California has led successful regionalized efforts for several time-critical medical conditions, including ST-segment elevation myocardial infarction (STEMI), but no specific mandated protocols exist to define regionalization of care. We aimed to study the trends in regionalization of care for STEMI patients in the state of California and to examine the differences in patient demographic, hospital, and county trends.Using survey responses collected from all California emergency medical services (EMS) agencies, we developed four categories – no, partial, substantial, and complete regionalization – to capture prehospital and inter-hospital components of regionalization in each EMS agency’s jurisdiction between 2005–2014. We linked the survey responses to 2006 California non-public hospital discharge data to study the patient distribution at baseline.STEMI regionalization-of-care networks steadily developed across California. Only 14% of counties were regionalized in 2006, accounting for 42% of California’s STEMI patient population, but over half of these counties, representing 86% of California’s STEMI patient population, reached complete regionalization in 2014. We did not find any dramatic differences in underlying patient characteristics based on regionalization status; however, differences in hospital characteristics were relatively substantial.Potential barriers to achieving regionalization included competition, hospital ownership, population density, and financial challenges. Minimal differences in patient characteristics can establish that patient differences unlikely played any role in influencing earlier or later regionalization and can provide a framework for future analyses evaluating the impact of regionalization on patient outcomes. Despite the relative decrease in the incidence of ST-segment elevation myocardial infarction (STEMI) over the past decade, STEMI still comprises about 25–40% of acute myocardial infarction incidents.3In an effort to optimize access to and delivery of care, policies from the American Heart Association (AHA) have advocated for establishing regional systems and networks.2Several regions in the United States have begun adopting innovative successful efforts to regionalize STEMI care, demonstrating feasible outcomes by integrating inter-hospital transfers and prehospital protocols;In California, EMS care is divided into 33 separate local EMS agencies (LEMSAs), and while all LEMSAs must have their EMS plans abide by general state EMS authority mandates, no specific mandated protocols exist to define regionalization of care.Previous work studying the effects of STEMI regionalization has not fully addressed demographic and clinical patient and hospital characteristics,What do we already know about this issue?Regionalization of care is a system that directs ambulances with STEMI patients to hospitals with PCI capabilities, which has been shown to improve treatment times.What was the research question?What are current trends in regionalization of care for STEMI patients in California?What was the major finding of the study?We found hospital and potential geographic differences between earlier and later regionalized counties.How does this improve population health?Our findings identify potential barriers to establishing regionalized systems of care, and may provide a framework for regionalization of care in other regions.International Classification of Diseases, Ninth Revision codes 410.0 through 410.6 and 410.8We collected survey responses from all California EMS agencies (details below), and linked the survey responses to 2006 California non-public hospital discharge data to understand the patient distribution at baseline. We obtained hospital information from annual surveys conducted by the AHA and annual reports submitted through the Healthcare Cost Report Information System maintained by the Centers for Medicare and Medicaid Services. We used county-level data from the U.S. Census for population demographics. We calculated the numbers of STEMI patients each year using Directors or STEMI coordinators from each EMS agency were asked to fill out a STEMI Regionalization Survey designed to evaluate the degree and duration of STEMI regionalization of care in each EMS jurisdiction. All 33 LEMSAs representing 58 counties in California responded and filled out the survey, resulting in a 100% response rate.Rokos et al.For the years evaluated, the survey assessed the degree of each EMS agency’s regionalized status by providing four multiple-choice options aimed at capturing the level of regionalization reached for each of the elements above: 1) Level A (none- 0%); 2) Level B (some- <50%); 3) Level C (most- 50–94%); and Level D . We developed four categories to capture both the prehospital (EMS devices and destination protocols) and inter-hospital  components of regionalization defined as follows: no regionalization; partial regionalization; substantial regionalization; and complete regionalization. In 2006, only eight out of 58 counties  had reached regionalization, with five counties (representing 13% of STEMI patients) considered completely regionalized . In 2011Of the counties that were not regionalized at baseline, 22 counties became partially regionalized, 12 became substantially regionalized, and 16 became completely regionalized in their first year of regionalization. The counties that became completely regionalized in their first year of regionalization generally were more populated than other counties. Not shown in the graph, a greater proportion of counties in Southern California reached complete regionalization by 2014, compared to Northern California, where the majority of counties reached only either partial or substantial regionalization. To collect the data we had to ensure the confidentiality of all counties and that no specific county would be identified in our results; thus, we are unable to provide maps of regionalization progress over time.At the hospital level, earlier regionalized counties had a higher proportion of for-profit hospitals (22% before 2006 v. 11% after 2011), while later regionalized counties had a higher proportion of teaching hospitals (14% after 2011 v. 8% before 2006) and hospitals part of a system (88% after 2011 v. 72% before 2006). Additionally, counties that regionalized by 2006 had a Herfindahl-Hirschman Index  mean of 0.14, indicating a low concentration of hospitals, 0.33 for counties regionalized by 2011, indicating a high concentration of hospitals, and 0.21 for counties that regionalized by 2014, indicating a moderate concentration of hospitals.At the county level, communities that regionalized later had a slightly higher mean per capita income  compared to those that regionalized earlier , consistent with earlier regionalization trends in communities with higher proportions of Medicaid-insured individuals. However, we found no apparent trends in the percentage of the population identified as living under the poverty line, part of a racial/minority group, or elderly (> 65 years).STEMI regionalization and its impact have been particularly well-documented in the literature for North Carolina,Our descriptive characteristics of hospitals show that counties with low concentrations of hospitals regionalized earlier than regions with high or moderate concentrations of hospitals, suggesting that competition may be a deciding factor in how quickly a county can regionalize. Generally, it has been well documented that competition between hospitals can prevent a region from reaching full regionalization.19Cardiac services such as PCI are considered one of the more profitable services a hospital can provide; therefore, it was not surprising when we found that generally, earlier regionalized counties had a greater percentage of for-profit hospitals. This could reflect the administrative and operational structural differences between for-profit and non-profit/government hospitals or the limitations facing non-profit/government hospitals to secure funding to become PCI-capable and regionalize. However, previous literature has suggested that our findings likely reveal the inclination of for-profit hospitals to offer PCI services and be motivated to regionalize faster as it would increase hospital revenue,Population density could potentially act as a barrier to earlier regionalization. Counties with denser cities may have a lesser need as the closest hospital likely is PCI-capable, even if the number of available PCI-capable hospitals is relatively small. On the other hand, counties with more spread-out cities may have a greater need as more residents are likely to be located nearest to a small hospital, without PCI capability. We found that Southern California counties generally regionalized faster as they geographically tend to be more “spread out,” which provides some evidence that population density could be a potential factor in adopting regionalization protocols. Our findings suggest that regionalization could be more beneficial in lower population density areas, especially in areas with relatively larger rural populations. Although high population density areas may benefit less from regionalization, they could be more susceptible to reduced PCI access due to crowding, and may benefit more from increasing the number of PCI-capable hospitals in the area or PCI capability within the existing hospitals.21Other potential factors that we did not specifically study but were mentioned by EMS administrators and medical directors may have contributed to the pace and degree to which each county regionalized. For instance, while regionalization of care provided the primary impetus to improve compliance with the 2004 AHA guidelines requiring a door-to-balloon time within 90 minutes,28Overall, examining trends in the regionalization of care has the potential to offer insight into how the reorganization of care can affect patient outcomes. These findings have important implications, as hospitals and EMS networks will require financial and organizational restructuring after adopting regionalization protocols. On a larger level, regionalized systems may allow providers and hospitals to benefit from reforms, such as the Medicare and CHIP Reauthorization Act of 2015,30Our descriptive characteristics establish that earlier and later regionalized counties do not differ dramatically in patient characteristics, and their differences would therefore not be expected to confound our later analysis of mortality outcomes. Furthermore, while hospital characteristics do vary considerably, previous literature suggests that although hospital ownership may influence the willingness and rate of regionalization, patient outcomes do not differ widely between for-profit hospitals and non-profit hospitals.Our study included several limitations. First, while we were able to receive responses to all questions from each EMS agency, we were only able to receive best approximate estimates in some responses of when each EMS agency reached a level of regionalization as in certain counties, the current administrator or medical director of the EMS agency might not have been present or have had knowledge of the state of cardiac care regionalization in 2005. Second, some directors and administrators could not provide records documenting the timeline of their protocols because these records did not exist or were erased. In these cases, we attempted to verify information by having external reviewers familiar with California’s regionalization trends examine the data and provide face validity checks for the provided responses. When questions arose for either limitation, we contacted the initial agency again and attempted to triangulate our information with other sources  to obtain more accurate information. Last, our data only covered California, meaning that while our findings may provide general insight into the regionalization of care, they may not be entirely generalizable to other states across the nation.Our survey results allowed us to identify prehospital and inter-hospital elements within each EMS agency’s STEMI program to assess the degree, duration, and trends of their regionalization-of-care efforts. We identified hospital competition, hospital ownership, population density, and financial barriers to be some potential factors in slowing down or preventing complete regionalization of care. We did not find any dramatic differences in the underlying population characteristics based on regionalization-of-care status in our study period, providing some reassurance for future studies evaluating the impact of regionalization that any findings would be less likely due to differences in patient characteristics. Our findings allow providers and policymakers to recognize the barriers to establishing and potentially reorganizing regionalized systems of care, and may serve as a framework for continued regionalization of care in other regions.


