach with respect to the state-of-the-art approaches ,22,23,24This paper comprehensively analyzed state-of-the-art user-authentication schemes in the context of smart home systems. Our analysis identified several limitations and security vulnerabilities in existing schemes, highlighting the need for an improved solution. To address these shortcomings, we propose a secure and enhanced user-authentication scheme tailored for smart home environments. We performed a thorough security analysis of our protocol using formal computational models such as BAN logic and ProVerif tools. The evaluation demonstrated that our scheme effectively mitigates various security vulnerabilities, providing robust protection against attacks. Furthermore, we conducted a performance analysis to assess the computational and communication costs of the proposed scheme. The results indicated that our protocol achieves efficiency in resource utilization, making it suitable for deployment in IoT-based smart home environments.Our future work will primarily focus on the dynamic aspects of user authentication within smart home environments. This entails exploring adaptive authentication mechanisms capable of accommodating changes in user profiles, roles, and permissions within the smart home system. Additionally, we plan to investigate techniques to improve the scalability and interoperability of user-authentication schemes, facilitating seamless integration with a diverse array of smart home devices and platforms. By addressing these areas, our objective is to bolster the security, usability, and flexibility of user authentication in smart homes. Ultimately, we aim to contribute to developing robust and efficient authentication solutions for future IoT applications, thereby safeguarding the privacy and security of smart home users."}
+{"text": "Acute myeloid leukemia (AML) can cause acute abdomen following adrenal insufficiency or adrenal infarction. Therefore, when diffusely enlarged adrenal glands and adrenal insufficiency of unknown cause are seen in a patient presenting with acute abdomen, adrenal infarction due to AML, or other hematologic diseases should be ruled out.A 49\u2010year\u2010old man developed acute abdominal pain following adrenal insufficiency and was diagnosed with acute myeloid leukemia (AML) with myelodysplasia\u2010related changes. Because AML can cause acute abdominal pain due to adrenal infarction following adrenal insufficiency, a patient with these conditions should be ruled out adrenal infarction due to AML or other hematologic diseases. Thoracoabdominal computed tomography with contrast enhancement on admission revealed diffusely enlarged bilateral adrenal glands with diffuse internal low\u2010attenuation areas. We report a case of abdominal pain and adrenal insufficiency due to AML\u2010induced adrenal infarction. Furthermore, endocrinologic diseases such as ketoacidosis and adrenal insufficiency, systemic diseases such as vasculitis, and dermatologic diseases should be considered as differential diagnoses when the cause of acute abdomen cannot be identified.The causes of adrenal insufficiency include autoimmune diseases; infectious diseases such as tuberculosis; metastases of malignancies such as lung cancer, breast cancer, stomach cancer, colon cancer, and lymphoma; adrenal hemorrhage; and drugs.We herein report a case of acute abdominal pain and adrenal insufficiency due to AML\u2010induced adrenal infarction in a patient who presented with diffuse adrenal enlargement on CT imaging.22.1A 49\u2010year\u2010old healthy Japanese man without a history of regular health check\u2010ups or hospital visits presented with a 2\u2010month history of general fatigue and constipation. He felt discomfort in the left side of his abdomen while sleeping 16\u2009days previously, and the discomfort gradually developed into pain and worsened. Although abdominal CT at another hospital revealed adipose tissue turbidity surrounding the left adrenal gland .2.5Although there is no established treatment for AML with myelodysplasia\u2010related changes,2.6Eventually, because of the improvement of adrenal function, hydrocortisone administration was discontinued on Day 26 at a final dosage of 30\u2009mg/day. Abdominal contrast\u2010enhanced CT imaging performed on Day 35 revealed an improvement in the bilateral adrenal enlargement. The previously observed areas of poor contrast enhancement were no longer present, and the surrounding fat tissue turbidity had also improved. However, subsequent treatment for AML showed limited effectiveness. On Day 40, the standard treatment regimen involved the administration of idarubicin at a dose of 18\u2009mg for 3\u2009days and cytarabine at a dose of 150\u2009mg for 7\u2009days. On Day 161, allogeneic peripheral blood stem cell transplantation was performed, resulting in the achievement of remission. However, the patient died 20\u2009months after the diagnosis.3The present report describes an uncommon case of acute abdomen following adrenal infarction caused by AML. Although adrenal diseases such as adrenal crisis and adrenal infarction can cause acute abdomen manifesting as acute back, chest, and upper abdominal pain,Our patient had adrenal insufficiency caused by adrenal infarction. The major causes of adrenal insufficiency are autoimmune diseases and infectious diseases such as tuberculosis, cytomegalovirus, and human immunodeficiency virus. Notably, however, adrenal infarction and hemorrhage  can also cause adrenal insufficiency, as in this case.Typical symptoms and findings of adrenal insufficiency, including adrenal crisis, are hypotension, shock, weight loss, fatigue, appetite loss, nausea and vomiting, diarrhea, abdominal pain, hypoglycemia, hyponatremia, elevated urea nitrogen, hyperkalemia, and eosinophilia.4AML can cause acute abdomen following adrenal insufficiency or adrenal infarction. Therefore, when diffusely enlarged adrenal glands and adrenal insufficiency of unknown cause are seen in a patient presenting with acute abdomen, adrenal infarction due to AML, or other hematologic diseases should be ruled out.Risa Hirata: Conceptualization; writing \u2013 original draft. Masaki Tago: Conceptualization; project administration; supervision; writing \u2013 original draft; writing \u2013 review and editing. Shun Yamashita: Writing \u2013 original draft. Satsuki Yamamoto: Writing \u2013 original draft. Shizuka Yaita: Writing \u2013 original draft. Yuka Hirakawa: Writing \u2013 original draft. Maiko Ono: Conceptualization; writing \u2013 original draft; writing \u2013 review and editing. Shu\u2010ichi Yamashita: Writing \u2013 original draft; writing \u2013 review and editing.There is no funding for this article.The authors state that they have no conflict of interest.This manuscript conforms to the provisions of the Declaration of Helsinki in 1995 (as revised in Brazil 2013).Written informed consent was obtained from the patient's wife to publish this report in accordance with the journal's patient consent policy."}
+{"text": "Polyacrylonitrile (PAN), with its unique chemical, electrical, mechanical, and thermal properties, has become a crucial acrylic polymer for the industry. This polymer has been widely used to fabricate ultrafiltration, nanofiltration, and reverse osmosis membranes for water treatment applications. However, it recently started to be used to fabricate thin-film composite (TFC) and fiber-based forward osmosis (FO) membranes at a lab scale. Phase inversion and electrospinning methods were the most utilized techniques to fabricate PAN-based FO membranes. The PAN substrate layer could function as a good support layer to create TFC and fiber membranes with excellent performance under FO process conditions by selecting the proper modification techniques. The various modification techniques used to enhance PAN-based FO performance include interfacial polymerization, layer-by-layer assembly, simple coating, and incorporating nanofillers. Thus, the fabrication and modification techniques of PAN-based porous FO membranes have been highlighted in this work. Also, the performance of these FO membranes was investigated. Finally, perspectives and potential directions for further study on PAN-based FO membranes are presented in light of the developments in this area. This review is expected to aid the scientific community in creating novel effective porous FO polymeric membranes based on PAN polymer for various water and wastewater treatment applications. The forward osmosis (FO) process has received considerable research attention in recent years and has been successfully applied in numerous applications, including desalination of seawater and brackish water, wastewater treatment and purification, the concentration and separation of food and pharmaceuticals, as well as other new fields ,2. This The membrane is a crucial component of the FO process. The membrane modules, and membrane properties play a major role in FO performance and applicability. Selecting proper production and modification procedures can result in high-performance membranes. The optimum membrane for the FO water treatment process must have high water permeability and solute retention, as well as excellent chemical stability, high mechanical strength, low concentration polarization, and low fouling tendency. Thus, achieving all these desired properties has become an important research focus of FO membrane development by various research groups ,6,7,8. TOver the past few years, numerous research studies by both academia and industry have been conducted on FO membrane fabrication and modification, using various polymers to achieve the best FO separation and selectivity performance. The most extensively utilized polymers for the preparation of the FO membrane support layer include cellulose acetate/triacetate (CA/CTA) , polyvinAmong various polymers, PAN polymer, with its unique characteristics such as low material cost, outstanding physicochemical properties, and wide commercial availability, has sparked a lot of research interest in FO membrane fabrication at the laboratory scale . Most ofTo the best of our knowledge, no comprehensive review of the use of PAN-based membranes in FO operation tests has been conducted. Therefore, this work aims to review all of the PAN-based porous FO membranes that were previously used, considering their fabrication, modification techniques, and FO performance results. Our research is anticipated to be beneficial to this field\u2019s researchers by providing new avenues for the successful development of new porous FO polymeric membranes based on PAN polymer.Osmosis is a natural process that experts in many fields of science and engineering have examined in depth. The osmosis phenomenon applied to natural materials was first examined by researchers, and starting in the 19th century, osmosis received particular attention . OsmosisIn FO, two solutions, a concentrated draw solution and a more diluted feed solution, are positioned within a semipermeable membrane . FO, in Several drawbacks of other membrane technologies using pressure, for example, RO, can be addressed by FO through utilizing the osmotic pressure differential to stimulate water through the membrane . There aOn the other hand, the membrane represents a crucial part of the FO process, and its properties could affect process efficiency . It can Depending on the applications for the forward osmosis process, fabrication methods and membrane materials can be altered and improved. Desalination and wastewater treatment are the two principal uses of forward osmosis . DifferePolyacrylonitrile has been widely used as a substrate for various membranes like UF, microfiltration (MF), NF, and RO because of its good physiochemical properties such as high chemical stability, solubility to common solvents , dioxanone, chloroacetonitrile, dimethyl phosphite, etc.), and high hydrophilicity in comparison to other membrane materials such as polysulfone, polyethersulfone, polyethylene, and polypropylene ,59. ThisPAN is a relatively active polymer, which makes it easy to alter and modify. As shown in Plasma treatment is based on the use of a lower ionization level and is generally referred to as a \u2018cold\u2019 plasma process due to the fact that it utilizes inert gases , where rGraft polymerization is one of the least used methods for PAN surface modification due to its complexity ,80. The The hydrolysis technique using an alkaline solution is considered one of the simplest and cheapest methods applied to PAN membranes due to its ability in facilitating the next modification processes and increasing PAN chemical stability in common solvents ,62,66. B6 orders of magnitude, leaving the reaction to be completely orthogonal. Other click chemistry reactions that have been used to modify the surface of PAN membranes include the thiol\u2013ene reaction and the strain-promoted azide\u2013alkyne cycloaddition (SPAAC) reaction. This reaction can be used to attach a wide range of functional groups to the surface of the membrane, allowing it to be customized for specific applications [Another different method of altering the PAN membrane\u2019s surface is through the use of \u201cclick chemistry\u201d, which refers to a set of chemical reactions that are fast, selective, and easy to perform. Click chemistry is an emerging technique as an alternative to hydrolysis and grafting treatments. The click reaction has been observed to be a quantitative and orthogonal reaction that is wide in scope, generating a result in linking byproducts that does not affect the polymers. The click reaction has three different methods of modification. First, diazo reagents imidiazole-1-sulfonyl azide hydrochloride, with either a basic copper-free condition or a copper-catalyzed condition, are used as transfer reagents. Both methods result in the conversion of amino groups on the surface to azides. Secondly, carboxyl groups on the polymersomes are activated by the reaction of EDC-NHS in the presence of amino-PEG-monoazide to form amine-terminated polymersomes. Immobilization of azide-functionalized polymersomes on the membranes converts alkyne groups to triazoles ,90. A clThe preparation of hydrophilic membranes frequently involves surface alterations, such as physical coating and chemical modification. Chemical and physical coatings have been used to study the characteristic traits affecting the morphology and performance of the membrane. Qin et al.  studied 2 