The TSLP forms can be produced by a number of cell types, including epithelial and dendritic cells (DCs). lfTSLP can activate mast cells, DCs, and T cells through binding to the lfTSLP receptor (TSLPR) and has a pro-inflammatory function. In contrast, sfTSLP inhibits cytokine secretion of DCs, but the receptor mediating this effect is unknown. Our recent studies have demonstrated that both forms of TSLP display potent antimicrobial activity, exceeding that of many other known antimicrobial peptides (AMPs), with sfTSLP having the strongest effect. The AMP activity is primarily mediated by the C-terminal region of the protein and is localized within a 34-mer peptide (MKK34) that spans the C-terminal α-helical region in TSLP. Fluorescent studies of peptide-treated bacteria, electron microscopy, and liposome leakage models showed that MKK34 exerted membrane-disrupting effects comparable to those of LL-37. Expression of TSLP in skin, oral mucosa, salivary glands, and intestine is part of the defense barrier that aids in the control of both commensal and pathogenic microbes.Thymic stromal lymphopoietin (TSLP) is a pleiotropic cytokine, hitherto mostly known to be involved in inflammatory responses and immunoregulation. The human  Thymic stromal lymphopoietin (TSLP) was first identified in the culture supernatant of a murine thymic stromal cell line and was shown to support B-cell growth and development . The humTwo variants of human TSLP peptides are expressed. Most studies hitherto have been focused on a long form of TSLP (lfTSLP), while translation of a short form (sfTSLP) has been reported only recently . lfTSLP Three transcript variants of human TSLP are annotated in the RefSeq (Reference Sequence) database , two long and one shorter variant, but only one of the long (variant 1) and the short (variant 2) variants give rise to coding RNA. Variant 1 consists of four exons and variant 2 is two 5′ exons shorter but contains an alternate 5′ exon compared to variant 1 A. The reThe long form (lfTSLP) encodes a 159 amino acid (aa) protein. The short form (sfTSLP) encodes a sequence that is identical in the C terminal region of long TSLP and consists of 63 aa (UniProt entry: G3XAM8) and/or a 60 aa (UniProt entry: Q96AU7) B. UniProThe N-terminal sequence of sfTSLP also contains a potential N-terminal signal sequence of 20 aa . The MWUse of variant-specific reagents is necessary to study the expression of the two human TSLP variants separately. At the mRNA level, this differentiation can be obtained by the use of variant-specific primers, which are constructed based on unique mRNA sequences in the two transcript variants. Detection of variant-specific protein expression, however, requires an indirect approach. As there is a total overlap of sfTSLP with the lfTSLP amino acid C-terminal sequence, antibodies raised against sfTSLP epitopes will recognize both variants and the production of sfTSLP-specific antibodies is, therefore, not possible. On the other hand, it is possible to generate antibodies specific for lfTSLP, either by immunization with peptide sequences that lie within the specific lfTSLP sequence, or by retrieving monoclonals that recognize such sequences. Thus, distinguishing between sfTSLP and lfTSLP has to be overcome by comparing the combined content of lfTSLP and sfTSLP in samples, using one antibody that recognizes both forms, and another antibody that binds to the unique sequence of lfTSLP (a long-specific anti-TSLP antibody). This approach has been used both for Western blotting and immunohistology ,6 Figur.Salmonellatyphimurium, shows upregulation of both mRNA and protein lfTSLP expression . U,6. U5,6]The divergent expression pattern for the two translated TSLP variants is consistent with the analysis of the human TSLP locus that reveals that the two variants are not alternatively spliced, but are derived from the activity of two separate, putative promotor regions . The sfTThe expression and regulation pattern of TSLP in mice overlaps to a large extend that of human lfTSLP. A role of TSLP in human allergic diseases is well supported by a variety of mouse models ,20,21,22+ DCs  and the TSLP receptor chain (TSLPR) ,26. The + DCs  [5,33,34+ T cells results in the generation of inflammatory Th2 cells producing classical Th2 cytokines including IL-4, IL-5, IL-13, but in contrast to conventional Th2 cells, these cells also produce TNF-α and not IL-10 [+ DCs potently augments allergen-specific TH2 memory responses [lfTSLP has an impact on several immune functions and has, as mentioned above, been associated with immune disorders, such as allergic diseases and intestinal inflammation. Co-culture of lfTSLP-stimulated DCs with allogeneic CD4ot IL-10 . This inot IL-10 ,35. Accoot IL-10 . In alleIn contrast to its role in inflammation, TSLP has also been suggested to have homeostatic, tolerogenic functions ,37. It wsfTSLP is constitutively expressed by several types of epithelial cells, as described above. sfTSLP appears to act on DCs on which it inhibits cytokine secretion . sfTSLP AMPs can be classified in a variety of approaches , fittingA common characteristic of AMPs is the propensity to form helical structure ,43. It hOur data displayed antimicrobial activity of TSLP against both bacteria and fungi ,7 FigurA,B. To fStreptococcus mitis, Escherichia coli, Enterococcus faecalis, Bacillus cereus, Staphylococcus epidermidis, and Candida albicans [S. mitis, reduced the antimicrobial activity by about half, showing that the reduction in colony-forming units per mL was specifically due to the action of sfTSLP [S. mitis showed that the effect of sfTSLP was stronger than that of LL-37  colonization at the human skin.AMPs are reported to possess different antibacterial spectrums. The well-characterized AMP LL-37 has a broad spectrum whereas psoriasin preferentially kills E. coli ,45. ConsE. coli , our finS. aureus and characterized by a chronic inflammatory infiltrate [P. aeruginosa elastase, S. aureus V8)  being derived from the C-terminal part and comprising 42 amino acids (Thr88-Lys129), encompassing the previously characterized synthetic MKK34 peptide. When the V8 degradation products were tested, the results showed similar antibacterial activity of the degradation product against the Gram-negative bacteria E. coli as the holoprotein TSLP [Several classical and recently discovered AMPs, such as LL-37 and AMPs derived from larger proteins, are generated by proteolytic processing resulting in bioactive fragments that exert antimicrobial effects ,47,48,49filtrate ,50. Therreus V8) . The S. E. coli, as well as on liposomes [AMPs are known to exert their effects by different mechanisms: some are membrane-active and others are not . To inveiposomes . MoreoveThe redox state of AMPs can be a determining factor for their activity: reduction of disulphide-bridges in human beta-defensin-1 (hBD-1) vastly potentiates its antimicrobial effect and free cysteines in the carboxy terminus seem important for the bactericidal effect . Of the Whether murine TSLP exhibits antimicrobial properties has not yet been investigated.Taken together, the latest studies on TSLP have shown (1) that human TSLP is translated in two forms; (2) that the short form is constitutively expressed in a steady-state, while the long form is absent; (3) that the long form is induced by inflammation, while the short form appears to be downregulated by inflammation; (4) that both forms exhibit potent AMP activity, with sfTSLP exhibiting the strongest activity; (5) that the AMP activity is primarily localized in the C-terminal part of both forms; and (6) that the C-terminal part of TSLP a has penetrating effect on bacterial membranes.As human sfTSLP is constitutively expressed at major barrier surfaces consisting of skin and mucosa, it is expected to play an important role against infection and regulation of inflammation at these sites. lfTSLP has a broader role in the defense against infection because it participates in the regulation of various immune activities, but it also preserves its antimicrobial functions.


Objectives To study the association between maternal use of antidepressants during pregnancy and autism spectrum disorder (ASD) in offspring.Design Observational prospective cohort study with regression methods, propensity score matching, sibling controls, and negative control comparison.Setting Stockholm County, Sweden.Participants 254 610 individuals aged 4-17, including 5378 with autism, living in Stockholm County in 2001-11 who were born to mothers who did not take antidepressants and did not have any psychiatric disorder, mothers who took antidepressants during pregnancy, or mothers with psychiatric disorders who did not take antidepressants during pregnancy. Maternal antidepressant use was recorded during first antenatal interview or determined from prescription records.Main outcome measure Offspring diagnosis of autism spectrum disorder, with and without intellectual disability.Results Of the 3342 children exposed to antidepressants during pregnancy, 4.1% (n=136) had a diagnosis of autism compared with a 2.9% prevalence (n=353) in 12 325 children not exposed to antidepressants whose mothers had a history of a psychiatric disorder . Propensity score analysis led to similar results. The results of a sibling control analysis were in the same direction, although with wider confidence intervals. In a negative control comparison, there was no evidence of any increased risk of autism in children whose fathers were prescribed antidepressants during the mothers’ pregnancy . In all analyses, the risk increase concerned only autism without intellectual disability.Conclusions The association between antidepressant use during pregnancy and autism, particularly autism without intellectual disability, might not solely be a byproduct of confounding. Study of the potential underlying biological mechanisms could help the understanding of modifiable mechanisms in the aetiology of autism. Importantly, the absolute risk of autism was small, and, hypothetically, if no pregnant women took antidepressants, the number of cases that could potentially be prevented would be small. Depression is common in women of childbearing age, and in Europe 3-8% of pregnant women are prescribed antidepressants during pregnancy.All antidepressants cross the placental barrier and are available to the developing fetus.In the absence of randomised controlled trials,We used data from the Stockholm youth cohort, an intergenerational record linkage study comprising all individuals aged 0-17 living in Stockholm County in 2001-11 (n=735 096). It contains prospectively recorded data on the cohort members and their first degree relatives collected by record linkage with a range of national and regional healthcare, social, and administrative registries.As mandated by the ethical permission, no attempts were made to contact any cohort members for any aspect of this record linkage study. As such, cohort members were not involved in setting the research question or the outcome measures, nor were they involved in developing plans for the design or implementation of the study. No patients were asked to advise on interpretation or writing up of results. There are no plans to directly disseminate the results of the research to cohort members, but dissemination to the general public and relevant patient groups will be undertaken by using presentations and social media.We derived information on maternal use of antidepressants in pregnancy from the medical birth register (since 1997) and supplemented it with the