is used as the pervaporation separation layer on the PAN membrane, which reduces membrane swelling and has high stability. The second layer on top of the pervaporation layer is the catalytic layer, which is the main esterification reaction. Alcoholic groups are reacted with acidic groups in the presence of a catalyst, which forms the catalytic layer and completes the surface modification [Esterification is another type of membrane surface modification technique . This tefication ,92. ThisChemical modification is the most commonly used surface modification technique, among which hydrazine hydrate is used as a cross-linking mechanism in the PAN membrane ,94. HydrOne of the most recent advancements in PAN-based membranes is their application in the FO process, in which PAN was employed as a polymer to fabricate the substrate of simple TFC membranes and nanofiber (NTFC) membranes by the phase inversion (casting) method and electrospinning method, respectively, as shown in Most of the studies have focused on utilizing PAN polymer in fabricating flat sheet and hollow fiber FO membranes via the nonsolvent-induced phase separation method (NIPS) rather than fabricating FO nanofiber membranes by electrospinning, as illustrated in A conventional TFC membrane based on casted PAN substrate and PA was prepared by Klaysom et al. , in whic2 as an FS and DS, respectively [2 and 0\u201310 mM NaCl as DS and FS, respectively [Furthermore, other studies have prepared a porous matrix substrate by incorporating nanoparticles with PAN polymer during the phase inversion method. It was proven that this technique has a high potential for increasing water flux and controlling ICP in osmotically driven membrane processes. For instance, the prepared metal\u2013organic framework (MOF)-PAN mixed porous matrix substrate has achieved a high FO water flux of 132 LMH using deionized water (DI) and 3.0 M MgClectively . While tectively ,108. Theectively . As a reectively . This saectively .Another method for incorporating NPs into PAN-casted substrates is during the IP process, where the NPs are embedded into one of the PA layer phases. This technique is considered one of the most effective ways to optimize the IP process of hydrophilic PAN support, which has a high affinity for the amine monomer required for the conventional PA selective layer IP reaction . This afResearchers also discovered that when an aromatic hydrocarbon solvent (toluene) is employed as an organic phase in the IP process instead of aliphatic hydrocarbons, a highly selective PA layer may be formed on hydrophilic support . This to2 reverse flux of 7.5 gMH using 0.5 M MgCl2 as the DS. [A layer-by-layer assembly technique using oppositely charged polyelectrolytes through electrostatic interaction to generate a thin selective film with a controlled structure at the nanoscale has been offered by several researchers as an alternate approach for producing high-performance casted PAN-based FO membranes ,123,124. the DS. . HoweverFor further enhancement of casted PAN-based FO membranes, a few works have been reported to enhance their chlorine and antimicrobial resistance through chemical grafting and surface coating. Li et al.  have graAccording to the great separation performance of TFC-casted PAN FO membranes, as stated above, they were utilized as a post-treatment in a study by D. Kwon et al. , treatedIt is worth noting that for improving membrane performance as well as achieving a desirable membrane structure and morphology, it is crucial to control the kinetic and thermodynamic mechanisms of phase separation. A detailed effect of both mechanisms has been discussed by Ahmad et al. . It was Electrospinning is a versatile and rapidly evolving technique for the fabrication of nanofiber-based membranes. PAN has received much research attention for fabricating nanofiber membranes with highly porous layers due to its excellent compatibility, good electrospinnability, and high tensile strength after electrospinning, as shown in The promised osmotic performance with a finely tuned pore structure of the support layer was achieved using hydrophilic PAN polymer for fabricating TFC hollow fiber membranes, via dry jet\u2013wet spinning . The preIt has been noticed that preparing PAN nanofiber by electrospinning is not suitable for the IP process directly. As a consequence, some researchers have improved the binding strength of PAN nanofibers with a PA selective layer, using a thin chitosan (CS) sublayer that was cast on top of hydrolyzed electrospun PAN nanofiber before carrying out the PA formation ,143,144.w = 12.6 LMH and Js = 11.6 gMH) [w = 13.8 LMH and Js = 8.83 gMH) [For more FO membranes enhancement, several studies have fabricated a new class of TFC-FO membranes via blending hydrophilic PAN polymer with another polymer material to merge their beneficial characteristics and create new electrospun nanofiber substrates. For example, through the electrospinning technique, polyvinylidene fluoride/polyacrylonitrile (PVDF/PAN) electrospun nanofiber was fabricated by Kallem et al.  and used1.6 gMH) . While P.83 gMH) . Unlike .83 gMH)  have des.83 gMH) . Further.83 gMH)  have inc.83 gMH)  during t.83 gMH) , natural.83 gMH) , polydop.83 gMH) , and gra.83 gMH) , have be.83 gMH)  have emb.83 gMH) . The sam.83 gMH) . This ne.83 gMH) .2 were used as the FS and DS, respectively [In the literature, very little work has been carried out on developing commercial polyacrylonitrile membranes to prepare a thin-film composite forward osmosis membrane, as shown in ectively . TherefoMost of the PAN-porous-based FO membranes were fabricated at lab scale mainly through the phase inversion method, as indicated by 33 studies. On the other hand, few membranes were prepared by electrostatic spinning to prepare the PAN nanofiber support, and this is represented by 24 studies; however, the number of studies that have used commercial PAN-based UF membranes under the FO process was limited to four studies only. By comparing the FO performance of the three types of PAN-based membranes, electrospun PAN-based nanofibers membranes exhibit great potential to compete with conventional phase inversion PAN-based membranes. As shown in In addition, PAN nanofibers\u2019 extremely large surface area enables them to maximize surface functionalization and hybridization by chemical substances and nanostructures, respectively. As a result of their surface functionalization, the applications for these nanofibers will significantly increase. However, with respect to the salt rejection performance, the TFC-phase inversion PAN-based membranes were showing better rejection performance (>90%), as shown in It is worth mentioning that the different water flux values of the same type of PAN-based membrane are related to the modification technique procedure using various additives and nanomaterials as well as to the applied operation parameters and conditions, such as hydrodynamic conditions and flow direction of the FS and DS as driving forces of water transportation during the process operation. Polyacrylonitrile polymer, with its unique properties, has confirmed its ability to fabricate FO support membranes as well as its applicability to a variety of module types and consequently