prescribed drug register (available from July 2005).We coded the medications as any antidepressant (ATC code N06A) and also divided them into selective serotonin reuptake inhibitors  and all other antidepressants. We also categorised antidepressants based on their affinities for the serotonin transporter into high versus medium or low affinity.We have described the multisource ascertainment of autism in previous publications.We used ICD-9 and 10 diagnoses recorded in the national patient register, which covers inpatient  and outpatient specialist care (since 2001) to ascertain depression and other maternal and paternal diagnoses of other psychiatric indications for antidepressants  any time before the birth of the child as described elsewhere.We used prospectively collected data on maternal and paternal age at birth of child (continuous variables used as restricted cubic splines), fifths of family income adjusted for year of ascertainment and family size, highest education of either parent , maternal country of birth , parity  as potential confounders. We also used several other variables to construct the propensity score as described below. These included maternal smoking and body mass index (BMI) recorded at first antenatal visit, number of diagnoses of maternal depression before birth, type of depression care (inpatient or outpatient), and a large range of maternal neurological and psychiatric conditions diagnosed before birth (see appendix).We used R-3.1.3  for analysis. After descriptive analyses, we used the following analytic strategies to assess the risk of offspring autism in mothers with antidepressant use during pregnancy.In our first analysis, we used logistic regression to derive odds ratios and their 95% confidence intervals as estimates of relative risks for autism in children of mothers who used antidepressants during pregnancy compared with those with a psychiatric disorder who did not use antidepressants. We used this stringent comparison group to better account for confounding by indication and because women without any psychiatric disorder would be ineligible in hypothesised randomised controlled trials of this issue. We adjusted the model for birth year to control for period effects in medication use and ascertainment of autism.In our second analysis, we calculated propensity score matched estimates for the above associations. Propensity score matching helps to minimise the potential for confounding from observed variables by comparing exposed and unexposed individuals with similar characteristics.In our third analysis, we examined associations between maternal antidepressant use in pregnancy and autism spectrum disorder in matched sets of outcome discordant siblings . Siblings share up to half of their genetic makeup and generally share their early postnatal environment. Sibling analysis can be a powerful method to control for unmeasured confounders, such as maternal genetic liability for neuropsychiatric conditions, when the confounders are shared more than the exposure.In our fourth analysis, we used paternal use of antidepressant as a negative control exposure with the assumption that the fathers would share many of the unmeasured confounders with mothers, and, if a similar heightened risk of autism was observable with paternal and maternal use of antidepressants, the associations would be unlikely to reflect an in utero effect of the medications.U increased the risk of autism and was more prevalent in antidepressant users than in non-antidepressant users. Given specified parameters such as the relation of U with autism spectrum disorder and the prevalence of U in antidepressant users and non-users, we estimated odds ratios and 95% confidence interval corrected for this unmeasured confounder, specified over a range of plausible parameters.In a sensitivity analysis, we estimated how robust the estimates of associations between antidepressant exposure and autism spectrum disorder were to unmeasured confounding in our propensity score matched sample. We assumed a binary confounder There were few missing data on the key variables in our main analysis . More data were missing in some variables used in the generation of propensity scores, but the boosted classification and regression tree model is able to incorporate missing values in the prediction (see appendix).Table 1Exposure to antidepressants during pregnancy was associated with a higher odds of a diagnosis of autism in offspring than exposure to a maternal psychiatric disorder without antidepressants . Tables C and D in the appendix show the rates and associations of autism in children exposed to the most commonly prescribed individual antidepressants, also grouped into SSRI and non-SSRI antidepressants as well as by their serotonin receptor affinity. Although imprecision because of small numbers is evident, the rates of autism seemed to be higher in children of mothers who used clomipramine and venlafaxine and lowest in users of paroxetine. Similar associations were observed for SSRI and non-SSRI antidepressants in relation to risk. The point estimates for the risk of autism in children of users of low/moderate SERT affinity seemed to be greater than that in children of users of high serotonin transporter affinity antidepressants, though the confidence intervals overlapped.In this large Swedish population based study, we carried out several analyses to further investigate the association between antidepressant use during pregnancy and autism in offspring. Our main findings were that children exposed to antidepressants during pregnancy seemed to be at a higher risk of autism, particularly autism without intellectual disability, than children of mothers with psychiatric disorders who were not treated with antidepressants during pregnancy. The findings seemed to be consistent across traditional regression methods, propensity score matching, and a sibling set comparison, and maternal exposure to antidepressants during pregnancy had a strong association with the outcomes whereas no such association was observed with paternal exposure. This points to a potential effect of antidepressant use on the risk of autism beyond any effect caused by confounding by the underlying condition. It is important to note, however, that the absolute risk was small, and 4.1% of children exposed to antidepressants in utero had autism compared with 2.9% of those with a maternal history of psychiatric disorder. We estimated that only about 2% of autism cases in this population would be theoretically prevented, if the association was causal and no women with psychiatric disorders used antidepressants during pregnancy.The main strengths of this study were the large sample size and the range of analyses carried out to strengthen causal inference beyond traditional methods. As the study included the total population of Stockholm County and benefited from multisource ascertainment of cases, as opposed to studies that rely on hospital discharge diagnoses, the findings are likely to have high external validity. The possibility of misclassification of exposure cannot be ruled out, but the availability of both self reported information from the medical birth register and dispensation information from the prescribed drug register was an advantage.This study builds on our previous case-control study,The different analyses we used have different strengths and limitations, but their findings seemed to triangulate, pointing towards an association between maternal antidepressant use in pregnancy and autism in offspring. On the other hand, the increased risk was seen largely for autism without intellectual disability, a phenotype that has been shown to be more heritable,So what should families and doctors making decisions about antidepressants during pregnancy make of such results? Firstly, this and other studies clearly suggest that there is an increased background risk of autism in children of women with psychiatric conditions, regardless of antidepressant treatment. Secondly, despite the observed relative risks, over 95% of women who took antidepressants during pregnancy did not have a child with autism. And, finally, if a causal link were robustly established, and if no pregnant women took antidepressants during pregnancy, only 2% of autism cases in this population would be prevented. It is known that pregnant women perceive such risks as greater than they are,Several observational studies have reported associations between antidepressant use during pregnancy and autism in offspringWhether this association is causal or confounded by indication is not clearThis large cohort study used various methods to deal with confounding, including traditional multivariable regression, propensity score matching, sibling controls, and a negative control designThe results of all these analyses, which used different assumptions, seemed to be consistent with each other, suggesting that the association between in utero exposure to antidepressants and autism might not be fully explained by confoundingThe absolute risks were small so results should not be considered alarming, but the findings could be useful in a further understanding of the aetiology of autism


The microspheres were prepared by multiple emulsion technique (water/oil/water) and characterized by the encapsulation efficiency, particle size, polydispersity index, and zeta potential. The acute toxicity of UA and UA-micro  was evaluated in mice. The anti-inflammatory activity of UA and UA-micro was evaluated by subcutaneous air pouch and carrageenan-induced paw edema in rat, with measurement of inflammatory cytokines and MPO levels. The UA presented encapsulation efficiency of 97.72%, particle size of 13.54 micrometers, polydispersity index of 2.36, and zeta potential of 44.5 ± 2.95 mV. The UA-micro presented lower acute toxicity  when compared to UA. UA-micro and UA (25 mg/kg) significantly reduced paw volume and decreased MPO levels, whereas only UA-micro (50 mg/kg) reduced significantly IL-1β, TNF-α, and NO levels in inflammatory exudate. These results suggest that controlled release systems, as microspheres, can be a promising alternative to reduce the toxicity of UA, making it a viable compound for inflammation therapy.Usnic acid (UA) has been studied by its pharmacological properties; however, it presents moderate toxicity, low solubility, and absorption by biological membranes. The aim of this study was to develop poly- Inflammation is a complex process involving several cell types and molecular mediators, whose main goals are to isolate, neutralize, and destroy harmful agents, following tissue regeneration stimuli . AlthougUsnic acid -benzofuran, C18H16O7), a secondary metabolite isolated from lichens , 5, has The controlled-release drug system was designed to slowly and continually release the drug, keeping the desired concentration in the blood for a longer period. One of the controlled-release drug systems is microspheres, which are polymeric solid particles  that deserve highlight due to physicochemical and biological stabilities. In addition, microspheres are utilized for encapsulation of several substances, such as peptides, proteins, DNA/RNA, and drugs , 14. EncMale Wistar rats (190–200 g) and female Swiss mice (30–35 g) from the vivarium of Antibiotics Department, Federal University of Pernambuco (UFPE), were used. Animals were kept in polypropylene boxes in a room with controlled temperature (22 ± 2°C) and light-dark cycle of 12 h/12 h. Water and food were available ad libitum. Experimental protocols were approved by the Animal Studies Committee (CEEA) of the UFPE, under process number 23076.052269/2012-35.