FO applications. This is because PAN is more easily modifiable than other polymeric materials used for membranes. In addition, PAN contains nitrile groups that enable a variety of chemical processes to enhance membrane structure and morphology, including hydrolysis, cyclization, and amination. Other than the polymer properties, the preparation conditions could also affect membrane structure and morphology. This paper has shown that different types of FO membranes have been fabricated based on the PAN polymer via phase inversion and electrospinning techniques. Numerous laboratory experiments have also been conducted to modify the physicochemical properties and performance of these PAN-based membranes, including membrane structure, permeability, mechanical strength, hydrophilicity, porosity, and surface charge, using various additives such as nanomaterials and polyelectrolytes during their fabrication stage or IP process. All lab-made PAN-based FO membranes have exhibited great potential for application under FO operating conditions at the lab scale. Their performances were described as achieving good water flux, comparable low reverse solute flux, high salt rejection, and high antifouling and mechanical strength. However, it should be noted that most of the nanofiber PAN-based membranes have demonstrated larger water permeable fluxes under the FO orientation mode compared to the phase inversion PAN-based membranes and even greater than those of commercial membranes in the range of (11\u201397 LMH). Although electrospun-PAN nanofiber-based substrates can provide significant potential in the FO process owing to their lower structural parameters and porous structure compared to PI PAN-based substrates, more research is required to improve their mechanical characteristics, optimize the pore size of ENs PAN substrates, and prevent the PA layer from delaminating from the ENs PAN substrates. Additionally, despite the significant increase in research on PAN-based FO membranes, their anti-fouling performance is still limited and needs to be addressed, as well as their performance at full-scale and pilot-scale systems, which still needs to be tested to better understand their FO performance."}
+{"text": "The recent legalization of sports wagering in many regions of North America has renewed attention on the practice of sports betting. Although considerable effort has been previously devoted to the analysis of sportsbook odds setting and public betting trends, the principles governing optimal wagering have received less focus. Here the key decisions facing the sports bettor are cast in terms of the probability distribution of the outcome variable and the sportsbook\u2019s proposition. Knowledge of the median outcome is shown to be a sufficient condition for optimal prediction in a given match, but additional quantiles are necessary to optimally select the subset of matches to wager on . Upper and lower bounds on wagering accuracy are derived, and the conditions required for statistical estimators to attain the upper bound are provided. To relate the theory to a real-world betting market, an empirical analysis of over 5000 matches from the National Football League is conducted. It is found that the point spreads and totals proposed by sportsbooks capture 86% and 79% of the variability in the median outcome, respectively. The data suggests that, in most cases, a sportsbook bias of only a single point from the true median is sufficient to permit a positive expected profit. Collectively, these findings provide a statistical framework that may be utilized by the betting public to guide decision-making. The practice of sports betting dates back to the times of Ancient Greece and Rome . With thA topic of obvious relevance to the betting public, and one that has also been the subject of multiple studies, is the efficiency of sports betting markets . While mon the side of the bettor. The classic paper by Kelly ). Examination of the sample quantiles suggests that NFL sportsbooks are very adept at proposing point totals that fall within 2.4 percentiles of the median outcome.The analysis was repeated for point totals . All butMaterials and Methods). The computation was performed separately within each stratified sample, and the height of each bar in when wagering on the team with the higher probability of winning against the spread. For the sake of clarity, only the four largest samples  are shown in the Figure, with data for all samples listed in In practice, it is desirable to have an understanding of how large of a sportsbook bias, in units of points, is required to permit a positive expected profit. To address this, the value of the empirically measured CDF of the margin of victory was evaluated at offsets of 1, 2, and 3 points from the true median in each direction. The resulting value was then converted into the expected value of profit (see \u03d5 \u2212 1)/2 = \u22120.045) when the spread equals the median (center column). Interestingly however, for 3 of the 4 largest stratified samples, a positive profit is achievable with only a single point deviation from the median in either direction . Averaged across all n = 21 stratifications, the expected profit of a unit bet is 0.022 \u00b1 0.011, 0.090 \u00b1 0.021, and 0.15 \u00b1 0.030 when the spread exceeds the median by 1, 2, and 3 points, respectively . Similarly, the expected return is 0.023 \u00b1 0.013, 0.089 \u00b1 0.026, and 0.15 \u00b1 0.037 when the spread undershoots the median by 1, 2, and 3 points respectively. This indicates that sportsbooks must estimate the median outcome with high precision in order to prevent the possibility of positive returns.The expected profit is negative . When the sportsbook underestimates the median, the expected profit on a unit bet is 0.015\u00b10.0071, 0.076\u00b1 0.014, and 0.14\u00b1 0.020, for deviations of 1, 2, and 3 points, respectively. Note that despite the dependent variable having a larger magnitude (compared to margin of victory), the required sportsbook error to permit positive profit is the same as shown by the analysis of point spreads.The analysis was repeated on the data of point totals. A deviation from the true median of only 1 point was sufficient to permit a positive expected profit in all four of the largest stratifications (Theorem 2), and if so, on which side (Theorem 1).The theoretical results presented here, despite seemingly straightforward, have eluded explication in the literature. The central message is that optimal wagering on sports requires accurate estimation of the outcome variable\u2019s quantiles. For the two most common types of bets\u2014point spread and point total\u2014estimation of the 0.476, 0.5 (median), and 0.524 quantiles constitutes the primary task of the bettor (assuming a standard commission of 4.5%). For a given match, the bettor must compare the estimated quantiles to the sportsbook\u2019s proposed value, and first decide whether or not to wager (Theorem 3). For a standard commission of 4.5%, the result is that if the sportsbook produces an estimate within 2.4 percentiles of the true median outcome, wagering always yields a negative expected profit\u2014even if consistently wagering on the side with the higher probability