ε-caprolactone, polyvinyl alcohol (MW: 30.000–70.000), polyethylene glycol (MW: 3.350), usnic acid, trehalose, indomethacin, and carrageenan type IV) were obtained from Sigma-Aldrich .All the reagents (poly-ε-caprolactone and 25 mg of usnic acid into dichloromethane (6 mL). The organic solution was emulsified  with 125 mg of polyethylene glycol into water for 1 min at 6848 ×g. The produced emulsion was added to an aqueous phase constituted by polyvinyl alcohol (1.25%) and was emulsified during 30 s at 6848 ×g, resulting in multiple emulsions. The final emulsion was kept under agitation at 17 ×g for 2 hours. Microspheres were centrifuged at 963 ×g for 10 minutes and dispersed into trehalose 1% (m/v). Subsequently, microspheres were frozen at −80°C and lyophilized afterwards.Microspheres were prepared using multiple emulsion method following solvent evaporation . One simμg·mL−1. Usnic acid was determined by spectrophotometry (280 nm) using a standard curve of 3–10 μg·mL−1 in acetonitrile.Usnic acid encapsulation efficiency in the microspheres was determined by spectrophotometry . Lyophilized microsphere samples (5 mg) were diluted in a dichloromethane : methanol (3 : 2) mixture and submitted to sonication for 10 min. An aliquot of this solution was diluted to a theoretical concentration of 6 d − d)/d, where d, d, and d, represent the percentage of microspheres with volume median diameter above 90%, 10%, and 50%, respectively.Microsphere particles size was determined through laser diffraction technique using a particle analyzer  . Lyophilζ) was measured after dilution into an aqueous solution of 1 Mm NaCl  at 25°C.Surface charge  was determined by electrophoretic mobility method using the Zetasizer Nano ZS90  equipment and Zetasizer software. Microsphere zeta potential . Experiments were repeated using the same animal quantity, with a total of 6 animals/group. The control group received vehicle  and experimental groups received usnic acid , poly-ε-caprolactone microsphere polymers containing usnic acid , or indomethacin . Animals were observed during the first 8 and 24 hours during 14 days after compound administration and had toxicity signals scored. On the last day of observation, animals were euthanized in a CO2 chamber and had their organs (liver and kidney) removed for histopathological analysis.Acute toxicity tests were designed following the “Acute Toxic Class Method” . Mice weμm. Histological slices were kept in an incubator at 37°C for 24 h in order to dry. Subsequently, slices were tinted with hematoxylin-eosin stain (H&E). Analysis was made using optical microscope.Necropsy of animals submitted to acute toxicity was performed and the organs (liver and kidney) were kept in neutral buffered formalin (10%) for 48 h at room temperature. Then, organs were submitted to dehydration using ethyl alcohol in different concentrations, diaphanized in xylol, impregnated, and included in paraffin. Samples were sliced into a microtome adjusted to 5 n = 5 animals/group) were treated orally with UA (25 mg/kg), UA-micro (25 and 50 mg/kg), indomethacin (10 mg/kg), or vehicle. One hour after treatments, 0.1 ml of a commercially prepared carrageenan was inoculated into the right hind of animals. The volume of the hind paw swelling (ml) was measured with an electronic water plethysmometer  at zero minutes (before) and 1, 2, 3, and 4 h after a carrageenan injection.Paw edema test was performed according to Winter et al. . Five WiμL of centrifuged supernatant and 200 μL of o-dianisidine dihydrochloride solution  following spectrophotometer reading (450 nm). Results were expressed in MPO units (UMPO/mg of tissue), considering that 1 UMPO corresponds to the quantity that one enzyme degrades with 1 μmoL per minute . Samples were centrifuged at 12.000 ×g for 2 minutes at 4°C. Each well in a 96-well plate was filled with 5 r minute . The tes2 chamber and had the pouch washed with 3 mL of PBS containing 3 μMol EDTA. Polymorphonuclear leukocytes quantification on the lavage fluid was performed using an ABX Micros 60 hematology analyzer [Anti-inflammatory activity was evaluated through air pouch formation by injection of 2.5 mL of sterile air on day 0 in the dorsal region of the mice, followed by a second injection of 2.5 mL of sterile air three days after the first. On the sixth day, according to their group, the animals received orally vehicle, UA , UA-micro (25 and 50 mg/kg), or indomethacin (10 mg/kg). One hour after treatments, inflammation was induced by injection of 1 mL carrageenan (1%) inside the air pouch. After 6 h, animals were euthanized in a COanalyzer , 21.μL was transferred into a microplate and incubated with 50 μL Griess reagent  for 10 min at room temperature in the dark. Absorbance (540 nm) was obtained using a microplate reader and nitrate concentration was calculated using a sodium nitrate standard curve [To access nitric oxide (NO) production, nitrate concentrations (NO stable metabolite) were quantified in the air pouch exudate through air pouch test. One sample aliquot of 50 rd curve .α and IL-1β quantification was accessed using specific ELISA kits for mice, following manufacturer's instructions . The lower limit of detection was 10 pg/mL.TNF-p” values lower than 0.05 (p < 0.05) were considered as indicators of statistical significance.Results were expressed as mean ± standard deviation for each experimental group using GraphPad Prism software . Data were analyzed through one-way Analysis of Variance (ANOVA) followed by Tukey's test, except for paw edema test, which was analyzed through two-way Analysis of Variance (ANOVA) followed by Bonferroni multiple comparison test. Confidence intervals of 95% and “y = 0.07309x + 0.00263, where y is the absorbance (nm) and x is the concentration (μg/mL) in usnic acid equivalents. The correlation coefficient was 0.99989, meaning that 99.98% of the total variation around the mean was explained by linear regression. Usnic acid was encapsulated into PCL microspheres with efficiency of 97.72 ± 0.0004%. UA-micro zeta potential (ζ) had a higher negative value (ζ = −44.5 ± 2.95 mV) when compared to PCL microspheres (ζ = −26.9 ± 0.58 mV)  .μm, while microspheres without usnic acid size had grain size of 9.37 μm  are presented in In control animals, no morphological kidney tissue damage was detected . The UA-Concerning hepatic tissue, no abnormalities were observed in control group . The UA-As can be seen in The treatment with UA , UA-micro , or indomethacin  significantly reduced leukocytes migration to injured paws . The neuα  microspheres (UA-micro) and free UA.Usnic acid, a secondary metabolite of lichen, is of great interest in pharmacological research. However, this compound has been associated with severe liver damage (hepatotoxicity) when taken as a dietary supplement. The anti-inflammatory activity of usnic acid has been described in the literature by several researchers using in vitro and in vivo assays , 23 , UA showed reduced toxicity degree and maintained its anti-inflammatory activity. The downregulated acute toxicity verified in UA-micro  can be explained by its incorporation into microspheres. According to Chen et al. [ε-caprolactone microparticles act as a drug delivery system, improving the therapeutic effect, prolonging the biological activity, controlling the drug release rate, decreasing the administration frequency, and so forth. These drug delivery systems protect certain labile active principles from degradation and/or inactivation by gastric juice, improve bioavailability by increasing the cellular penetration of substances, and provide release of the drug at the desired site of action, decreasing the toxic effects that normally accompany conventional therapy [Consistently, as incorporated into poly-n et al. , poly-ε- therapy . Regardi therapy  which re therapy  found th therapy . Accordi therapy , this ki therapy  study liα, and IL-1β [α, IL-1β, and NO levels in the inflammatory exudate from subcutaneous air pouch was also registered. TNF-α and IL-1β are related to increase of vascular permeability and migration of cells that produce chemokines [β, and TNF-α. Overall, the results of the current study indicate that equivalent or greater anti-inflammatory activity was observed as UA was incorporated into poly-ε-caprolactone microsphere polymers (UA-micro).The results in the present study indicated that free UA and UA-micro attenuated the carrageenan-induced rat paw edema and the levels of MPO  into injured paw. Neutrophil granules myeloperoxidase (MPO) is involved in reactive oxygen production, free radicals, and membrane oxidation, and tissue MPO levels become higher during inflammatory conditions, which are commonly related with bacterial lysis and tissue oxidative injury . Inflammnd IL-1β . In addiemokines . The resemokines , which semokines , which dε-caprolactone microsphere polymers (UA-micro) compared to free UA, with greatly reduced acute toxicity.UA is now shown to achieve equivalent or greater anti-inflammatory activity as it is incorporated into poly-


Consistently, cotton JAZ can partially rescue TOR-suppressed phenotypes in Arabidopsis. These evidences revealed that the crosstalk between TOR and JA pathway operates in cotton and Arabidopsis.Target of rapamycin (TOR) acts as an important regulator of cell growth, development and stress responses in most examined diploid eukaryotes. However, little is known about TOR in tetraploid species such as cotton. Here, we show that TORC1-S6K-RPS6, the major signaling components, are conserved and further expanded in cotton genome. Though the cotton seedlings are insensitive to rapamycin, AZD8055, the second-generation inhibitor of TOR, can significantly suppress the growth in cotton. Global transcriptome analysis revealed that genes associated with jasmonic acid (JA) biosynthesis and transduction were significantly altered in AZD8055 treated cotton seedlings, suggesting the potential crosstalk between TOR and JA signaling. Pharmacological and genetic approaches have been employed to get further insights into the molecular mechanism of the crosstalk between TOR and JA. Combination of AZD8055 with methyl jasmonate can synergistically inhibit cotton growth, and additionally JA levels were significantly increased when cotton seedlings were subjected to AZD8055. JA biosynthetic and signaling mutants including  Streptomyces hygroscopicus, and has aided a wealth of studies in animals and yeasts1Arabidopsis and Vicia faba, likely due to the altered structural aspects of FKBP12 (FK506 binding protein of 12 kDa) in the formation of functional TOR-rapamycin-FKBP12 ternary complex5Target of rapamycin (TOR) is a serine/threonine protein kinase and is evolutionally conserved from the last eukaryotic common ancestor (LECA) to humans1231FKBP12-based Arabidopsis transgenic plants (BP12-2) were produced, which confer conditional sensitivity to rapamycin89Arabidopsis seedlings exposed to AZD showed that TOR regulated photosynthesis and phytohormone signaling pathways including jasmonic acid (JA) signaling pathwayCompared to great progress made in the understanding of TOR signaling in animals and yeasts, relatively less is known about this ancient and important regulatory system in higher plants. To explore TOR functions using RAP in higher plants, yeast COI1 (Skp1/Cullin/F-box) complex that contains CORONATINE-INSENSITIVE1 (COI1) F-box protein and the transcript repressor JASMONATE-ZIM DOMAIN (JAZ) protein161718192021The phytohormone JA regulates a broad spectrum of biological processes, including cell growth and development, as well as defense responses to biotic and abiotic stresses12131314Gossypium hirsutum) provides the major resource of textile fiber worldwide. Environmental stresses are major threats to cotton crop yields. To deal with environmental stresses, crop plants like cotton frequently encounter yield penalties resulting from effects on growth and development. Relevant to these, TOR is involved in regulating plant growth and JA plays a key role in response to environmental stresses1Arabidopsis. Our results provide new evidences showing a connection between TOR and JA signaling pathways and demonstrate that TOR has a negatively effect on JA signaling pathway.Cotton  spans 20,012 bp and contains 58 exons, and GhTOR2 (Target of Rapamycin 2) has 21,569 bp with the same number of exons as observed in GhTOR1. GhTOR1 protein sequences showed similar organization of domains as Arabidopsis, humans and yeasts, whereas the FATC domain was not detected in GhTOR2 protein  analysis revealed that the expression of these two GhTOR genes can be detected with similar profiles in all cotton tissues , RPS6 , α4, E2F3 (E2F transcription factor 3), ATPBD3 (Repressor of LRR-extension 1), and TCTP ; however, the homologs of VPS34 (Vacuolar protein sorting 34), 4EBP1 (Translation initiation factor 4E-binding protein 1), and SGK1 (Serum/glucocorticoid-regulated kinase 1) were not detected (S6K (6 copies) and RPS6 (ten copies) have been found, suggesting the expansion of these genes in cotton genome compared to the other TOR components like α4 (1 copy), E2F3 (2 copies), ATPBD3 (1 copy) and TCTP (2 copies) in cotton -PI3K (Phosphoinositide 3-kinase)-PDK (3′-phosphoinositide-dependent protein kinase)-Akt-TSC1 (Tuberous sclerosis 1)/TSC2 (Tuberous sclerosis 2) signaling cascade is a crucial upstream regulatory input of TOR in humans126homologs , indicatdetected . It shoun cotton . The extGhFKBP12 in the cotton genome (http://cgp.genomics.org.cn). Only a single copy of FKBP12 (CotAD_57498) was present and was evolutionary conserved among species  and AZD were used for RNA-seq analysis. After trimming for quality, 87.77 and 96.96 million RNA-seq clean reads were obtained under the treatment of DMSO and AZD (representing the average of three replicates), respectively . The corNext, we examined whether the potential crosstalk between TOR and JA exists in cotton by applying MeJA (Methyl jasmonate), AZD, and a pairwise combination of MeJA + AZD to cotton seedlings . The rooArabidopsis to study the potential interactions between TOR and JA. RAP selectively inhibits TORC1-S6K signaling while AZD is a pan-TOR inhibitor which can directly interact with kinase domain of TOR by competing with ATP. These different properties of RAP and AZD may provide alternative and complementary approaches to investigate the crosstalk between TOR and JA in Arabidopsis. WT and BP12-278Due to the lack of mutant collections and difficulties in genetic transformation in cotton, we used the model plant To quantitatively measure the synergism generated by the interaction between TOR inhibitors and MeJA, CI (Combination index) values were used in this study. First, to further examine whether the synergism can be generated by the combination of TOR inhibitors (RAP or AZD) with MeJA, we investigated the GI50 (50% growth inhibitory dose) values of each drug alone and pairwise combinations in BP12-2 seedlings . The GI5AOC1 (Allene oxide cyclase1) and OPR3 (OPDA reductase3)32JAZ1 and JAZ71718Arabidopsis seedlingsCYCB1;1 MYC2Arabidopsis  and GhTIFY10a (JAZ10) showed a relative better growth condition under TOR inhibition than other transgenic lines, thus GhTIFY7b and GhTIFY10a were selected for further analysis  were generated and exposed to MeJA and TOR inhibitors , we identified the putative components of TOR complexes, the upstream and downstream factors of TOR signaling pathway for the first time. Unlike the FATC domain’s essential functions for TOR protein kinase in yeast and animalsArabidopsis, this domain appears dispensable and no clear phenotype can be observed with its absenceTOR genes and the canonical TOR components in cotton, which might point to new additional functions for TOR signaling.Most examined heterotrophic organisms contain two distinct TOR complexes: TORC1 and TORC2, and TOR signaling pathway has been well documented in several model organisms123Arabidopsis BP12-2 line was successfully applied to detect the functions of GhTIFYs in TOR signaling by using RAP. These results demonstrate that RAP and AZD have many overlapping and complementary advantages for TOR signaling research in cotton and Arabidopsis.Rapamycin and AZD were employed to probe TOR signaling in this study. They both target TOR protein, but the modes of action, molecular mechanism, inhibitory spectrum and drug potency of rapamycin are quite different from that of AZD. Rapamycin is the first generation TOR inhibitor and specifically inhibit TORC1-S6K signaling branch, whereas AZD is the second generation of TOR inhibitor and can suppress most outputs of TORC1 and TORC2. Since TOR has many upstream signaling inputs and downstream signaling outputs in eukaryotic organisms and different signaling cascades operated through these have different functions, both selective TOR inhibitor (rapamycin) and pan-TOR inhibitor (AZD) based approaches are useful for defining TOR signaling in plants. Rapamycin/FKBP12/TOR ternary complex provides a conditional and highly inducible selective system to decipher TORC1-S6K-RPS6 signaling. Considering TORC1-S6K-RPS6 is the mainstay of cotton TOR signaling, rapamycin should be a very important tool to dissect TOR signaling if GhFKBP12 is able to bridge the interaction between RAP and TOR. However, our observations suggest that GhFKBP12 is defective to interact with RAP and not effective to produce TOR inhibition. As an alternative, we have successfully employed AZD to probe TOR signaling in cotton; and the Arabidopsis, the expression profiles of representative JA related genes were also altered under TOR suppression by AZDArabidopsis, TOR inhibition can be partially rescued by a mutation in ABI4 (ABA Insensitive 4), which encodes an ABA-regulated AP2 domain transcription factorArabidopsis. In the DEGs of our cotton RNA-seq data, we didn’t find these two putative homologs, potentially either due to the loss of the evolutionary functional conservation for these genes or they do not represent the true orthologs. Besides, the transcript levels of the other key node genes of ABA signaling pathway were changed in cotton for instance PYL (PYRABACTIN resistance 1-like), ABF (Abscisic acid responsive elements-binding factor) and PP2C (Type 2C protein phosphatases) family protein  was used as the wild-type plants in this study. The transgenic Arabidopsis BP12-2 was generated in a previous studyArabidopsis mutant lines jar1, coi1-2, myc2-2 and jaz10 were kindly provided by Dr. Chuanyou Li of the Institute of Genetics and Developmental Biology, Chinese Academy of Sciences. Arabidopsis seeds were surface-sterilized with 70% ethanol for 3 min; followed by 10% bleach containing 0.3% Tween-20 for 10 min; and then rinsed by sterile water for three times. The sterilized seeds were suspended in 0.1% sterile agrose and kept at 4 °C for two days, and then germinated on 1/2 MS (Murashige and skoog) medium plates supplemented with reagents as indicated in Figures. All Arabidopsis were grown at 22 °C under long-light conditions (16 h light/8 h dark). Wild-type G. hirsutum (CCRI24) is used in this study. Cotton seeds were surface-sterilized with 0.1% mercuric chloride for 5 min, rinsed with sterile water. The sterilized seeds were sown in MS medium containing different solutions and 1% agar, and grown in a growth chamber at 25 °C under 16 h light/8 h dark condition. Plant growth showed no difference between MS and MS + DMSO medium, and all drugs were dissolved in DMSO, MS + DMSO medium was set as a control in this study.2s at 22 °C. Fresh weight of 6- and 10-day-old seedlings in every well was measured to calculate growth inhibition. Experiments were repeated three times. Percentage of growth value was calculated by using the fresh weight of 6-day-old seedlings (W6), DMSO-treated control plants (C), and drug-treated plants (D) as follows: [(D-W6)/(C-W6)] × 100. The 50% growth inhibitory dose (GI50) and CI values were calculated using the CompuSyn software programCombination index (CI) values were used to quantitatively measure the interaction between MeJA and TOR inhibitorsArabidopsis TOR and FKBP12 protein sequences were used as quests searching for cotton homologs in the cotton genome database (http://cgp.genomics.org.cn/)Arabidopsis TOR signaling pathways components were used to identify cotton homologs in the same database. The 21 G. hirsutum TIFY genes were identified by aligning the TIFY domain sequences with the cotton genome (http://cgp.genomics.org.cn/). E-value (1e–10) was set to obtain the final TIFY proteins. Phylogenetic tree was constructed by MEGA4.0 using the neighbor-joining method39http://meme.sdsc.edu/meme)Agrobacterium-mediated transformation using the floral dip methodFull-length CDS were amplified by PCR using the TransStar Taq Polymerase Mix kit (TRANSGEN) following the manufacturer’s instructions. The list of primers is shown in Arabidopsis seedlings as indicated in Figures using the RNAprep Pure Plant kit (TIANGEN). PrimeScript RT reagent kit (TAKARA) was used to generate first-strand cDNA products. The CFX96 real-time PCR system (BIO-RAD) was used to measure relative transcript expression levels by using an SYBR Premix Ex Taq II (TAKARA) kit. Primers used for QRT-PCR were listed in AtACTIN2 and GhHISTONE3 in Arabidopsis and cotton, respectively. The relative quantitative method (2−ΔΔCT) was used to calculate the quantitative variationTotal RNA was isolated from cotton or X-Gluc was used in GUS staining following a previously reported procedureArabidopsis or six-day-old cotton seedlings were transferred to 1/2 MS medium containing DMSO or various TOR inhibitors for 48 hours. Each treated Arabidopsis or cotton seedling was harvested for measurements of JA contents. JA was extracted following the exact extraction procedureTwo-week-old G. hirsutum transcripts with the help of genome annotation482Ratio| value ≥ 1 were identified as significant differentially expressed genes. KOBAS software was used to test the enrichment of DEGs in KEGG pathways524-d-old cotton seedlings were transferred into MS medium containing DMSO and 5 μΜ AZD for 24 h, respectively. Total RNA was isolated as described above. Three biological repeats were employed to transcriptome analysis. Sequencing libraries were generated by NEB Next Ultra™ Directional RNA Library Prep Kit for Illumina (New England Biolabs) following the manufacturer’s recommendations. An Illumina Hiseq 4000 platform was used to sequence these libraries, and 100 bp paired-end reads were generated. TopHat2 was used to map these reads to the SPSS 20.0 software  was used to perform all statistical analysis of data. The data represent the mean ± SD of n = 3 independent experiments. ANOVA analysis and a Tukey-Kramer multiple comparison test was used for data analysis. P < 0.05 was considered as statistically significant.How to cite this article: Song, Y. et al. The crosstalk between Target of Rapamycin (TOR) and Jasmonic Acid (JA) signaling existing in Arabidopsis and cotton. Sci. Rep.7, 45830; doi: 10.1038/srep45830 (2017).Publisher's note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.