of winning the bet. This finding underscores the importance of not wagering on matches in which the sportsbook has accurately captured the median outcome with their proposition. In such matches, the minimum error rate is lower bounded by 47.6%, the maximum error rate is upper bounded by 52.4%, and the excess error rate (Theorem 4) is upper bounded by 4.8%.The sportsbook\u2019s proposed spread  effectively delineates the potential outcomes for the bettor  regression yields estimates of the mean of a random variable, conditioned on the predictors. This is achieved by minimizing the mean squared error between the predicted and target variable.The findings presented here suggest that conventional regression may be a sub-optimal approach to guiding wagering decisions, whose optimality relies on knowledge of the median and other quantiles. The presence of outliers and multi-modal distributions, as may be expected in sports outcomes, increases the deviation between the mean and median of a random variable. In this case, the dependent variable of conventional regression is distinct from the median and thus less relevant to the decision-making of the sports bettor. The significance of this may be exacerbated by the high noise level on the target random variable, and the low ceiling on model accuracy that this imposes.Therefore, a more suitable approach to quantitative modeling in sports wagering is to employ quantile regression, which estimates a random variable\u2019s quantiles by minimizing the quantile loss function . Any fea\u03d5h and \u03d5v do not substantially vary across matches. As a result, one can train a model on historical data to generate estimates of these canonical quantiles for future matches. Alternatively, one can develop a model to estimate the median and utilize it in conjunction with knowledge of how many points represents the requisite 2.4 percentile deviation. However, in the case of moneyline wagering, the payouts \u03d5h and \u03d5v do vary greatly across matches, meaning that one needs to estimate variable quantiles for different matches. This poses a challenge to predictive modeling for moneyline wagering, which will require estimating either very many quantiles or the entire distribution of the outcome variable. This seems to suggest a potential advantage of point spread and point total wagering: quantitative models can be trained to predict one or a few nominal quantiles, without the need to estimate the entire distribution of the outcome variable.Optimal wagering requires knowledge of the Theorem 4 is that estimators of the median outcome in sports betting need not be more precise than the sportsbook\u2019s proposition in order to achieve a positive expected profit. Rather, the goal of the statistical model is to produce estimates that yield sampling distributions with mass on the same side of the sportsbook proposition as the true median. Variations on this fundamental result have been previously presented in Reviewers' comments:Reviewer's Responses to Questions <font color=\"black\"> Comments to the Author1. Is the manuscript technically sound, and do the data support the conclusions? </font> The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented. Reviewer #1:\u00a0Partly********** <font color=\"black\"> 2. Has the statistical analysis been performed appropriately and rigorously?  </font> Reviewer #1:\u00a0No********** <font color=\"black\"> 3. Have the authors made all data underlying the findings in their manuscript fully available?PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data\u2014e.g. participant privacy or use of data from a third party\u2014those must be specified. </font> The Reviewer #1:\u00a0Yes********** <font color=\"black\"> 4. Is the manuscript presented in an intelligible fashion and written in standard English? </font> PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.Reviewer #1:\u00a0Yes********** <font color=\"black\"> 5. Review Comments to the Author </font> Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. Reviewer #1:\u00a0Report on \u201cStatistical Principles of Optimal Decision-Making in Sports Wagering\u201dThis paper analyzes the efficiency of betting markets for American professional football games from 2002-2022. The analysis includes bets on the margin of victory (point spread) and on the total number of points scored.In each case, bets are analyzed that are approximately even money bets. A casino specifies a point spread or point total, and the bettor can either choose the team to bet on (in the case of a point spread) or choose to bet that the total will higher or lower than the specified \u201cover/under\u201d. The most common arrangement is that a bettor wins the amount bet if they win but loses 1.1 times that amount if they lose. Therefore a bettor would need to win 11/21 = 0.524 or more of their bets over the long run to earn positive profits.The analysis is conducted non-parametrically. Games are placed in bins according to their point spread and again according to their point totals. Assuming that a bin ends up containing 100 games or more, the paper calculates the 47.6th and 52.4th percentiles of the outcome distribution and compares them to the \u201cconsensus\u201d point spread or point total offered by the casinos. This is equivalent to calculating the mean binary outcome for bets in each direction for the bin and comparing it to the profitability thresholds of 0.524 or 1 \u2013 0.524 = 0.476.The paper finds that many bins have sample means that are outside the 0.476 \u2013 0.524 range . It highlights this finding in the abstract: \u201capproximately two-thirds of matches permit a positive expected profit .\u201d Of course, sample means will vary around population means; with 100 binary outcomes, we would expect the difference between the sample mean and population mean to be approximately 0.5(1 \u2013 0.5)/sqrt(100) = 0.05. There is no analysis of whether we can reject a null hypothesis that the population means fall within the 0.476 \u2013 0.524 interval.Put another way, in order for a bettor to have earned a positive profit, they would have had to know in advance of the sample period which specific bins would yield outcome probabilities below 0.476 or above 0.524. The statement that \u201capproximately two-thirds of matches permit a positive expected profit\u201d assumes an ability to cherry pick bins after the fact.The general patterns in Figure 1, that home underdogs perform better relative to the point spread than home favorites and that outcomes tend to be less extreme than predicted by extreme over/unders, are well-documented in the literature. The literature in this area is quite vast (search \u201cNFL betting\u201d in Google Scholar to see what I mean). I do not find the incremental contribution of this paper to be compelling enough for PLOS-ONE.In this context, I should also mention that I find the claim in the abstract that \u201chere it is shown that sports wagering is effectively a problem of quantile estimation, with the median outcome having a crucial role in optimal decision making\u201d to be a bit strange. That the probability of an outcome being above or below a specified point spread or total, and therefore the quantiles of the distribution of the point spread or total, is central to the profitability of sports betting strikes me as completely obvious to everyone.