The aim of this study was to examine the relationship between insulin-like growth factor-1 (IGF-1) and serum uric acid (UA) in Chinese children and adolescents with idiopathic short stature (ISS). A cross-sectional study of 91 Chinese children and adolescents with ISS was performed. Anthropometric measurements and biochemical parameters were tested. The standard deviation score of IGF-1 (IGF-1 SDS) was calculated.P = 0.004). In multivariate piecewise linear regression, the levels of IGF-1 SDS increased with the elevation of UA when UA was between 168 μmol/L and 301 μmol/L . The levels of IGF-1 SDS decreased with the elevation of UA when UA was either less than 168 μmol/L  or more than 301 μmol/L . A univariate analysis displayed a significant positive correlation between IGF-1 SDS and UA ( This study demonstrated a nonlinear relationship between IGF-1 and UA levels in Chinese children and adolescents with ISS. This finding suggests that either high or low levels of UA may have an adverse effect on IGF-1, whereas appropriate UA levels have a beneficial effect. The growth hormone-insulin-like growth factor-1 (GH-IGF-1) axis is central to the regulation of growth and development. Low levels of GH or IGF-1 can result in short stature in childhood. Usually, children with idiopathic short stature (ISS) show normal IGF-1 levels, but in some other cases IGF-1 levels can be lower  . Kim et Related studies have shown that the generation of IGF-1 was influenced by diet and nutritional status , 5. MoreUric acid (UA) was the final product of purine metabolism of nucleic acids. The majority of serum UA was freely filtered through the kidney glomeruli. According to preliminary estimates, approximately 90% of the filtered UA was reabsorbed by renal tubule , which iUntil now, the association between IGF-1 levels and UA concentrations in children with ISS has not been studied. The aim of this study was to examine the relationship between IGF-1 and UA in Chinese children and adolescents with ISS.We retrospectively reviewed the medical records of short-statured children and adolescents from the Department of Endocrinology, Affiliated Hospital of Jining Medical University, between January 2015 and December 2016. Ninety-one children and adolescents with ISS  aged 10.1 ± 3.8 years were enrolled in this study. The subjects were selected based on the following inclusion criteria: short stature, a condition wherein the height of an individual was more than 2 SD scores (SDSs) below the corresponding mean height for a given age, sex, and population group; normal weight and height at birth; peak GH levels > 10 ng/ml in at least two GH stimulation tests. The exclusion criteria included the presence of chronic organic diseases, GH deficiency, chromosomal abnormalities, skeletal dysplasia, genetic metabolic diseases, thyroid dysfunction, and abnormal liver function, as well as the use of medication that interferes with GH secretion or function .The study was approved by the Human Ethics Committee of the Affiliated Hospital of Jining Medical University . All of the families of the patients were informed of the aims of the study, and informed consent was written and obtained from all patients of the subjects.Heights were measured by a specially designated individual using the same measuring instrument  in the morning with an allowable error range of 0.1 cm. Height SDS was calculated based on normal range of Chinese children . BMI wasμg/L. Serum IGF-1 and IGFBP-3 levels were measured by the chemiluminescence immunometric method  with intra- and interassay CVs for IGF-1 of 3.0% and 6.2%, respectively, and intra- and interassay CVs for IGFBP-3 of 4.4% and 6.6%, respectively. Measures of liver function , AST, and gamma-glutamyl transferase (GGT)), kidney function , and UA), lipid profiles , high-density lipoprotein cholesterol (HDL-C), LDL-C, and triglycerides (TG)), and fasting plasma glucose (FPG) were tested by a biochemical autoanalyzer . Measures of thyroid function, including Free T3 (FT3), Free T4 (FT4), thyroid-stimulating hormone (TSH), gonadotropin, cortisol rhythm, and adreno–cortico–tropic hormone (ACTH), were tested by a luminescence immunoassay system . The standard deviation score of IGF-1 (IGF-1 SDS) was calculated according to a previous study [To assess GH secretion, L-dopa  and insulin  were administered orally or subcutaneously after overnight fasting. Blood samples were collected 0, 30, 60, 90, and 120 min later to obtain the serum GH concentration for each time point. GH was measured using a chemiluminescence method  with an analytical sensitivity of 0.010 us study .https://www.r-project.org) and EmpowerStats . Normally distributed variables are expressed as the mean ± standard deviation (SD), abnormally distributed variables are shown as the median (quartile), and non-parametric tests were used. Qualitative data are expressed as the numbers and percentage . First, we describe the demographic characteristics and biochemical values of the subjects  below 0.05 were considered statistically significant.All of the statistical analyses were performed with R statistical software , and UA (P = 0.004), whereas a significant negative correlation was found between the IGF-1 SDS and LDL-C (P = 0.015) and AST (P = 0.001).Univariate analysis was performed to analyze the relationship between IGF-1 SDS and each variable. As shown in μmol/L and 301 μmol/L. Specifically, the IGF-1 SDS decreased with the elevation of UA when UA was less than 168 μmol/L  or more than 301 μmol/L , whereas the IGF-1 SDS increased with the elevation of UA when UA was between 168 μmol/L and 301 μmol/L .Covariate screening was used to screen for possible confounders. The screening criteria included effect factors producing a >10% change when introducing covariates in the basic model or eliminating covariates in the regression model. The results revealed that body weight, pubertal stage, Cr, LDL-C, and TG met the filter criteria. In addition, smooth curve fitting was performed after adjustment for all variables, and the resultant curve exhibited a three-stage change and two breakpoints . Namely,μmol/L and 301 μmol/L. In contrast, there was an inverse relationship between IGF-1 SDS and UA when the concentration of UA was either less than 168 μmol/L or more than 301 μmol/L.In this cross-sectional study, we demonstrated a nonlinear association between IGF-1 SDS and serum UA in Chinese children and adolescents with ISS. Namely, there was a positive relationship between IGF-1 SDS and UA when the concentration of UA was between 168 μmol/L. However, there has been no research on the relationship between IGF-1 and the low levels of UA until now. Given the small sample size of this group, it is necessary to carry out related studies to explore the correlation between IGF-1 and low levels of UA and its potential mechanism. In addition, although the difference was not statistically significant, our research showed an inverse trend between IGF-1 SDS and UA when the concentration of UA was greater than 301 μmol/L. This relationship was consistent with findings by Sesti et al. [μmol/L could induce the downregulation of GH-stimulated IGF-1 expression [UA is the end product of purine metabolism in humans. It is known that high levels of UA are associated with many metabolic diseases –16, wheri et al. . Furtherpression . The potpression . Additiopression .μmol/L and 301 μmol/L, which was inconsistent with previous findings [Notably, this study also described a positive relationship between IGF-1 SDS and UA when the concentrations of UA were between 168 findings . To the findings . This imfindings . As far findings , 26, andfindings . UA may findings . Therefoμmol/L (n = 5). Therefore, we should further verify our results by expanding the sample size in any future studies. Finally, because of the study's cross-sectional nature, the present findings only exhibited a nonlinear relationship between IGF-1 levels and the serum UA concentration. Prospective basic and clinical studies are thus required to further confirm causality.Several potential limitations of this study should be considered. First, we failed to collect information about the diet of the subjects and did not perform a diet analysis of our study group. Therefore, we intend to design the dietary questionnaire to further analyze the effects of diet on IGF-1. Second, the levels of serum UA were influenced by diet, metabolism, and other factors. Therefore, it is necessary to measure enzymes related to UA synthesis or decomposition, such as xanthine oxidase or adenosine deaminase. In addition, the present study did not measure urinary UA levels for 24 hours, which was a stable indicator to assess the excretion of UA. Third, the number of samples in this study was relatively small, especially in the group of UA < 168 In conclusion, in this present study, we described a nonlinear relationship between IGF-1 and UA levels in Chinese children and adolescents with ISS after adjusting for several confounders. This finding suggests that either high or low levels of UA may have an adverse effect on IGF-1, whereas appropriate UA levels have a beneficial effect on IGF-1.


RNA (miR)‐140‐5p is known to be reduced in both pulmonary arterial hypertension (PAH) patients and monocrotaline‐induced PAH models in rat. Identification of target genes for miR‐140‐5p with bioinformatics analysis may reveal new pathways and connections in PAH. This study aimed to explore downstream target genes and relevant signaling pathways regulated by miR‐140‐5p to provide theoretical evidences for further researches on role of miR‐140‐5p in PAH. Multiple downstream target genes and upstream transcription factors (TFs) of miR‐140‐5p were predicted in the analysis. Gene ontology (GO) enrichment analysis indicated that downstream target genes of miR‐140‐5p were enriched in many biological processes, such as biological regulation, signal transduction, response to chemical stimulus, stem cell proliferation, cell surface receptor signaling pathways. Kyoto Encyclopedia of Genes and Genome (KEGG) pathway analysis found that downstream target genes were mainly located in Notch, TGF‐beta, PI3K/Akt, and Hippo signaling pathway. According to TF–miRNA–mRNA network, the important downstream target genes of miR‐140‐5p were PPI, TGF‐betaR1, smad4, JAG1, ADAM10, FGF9, PDGFRA, VEGFA, LAMC1, TLR4, and CREB. After thoroughly reviewing published literature, we found that 23 target genes and seven signaling pathways were truly inhibited by miR‐140‐5p in various tissues or cells; most of these verified targets were in accordance with our present prediction. Other predicted targets still need further verification in vivo and in vitro.The expression of micro GOgene ontologyKEGGkyoto encyclopedia of genes and genomePAHpulmonary arterial hypertensionPASMCpulmonary arterial smooth muscle cellTFtranscription factorin situ thrombosis Pulmonary arterial hypertension (PAH) is a chronic progressive disease of pulmonary vasculature characterized by sustained elevation of pulmonary vascular resistance and pulmonary arterial pressure, consequently leading to right heart failure and eventual death http://mirbase.org/index.shtml).Mature sequences of miR‐140‐5p in various species were obtained in the miRBase database (http://www.microrna.org/), TargetScan (http://www.targetscan.org/), RNAhybrid (https://bibiserv.cebitec.uni-bielefeld.de/rnahybrid/submission.html), and miRDB (http://www.mirdb.org/) databases were used to predict the target genes of miR‐140‐5p. To make our predicted target genes more convincible, only the target genes predicted by at least three databases were selected for further analyses.Identification of target genes is critical for characterizing the functions of miRNAs. In this study, miRanda (http://david.abcC.ncifcrf.gov), was used to perform GO analysis to identify biological processes, cellular components, and molecular functions of these target genes. At the same time, the probable signaling pathways in which these target genes were enriched were analyzed by KEGG database (http://www.genome.jp/kegg/). The P‐value <0.05 was considered significant.Target mRNA of miR‐140‐5p supported by at least three databases were used for GO analysis to predict gene functions. Integration Discovery (DAVID) software, version 6.7 (http://jaspar.binf.ku.dk/), which includes various vertebrate TFs. The P‐value <0.0001 was considered significant.Human miR‐140‐5p precursor was obtained in the miRBase database and its 5000 bp upstream was defined as the miR‐140‐5p promoter. The TFs of miR‐140‐5p were predicted using MOODS‐python software (version 1.9.3) in JASPAR database (http://gephi.github.io/).By merging the regulatory relationships between TFs and miR‐140‐5p, miR‐140‐5p and target genes, genes and genes , we constructed a comprehensive TF–miR‐140‐5p–mRNA regulatory network using Gephi software  and participated in the molecular functions of protein binding, activating transcription factor binding, ion binding, lipid binding, and so on (P < 0.05). In addition, the target genes of miR‐140‐5p were involved in various biological processes, including biological regulation, metabolic process, cell communication, signal transduction, response to chemical stimulus, stem cell proliferation, cell surface receptor signaling pathway (P < 0.05). Fig. GO enrichment analysis was conducted for the target genes of miR‐140‐5p predicted by at least three databases. As shown in Table P‐values. As shown in Table P < 0.05); among them, Notch, TGF‐beta, PI3K/Akt, and Hippo signaling pathways were well known to be associated with the pathogenesis of PAH. Fig. Enriched signaling pathways for the target genes of miR‐140‐5p identified by KEGG pathway analysis were ranked according to the P‐value <0.0001 was 393. To reduce false‐positive results, TFs with a quality score (Q‐score) less than 10 were filtered. As shown in Table The number of predicted TFs for miR‐140‐5p with A comprehensive electronic search of Web of Science and PubMed databases was performed until April 20, 2017, to obtain target genes and signaling pathways modulated by miR‐140‐5p in published studies. Finally, a total of 26 papers including 23 target genes and seven signaling pathways inhibited by miR‐140‐5p were obtained; most of them focus on the functions of miR‐140‐5p suppressing tumor growth, migration, and invasion in various tumor tissues and cells. Two recent studies have found that SMURF1 and Dumt1 are direct target genes of miR‐140‐5p in pulmonary arterial smooth muscle cells (PASMCs) and are involved in the pathogenesis of PAH. The details are shown in Table Pulmonary arterial hypertension is a chronic life‐threatening condition requiring long‐term management In this study, we found that the target genes of miR‐140‐5p were enriched in many biological processes, such as biological regulation, metabolic process, cell communication, signal transduction, response to chemical stimulus, stem cell proliferation, cell surface receptor signaling pathway. In KEGG pathway analysis, the target genes of miR‐140‐5p were mainly located in Notch, TGF‐beta, PI3K/Akt, and Hippo signaling pathways. According to the TF–miRNA–mRNA network, the important genes potentially regulated by miR‐140‐5p included PPI, TGF‐betaR1, smad4, JAG1, ADAM10, FGF9, PDGFRA, VEGFA, TLR4, LAMC1, CREB, and the upstream TFs, which might regulate miR‐140‐5p expression including TAX5, FOXI, IRF1, GATA6, RUNX2. After thoroughly reviewing published literature, we found that 23 target genes and seven signaling pathways were truly inhibited by miR‐140‐5p in various tissues or cells; most of these downstream targets were in accordance with our present prediction.Several studies have shown that activation of Notch3 pathway is involved in the pathogenesis of PAH in vivo and in vitro.Our predicted network provided potential target genes and relevant signaling pathways that might be modulated by miR‐140‐5p contribution to the development of PAH. Several targets and pathways predicted in our analysis, such as TGF‐betaR1, ADAM10, FGF9, PDGFRA, VEGFA and Notch, PI3K/Akt, TGF‐beta cascades, have been demonstrated to mediate the effects of miR‐140‐5p on antiproliferation and prodifferentiation in several cell types in published studies ML and FL designed the study; WS, YW, LC, and QW analyzed and interpreted the data; WF, XY, QZ, and JW organized the results; FL wrote the manuscript.


The Human T-Lymphotropic Virus type 1c subtype (HTLV-1c) is highly endemic to central Australia where the most frequent complication of HTLV-1 infection in Indigenous Australians is bronchiectasis. We carried out a prospective study to quantify the prognosis of HTLV-1c infection and chronic lung disease and the risk of death according to the HTLV-1c proviral load (pVL).840 Indigenous adults  were recruited to a hospital-based prospective cohort. Baseline HTLV-1c pVL were determined and the results of chest computed tomography and clinical details reviewed. The odds of an association between HTLV-1 infection and bronchiectasis or bronchitis/bronchiolitis were calculated, and the impact of HTLV-1c pVL on the risk of death was measured.5 peripheral blood leukocytes (log-rank χ2 (2df) = 6.63, p = 0.036) compared to those with lower HTLV-1c pVL or uninfected subjects. Excess mortality was largely due to bronchiectasis-related deaths .Radiologically defined bronchiectasis and bronchitis/bronchiolitis were significantly more common among HTLV-1-infected subjects . Median HTLV-1c pVL for subjects with airways inflammation was 16-fold higher than that of asymptomatic subjects. There were 151 deaths during 2,140 person-years of follow-up (maximum follow-up 8.13 years). Mortality rates were higher among subjects with HTLV-1c pVL ≥1000 copies per 10Higher HTLV-1c pVL was strongly associated with radiologically defined airways inflammation and with death due to complications of bronchiectasis. An increased risk of death due to an HTLV-1 associated inflammatory disease has not been demonstrated previously. Our findings indicate that mortality associated with HTLV-1c infection may be higher than has been previously appreciated. Further prospective studies are needed to determine whether these results can be generalized to other HTLV-1 endemic areas. The Human T-Lymphotropic Virus type 1 (HTLV-1) infects up to 20 million people worldwide who predominantly reside in resource-limited areas. The virus is associated with a haematological malignancy , and inflammatory diseases involving organ systems including the spinal cord, eyes and lungs. Determining the outcomes of infection in most HTLV-1 endemic areas is extremely difficult; however, the virus is highly endemic to central Australia where the Indigenous population has access to sophisticated medical facilities. We prospectively followed a large hospital-based cohort of Indigenous Australian adults that was well characterized with regard to base-line comorbid conditions, HTLV-1 serostatus and HTLV-1 proviral load (pVL). A higher baseline HTLV-1 pVL was strongly associated with an increased risk of airway inflammation (bronchitis/bronchiolitis and bronchiectasis) and death, which most often resulted from complications of bronchiectasis. Increased mortality due to an HTLV-1-associated inflammatory condition has not been demonstrated previously. The morbidity and mortality associated with HTLV-1 infection may therefore be substantially higher than has been assumed from an analysis of cohorts of subjects with adult T-cell leukaemia or HTLV-1-associated myelopathy. These findings have important implications for epidemiological research and for determining health care priorities in resource-limited settings. The Human T-Lymphotropic Virus type 1 (HTLV-1) is an oncogenic retrovirus that preferentially infects CD4+ T cells. An endeClinically significant sequelae of HTLV-1 infection include a haematological malignancy, Adult T cell Leukemia/Lymphoma (ATL), and inflammatory diseases, such as HTLV-1 associated myelopathy/tropical spastic paraparesis (HAM/TSP),6. ChestIn endemic areas in Japan and Africa, HTLV-1 seropositivity is associated with increased mortality–12, whic2. Critically ill patients are transferred by air to ASH, which has sophisticated diagnostic capabilities.Alice Springs Hospital (ASH) is the only medical facility serving central Australia, an area of >1,000,000 kmst June 2008 to 31st December 2013, were identified from the ASH patient management database, which coordinates all in-patient and out-patient hospital activities. Indigenous status was determined from