********** <font color=\"black\"> 6. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.If you choose \u201cno\u201d, your identity will remain anonymous but your review may still be made public.Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy. </font> Reviewer #1:\u00a0No**********- - - - -For journal use only: PONEDEC3 7 Mar 2023Please see the \"Response to Reviewers\" document.AttachmentPONE-D-22-34727 - Response to Reviewers PONE-D-22-34727.pdfSubmitted filename: Click here for additional data file. 24 Apr 2023 <div> PONE-D-22-34727R1 </div>  <div> A statistical theory of optimal decision-making in sports wagering </div>  <div> PLOS ONEDear Dr. Dmochowski,Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE\u2019s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.We recommend that it should be revised by taking into account the changes requested by Reviewers. I want to give you a chance to revise your manuscript. The Academic Editor will only review the manuscript in the next round to speed the review process.plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.Please submit your revised manuscript by Jun 08 2023 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at\u00a0 </div> A rebuttal letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'.A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'.An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'. <div> If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.Please include the following items when submitting your revised manuscript:https://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols. Additionally, PLOS ONE offers an option for publishing peer-reviewed Lab Protocol articles, which describe protocols hosted on protocols.io. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols.If applicable, we recommend that you deposit your laboratory protocols in protocols.io to enhance the reproducibility of your results. Protocols.io assigns your protocol its own identifier (DOI) so that it can be cited independently in the future. For instructions see: We look forward to receiving your revised manuscript.Kind regards,Baogui Xin, Ph.D.Academic EditorPLOS ONEJournal Requirements: <span id=\"CommentsGrid_ctl02_shortText\"> When submitting your revision, we need you to  </span>  <span id=\"CommentsGrid_ctl02_longText\" style=\"\"> address these additional requirements.https://journals.plos.org/plosone/s/file?id=wjVg/PLOSOne_formatting_sample_main_body.pdf and https://journals.plos.org/plosone/s/file?id=ba62/PLOSOne_formatting_sample_title_authors_affiliations.pdf.1. Please ensure that your manuscript meets PLOS ONE's style requirements, including those for file naming. The PLOS ONE style templates can be found athttps://journals.plos.org/plosone/s/materials-and-software-sharing#loc-sharing-code and ensure that your code is shared in a way that follows best practice and facilitates reproducibility and reuse.2. Please note that PLOS ONE has specific guidelines on code sharing for submissions in which author-generated code underpins the findings in the manuscript. In these cases, all author-generated code must be made available without restrictions upon publication of the work. Please review our guidelines at http://journals.plos.org/plosone/s/data-availability.3. In your Data Availability statement, you have not specified where the minimal data set underlying the results described in your manuscript can be found. PLOS defines a study's minimal data set as the underlying data used to reach the conclusions drawn in the manuscript and any additional data required to replicate the reported study findings in their entirety. All PLOS journals require that the minimal data set be made fully available. For more information about our data policy, please see http://journals.plos.org/plosone/s/data-availability#loc-recommended-repositories. Any potentially identifying patient information must be fully anonymized.Upon re-submitting your revised manuscript, please upload your study\u2019s minimal underlying data set as either Supporting Information files or to a stable, public repository and include the relevant URLs, DOIs, or accession numbers within your revised cover letter. For a list of acceptable repositories, please see http://journals.plos.org/plosone/s/data-availability#loc-unacceptable-data-access-restrictions. Note that it is not acceptable for the authors to be the sole named individuals responsible for ensuring data access.Important: If there are ethical or legal restrictions to sharing your data publicly, please explain these restrictions in detail. Please see our guidelines for more information on what we consider unacceptable restrictions to publicly sharing data: We will update your Data Availability statement to reflect the information you provide in your cover letter.http://journals.plos.org/plosone/s/data-availability#loc-unacceptable-data-access-restrictions.In your revised cover letter, please address the following prompts:a) If there are ethical or legal restrictions on sharing a de-identified data set, please explain them in detail  and who has imposed them . Please also provide contact information for a data access committee, ethics committee, or other institutional body to which data requests may be sent.b) If there are no restrictions, please upload the minimal anonymized data set necessary to replicate your study findings as either Supporting Information files or to a stable, public repository and provide us with the relevant URLs, DOIs, or accession numbers. For a list of acceptable repositories, please see http://journals.plos.org/plosone/s/data-availability#loc-recommended-repositories.We will update your Data Availability statement on your behalf to reflect the information you provide.4. We note that you have indicated that data from this study are available upon request. PLOS only allows data to be available upon request if there are legal or ethical restrictions on sharing data publicly. For more information on unacceptable data access restrictions, please see http://journals.plos.org/plosone/s/latex. </span> 5. Please update your submission to use the PLOS LaTeX template. The template and more information on our requirements for LaTeX submissions can be found at Please review your reference list to ensure that it is complete and correct. If you have cited papers that have been retracted, please include the rationale for doing so in the manuscript text, or remove these references and replace them with relevant current references. Any changes to the reference list should be mentioned in the rebuttal letter that accompanies your revised manuscript. If you need to cite a retracted article, indicate the article\u2019s retracted status in the References list and also include a citation and full reference for the retraction notice.Additional Editor Comments (if provided):[Note: HTML markup is below. Please do not edit.]Reviewers' comments:Reviewer's Responses to Questions <font color=\"black\"> Comments to the Author </font> 1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the \u201cComments to the Author\u201d section, enter your conflict of interest statement in the \u201cConfidential to Editor\u201d section, and submit your \"Accept\" recommendation.Reviewer #2:\u00a0All comments have been addressedReviewer #3:\u00a0(No Response)********** <font color=\"black\"> 2. Is the manuscript technically sound, and do the data support the conclusions? </font> The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented. Reviewer #2:\u00a0YesReviewer #3:\u00a0Yes********** <font color=\"black\"> 3. Has the statistical analysis been performed appropriately and rigorously?  </font> Reviewer #2:\u00a0YesReviewer #3:\u00a0Yes********** <font color=\"black\"> 4. Have the authors made all data underlying the findings in their manuscript fully available?PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data\u2014e.g. participant privacy or use of data from a third party\u2014those must be specified. </font> The Reviewer #2:\u00a0YesReviewer #3:\u00a0Yes********** <font color=\"black\"> 5. Is the manuscript presented in an intelligible fashion and written in standard English? </font> PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.Reviewer #2:\u00a0YesReviewer #3:\u00a0Yes********** <font color=\"black\"> 6. Review Comments to the Author </font> Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. Reviewer #2:\u00a0Please remove the heading \"Results\" prior to problem formulation. All the subheadings with a question mark should be revised as normal heading (page 6-7). Subsections should be named without any a punctuation mark . Materials and methods should be placed as appendix. Discussion should be named as discussion and conclusion.Reviewer #3:\u00a0For a better overview, please find my comments also attached as pdf.Review PONE-D-22-34727R1Thank you for giving me the opportunity to review this manuscript. I would like to underline that \u2013 in my opinion \u2013 the manuscript has merit and I am very confident that it will be worth publishing if several revisions are made to the manuscript.Positive aspectsI will only very briefly mention the positive aspects of the manuscript as this review is intended to put more effort on possibilities for improvement. However, I would like to underline that I really enjoyed reading the manuscript. I like the fact that theoretical considerations are combined with empirical data. Moreover, it is interesting to see a manuscript in the domain of forecasting/sports betting that is able to answer relevant questions without the use of a concrete forecasting model. The theory is explained intuitively and is easy to follow, the results are well-explained and graphically represented. I\u2019d like to express my respect for the effort done by the author with regard to this manuscript. Please see my several critical comments as an effort to further improve the manuscript.ShortcomingsMy major point of criticism is the integration of the present results into the existing literature. Some claims are too strong and overselling the results, moreover in several parts I\u2019d strongly suggest to consider additional relevant literature. Please find more details on this point belowAbstract: I find the claim \u201cthe principles governing optimal wagering have not been presented\u201d way too strong. Please be more careful and precise here. Please also find more information on this point below.p. 1 Introduction l. 5: The author states \u201cimportant insights into market efficiency\u201d. In my mind this is simplifying as the results of market efficiency papers can point into different directions. So I would suggest to be more careful here by stating that market efficiency has been the subject of investigation or more precise by saying what the important insights were. Moreover, the author states three papers from 1968, 1997 and 2004. Below, I have summarised some more recent papers that might be worth considering:Angelini, G., & De Angelis, L. (2019). Efficiency of online football betting markets. International Journal of Forecasting, 35(2), 712-721.Bernardo, G., Ruberti, M., & Verona, R. (2019). Semi-strong inefficiency in the fixed odds betting market: Underestimating the positive impact of head coach replacement in the main European soccer leagues. The Quarterly Review of Economics and Finance, 71, 239-246.Meier, P. F., Flepp, R., & Franck, E. P. (2021). Are sports betting markets semistrong efficient? Evidence from the COVID-19 pandemic. International Journal of Sport Finance, 16(3).p. 1 Introduction 2nd paragraph: The author claims that literature on \u201coptimal decision-making from the bettor\u2019s perspective\u201d is missing. From my point of view, this is overselling the novelty of the current manuscript. While I certainly see what the manuscript adds to the literature, I would like to see acknowledged that decisions from a bettor\u2019s perspective have been considered in the literature explicitly and implicitly.Examples are the well-known Kelly betting strategy for optimal stake sizes going back toKelly, J. L. (1956). A new interpretation of information rate. the bell system technical journal, 35(4), 917-926.or literature focusing on the effects of non-optimal decisions of bettorsSnowberg, E., & Wolfers, J. (2010). Explaining the favorite\u2013long shot bias: Is it risk-love or misperceptions?. Journal of Political Economy, 118(4), 723-746.Moreover, it is well established in the forecasting literature to use and test several models for deciding on bets such as several stake sizes  in this example:Hvattum, L. M., & Arntzen, H. (2010). Using ELO ratings for match result prediction in association football. International Journal of forecasting, 26(3), 460-470.p.3 Theorem 2: I am surprised to not see the following paper cited.Wunderlich, F., & Memmert, D. (2020). Are betting returns a useful measure of accuracy in (sports) forecasting?. International Journal of Forecasting, 36(2), 713-722.For example, Theorem 2 is highly related to the area of no profitable bet presented in the aforementioned paper. In general, I see a lot of overlap between the two papers, both analysing betting decisions both from a theoretical point and based on real-world data. The author could also describe how the current manuscript is different from the aforementioned paper, e.g. by using point spreads and by analysing quantiles.p.8 Bias-variance in sports wagering: I really like the statement that bettors just need their estimation to be on the correct side of the spread, a fact that is often overlooked in profitable forecasting . You might discuss that the aforementioned paper of Wunderlich & Memmert and the paper of Hubacek & Sir below make similar statements.Hub\u00e1\u010dek, O., & \u0160\u00edr, G. (2023). Beating the market with a bad predictive model. International Journal of Forecasting, 39(2), 691-719.Further pointsTitle: Why do you use the wording \u201csports wagering\u201d? As far as I am concerned, this is pretty uncommon in the literature and \u2013 unless there is a specific reason that I am not aware of \u2013 I would rather expect the wording \u201csports betting\u201d.Results Problem formulation point spread: You mention the word point spread betting before giving an explanation on how such bets work. You might want to give a very brief explanation on this before, particularly as a lot of literature in this domain is concentrated on European sports betting markets, where point spreads are not that pronounced.p.2: Please define or introduce Phi_h, Phi_v is before the first usage.p. 4 Optimal estimation of the margin of victory: It is assumed that the difference between home and away team m and its estimation are independent. I am neither convinced that this is true nor convinced that this is false. But I am a bit sceptical as this is obviously a strong assumption needed for the further proof. Could you discuss this issue and explain in more detail why this assumption is reasonable?p.5 Optimality in moneyline wagering: At this point, again, I would suggest to add a reference to European betting markets, where bets without spread (i.e. s=0) are the most common bets. This is also reflected in the literature, which (for example in soccer) is highly concentrated on home, draw, away betting. I would also suggest to state possible differences between (European) home, draw, away and (North American) moneyline betting.Koopman, S. J., & Lit, R. (2019). Forecasting football match results in national league competitions using score-driven time series models. International Journal of Forecasting, 35(2), 797-809.Hvattum, L. M., & Arntzen, H. (2010). Using ELO ratings for match result prediction in association football. International Journal of forecasting, 26(3), 460-470.Constantinou, A. C., & Fenton, N. E. (2013). Determining the level of ability of football teams by dynamic ratings based on the relative discrepancies in scores between adversaries. Journal of Quantitative Analysis in Sports, 9(1), 37-50.\u2026among many many othersp. 6 Empirical results: I would strongly suggest to state the source of data where you obtained information on > 5.000 matches. Is it data provided by a company or data openly available online? On p. 9 Materials and Methods it appears that the source is the company usually not opening up on the data. However, I would like to see this explained more clearly.Same paragraph: This paragraph states means and medians from the data. While the manuscript generally correctly underlines the potential difference between mean and median (e.g. due to strongly skewed distributions), the numbers seem to suggest that the real-world distributions are only very weakly skewed and as such mean and median are closely related . I would highly like to see this aspect explained and acknowledged.Same paragraph: The paragraph says that \u201cThe standard deviation [\u2026] is nearly 7x the mean, indicating the frequent occurrence of outliers (\u201cblowouts\u201d). This claim, in my mind, is at least misleading. While I agree that a high standard deviation indicated frequent blowouts, this has nothing to do with the mean margin of victory which is rather an estimate of home advantage. If the data would show 0.12 + 14.68 instead of 2.19 + 14.68, would this be a sign of even more blowouts???p. 7 last paragraph: I really like that the author states that (besides correct forecasting) the bookmaker might have other incentives such as risk management . I wonder and I would like to see discussed whether there might be additional incentives of the bookmaker that contradict perfect forecasting. Just as one example, bookmakers might chose higher odds than reasonable for marketing reasons in some specific games. Might bookmakers in spread betting favour completely equal odds over slightly different odds, although not representing their true belief in the probabilities? To be very precise here, this is a true question from my side, i.e. I don\u2019t want to express that this is actually the case.p. 8 The case for quantile regression: The manuscript states \u201csubstantial deviation between the mean and median\u201c. This seems to be in contradiction to the data presented in the results section (see three points before). Please adjust this statement or give a better explanation on why you think this is the case.General point: Not related to any specific part of the manuscript, but you might want to discuss differences between American football and other sports. In terms of forecasting and statistical modelling American football is a very specific challenge as it has different possibilities to score  while other sports usually have only one possibility to score.********** <font color=\"black\"> 7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.If you choose \u201cno\u201d, your identity will remain anonymous but your review may still be made public.Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy. </font> Reviewer #2:\u00a0NoYes:\u00a0Dr. Fabian WunderlichReviewer #3:\u00a0**********https://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at\u00a0figures@plos.org. Please note that Supporting Information files do not need this step.While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool,\u00a0AttachmentReview PONE-D-22-34727R1.pdfSubmitted filename: Click here for additional data file. 6 Jun 2023Please refer to the attached PDF.Attachmentresponse_to_reviewers_060123.pdfSubmitted filename: Click here for additional data file. 8 Jun 2023A statistical theory of optimal decision-making in sports bettingPONE-D-22-34727R2Dear Dr. Dmochowski,We\u2019re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.Within one week, you\u2019ll receive an e-mail detailing the required amendments. When these have been addressed, you\u2019ll receive a formal acceptance letter and your manuscript will be scheduled for publication.http://www.editorialmanager.com/pone/, click the 'Update My Information' link at the top of the page, and double check that your user information is up-to-date. If you have any billing related questions, please contact our Author Billing department directly at authorbilling@plos.org.An invoice for payment will follow shortly after the formal acceptance. To ensure an efficient process, please log into Editorial Manager at onepress@plos.org.If your institution or institutions have a press office, please notify them about your upcoming paper to help maximize its impact. If they\u2019ll be preparing press materials, please inform our press team as soon as possible -- no later than 48 hours after receiving the formal acceptance. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact Kind regards,Baogui Xin, Ph.D.Academic EditorPLOS ONEAdditional Editor Comments :Reviewers' comments: 19 Jun 2023PONE-D-22-34727R2 A statistical theory of optimal decision-making in sports betting Dear Dr. Dmochowski:I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now with our production department. onepress@plos.org.If your institution or institutions have a press office, please let them know about your upcoming paper now to help maximize its impact. If they'll be preparing press materials, please inform our press team within the next 48 hours. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information please contact plosone@plos.org. If we can help with anything else, please email us at Thank you for submitting your work to PLOS ONE and supporting open access. Kind regards, PLOS ONE Editorial Office Staffon behalf ofProfessor Baogui Xin Academic EditorPLOS ONE"
\ No newline at end of file