self-reported data obtained at admission, as recorded in the patient information database. Potential subjects were offered enrolment when next admitted for >48 hours. Among 165 eligible cases, 154 were recruited . Written reports for chest high-resolution computed tomography (cHRCT) were reviewed for all subjects, confirming bronchiectasis in 104 cases and bronchitis with or without bronchiolitis in 33 cases . Patientalone, 1).Demographic and clinical details were extracted from medical records at the time of recruitment using a standardized data-collection form. HTLV-1 associated conditions were identified from medical records at baseline and study end. No control patient developed chronic pulmonary disease during the study period. Mortality data was obtained at study end from the ASH patient management database, and the cause of death was determined from death certificates held in Registries in the Northern Territory of Australia and South Australia. Death certificates were not available for four subjects who died in remote communities in Western Australia, for whom a cause of death was sought from the responsible remote clinic.Bronchitis was diagnosed where cHRCT revealed bronchial wall thickening or dilatation not fulfilling criteria for bronchiectasis, and bronchiolitis where cHRCT revealed multiple centrilobular nodules or a ‘tree-in-bud’ pattern. ChronicThe study was approved by the Central Australian Human Research Ethics Committee. All patients, and their parents/guardians if aged <18 years, gave written informed consent in primary languages.gag gene in the p19 coding region of the Australo-Melanesian HTLV-1 subtype C[5 peripheral blood leukocytes; PBL) was calculated. Samples and standards were extracted using the Qiagen QIA blood Mini Extraction kit and the extracts amplified on a Stratagene Mx3000p Real Time PCR Instrument (Integrated Sciences). The extract (5 μL) was added to 20 μL of Master mix containing 2 x Brilliant Multiplex QPCR Master Mix (Agilent Technologies) 0.3 μM of each primer (Gene works) and 0.16 μM of each probe (Sigma-Aldrich) and amplified at 95°C for 10 minutes, 45 cycles at 95°C for 30 seconds, 65°C for 60 seconds and 72°C for 60 seconds.Whole blood samples were collected from each participant at the time of recruitment. Peripheral blood buffy coats (PBBC) were prepared, and plasma and PBBCs were stored at ASH at -80° C until transfer to the National Serology Reference Laboratory, Melbourne. Samples were screened for antibodies to HTLV-1 using both an enzyme immunoassay  and a particle agglutination assay . Any sample reactive on either screening assay was tested by Western blot  and HTLV-1c PCR. Primers and fluorescently labelled hydrolysis probes were designed to target a highly conserved 88 bp fragment of the subtype C and mult5 PBL, were HBsAg negative and strongyloides seronegative. Although 53 subjects met these criteria, technical difficulties prevented analysis for nine subjects . The OCI was therefore compared between 29 asymptomatic and 15 symptomatic subjects .The clonality of HTLV-1-infected PBLs was determined by high-throughput sequencing of PBBC cell genomic DNA. The oligoclonality index (OCI) was calculated as previously described, and the5 PBL, a cut-off that has been associated with an increased risk of HAM/TSP[th March 2015. The association between overall mortality and HTLV-1 pVL was assessed using log-rank tests and Kaplan-Meier curves in univariate analysis and using Cox regression for multivariate analysis. Associations with cause-specific mortality were assessed using competing risks analysis with all causes except the specific cause of interest treated as a competing risk. Where HTLV-1 pVL was treated as a categorical variable we also tested for a trend by creating a continuous variable with value zero for those uninfected, and with the median value of HTLV-1 pVL for those in the low and high pVL categories. Predictors of chronic airways inflammation were assessed using multivariate binary logistic regression. A 2-sided Type 1 error rate of p<0.05 was regarded as indicating statistical significance in each analysis.All analysis was performed using Stata version 14.2 . HTLV-1 pVL was log-transformed and also categorized as low if <1000 and high if ≥1000 per 10f HAM/TSP. DiffereDemographic and clinical characteristics are presented in Radiologically defined airways inflammation was more common among HTLV-1c-infected subjects . Bronchi5 PBL , those for subjects with bronchiectasis  (p = 0.001) and bronchitis/bronchiolitis (p = 0.042) were 16-fold higher, was 16-fold higher than that of asymptomatic subjects (p<0.001). Few other recognised causes of bronchiectasis were found   , while H= 0.006) .5 PBL for subjects who were strongyloides seronegative , seropositive  and those with equivocal strongyloides serological results .Other HTLV-1 associated conditions included infective dermatitis (4), crusted scabies (4), probable HAM/TSP (2) and uveitis (2). Four subjects with HTLV-1-associated bronchiectasis had other sequelae of HTLV-1 infection . There was no difference in strongyloides seropositivity between groups , nor was5 PBL) developed ATL during follow-up, one developed penile cancer  and another metastatic anal cancer .Although the risk of malignancy was not increased among HTLV-1c infected subjects, one subject  and those with airways inflammation (p = 0.20).The median OCI did not differ between asymptomatic  and symptomatic groups  (p = 0.51) or when symptomatic subjects with uveitis were excluded from the analysis  (p = 0.341). Median logDuring 2140 person-years of follow-up, 155 deaths were recorded . Non-bronchiectasis causes of death were infections (37), cardiovascular disease (37), malignancy (15), end-stage kidney disease (ESKD) (15), chronic liver disease (10), intracerebral haemorrhage (4), primary pulmonary hypertension (2) and amyloidosis (1).Subjects with high HTLV-1c pVL were more likely to die during the study period . MortaliRelative to HTLV-1c uninfected subjects, unadjusted hazard ratios for a bronchiectasis-related death were 4.73  and 1.44  for subjects with high and low HTLV-1c pVL, respectively .5 PBL when median values for proviral load were treated as a continuous variable in an adjusted model (p = 0.009 for trend). This effect was not apparent among the 736 subjects without bronchiectasis  when other causes of death were treated as competing risks and after adjusting for alcohol, place of residence, age at test, and gender  Fig 4)..Fig 4). iectasis . Causes Mean (±SD) age at death for bronchiectasis-related deaths (49.5±15.3 years) was lower than that of subjects who died due to cardiovascular disease (55.0±11.5 years)(p = 0.073), sepsis (56.7±16.7)(p = 0.044) or malignancy (60.1±9.3) (p = 0.007).In a large hospital-based cohort of Indigenous Australian adults, a higher baseline HTLV-1c pVL prospectively predicted a bronchiectasis-related death, which occurred at a mean age of only 49.5 years. In addition to confirming a previously reported association between HTLV-1c infection and bronchiectasis,15. The tax/rex mRNA[HTLV-1 associated inflammatory diseases are thought to result from a genetically determined, inefficient cytotoxic T lymphocyte response, permitting widespread dissemination of the virus in a large number of HTLV-1-infected T-cell clones, which is reflected in a high HTLV-1 pVL. Organ i/rex mRNA expressi/rex mRNA. In larg/rex mRNA,8. The p/rex mRNA. Persist/rex mRNA,8, and a/rex mRNA and Braz/rex mRNA suggest /rex mRNA, publishConsistent with other HTLV-1 associated inflammatory diseases, the medIncreased mortality due to a specific HTLV-1-associated inflammatory disease has not been prospectively demonstrated previously. However, an excess mortality that is not attributable to ATL or currently recognized HTLV-1-associated inflammatory diseases has been reported in other endemic areas. Adjusted hazard ratios of death are 1.3 to 1.77–The strengths of this prospective cohort study include the recruitment of nearly all eligible subjects with a discharge diagnosis of bronchiectasis, the use of cHRCT for diagnosis and the blinding of ASH researchers to the HTLV-1 serostatus of subjects and of those who performed HTLV-1 studies to their clinical state. Nevertheless, some design limitations must be recognized. First, investigations to exclude other causes of airways disease could not be performed in all cases. However, consistent with previous studies,6, a speIn summary, HTLV-1c infection and higher HTLV-1c pVL were strongly linked to airways inflammation in a hospital-based cohort of Indigenous Australian adults. Furthermore, higher baseline HTLV-1c pVL prospectively predicted death due to bronchiectasis, which may result from more extensive disease,6, prediS1 Checklist(DOC)Click here for additional data file.


Increased HTLV-1 Tax expression induces the expression of various cellular genes such as IL-2 and IL-15, which directly contributes to lymphocyte activation and immunopathogenesis in HAM/TSP patients. However, little is known about the molecular and cellular mechanism of HBZ in development of HAM/TSP. It has been reported that HBZ mRNA expression was detected in HAM/TSP patients higher than in asymptomatic carriers and correlated with proviral load and disease severity. Unlike HTLV-1 tax, HBZ escapes efficient anti-viral immune responses and therefore these reactivities are difficult to detect. Thus, it is important to focus on understanding the function and the role of HTLV-1 tax and HBZ in disease development of HAM/TSP and discuss the potential use of these HTLV-1 viral gene products as biomarkers and therapeutic targets for HAM/TSP.Human T cell lymphotropic virus type 1 (HTLV-1) infection can lead to development of adult T cell leukemia/lymphoma (ATL) or HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP) in a subset of infected subjects. Understanding the interaction between host and HTLV-1 and the molecular mechanisms associated with disease pathogenesis is critical for development efficient therapies. Two HTLV-1 genes,  Several disease . The spi disease . As the  disease . Recent  disease . In HAM/ disease . A highe disease . High le disease . Intrathgag, pol, and env flanked by long terminal repeat at both ends. HTLV-1 genome also contains a pX region between env and 3′ LTR encoded several accessory genes including tax, rex, p12, p21, p30, p13 and HTLV-1 basic leucine zipper factor (HBZ)  . The virdisorder . There i+ T cells in vivo  and are engaged in the maintenance of immunologic self-tolerance by suppressing the activation and expansion of self-reactive lymphocytes that may cause autoimmune diseases , play critical roles in suppressing the immune response, such as inhibition of inflammatory responses and promotion of Treg generation and function . Dysregu culture . Higher  culture . Thus, d+ T cells and also HTLV-1 Tax-specific cytotoxic CD8+ T cells , which consists primarily of a continuous endothelium with tight junctions. HAM/TSP develops upon infiltration of HTLV-1-infected lymphocytes into the CNS, mostly within the thoracic spinal cord. The tight junctions of the BBB endothelium in HAM/TSP patients are locally disorganized , 2008. M+CD25+ T cells from HAM/TSP patients , vascular cell adhesion molecule 1 (VCAM-1) and cell adhesion molecule 1 (CADM1/TSLC1) . Activatpatients . ALCAM epatients . These rtax mRNA and Tax protein are rarely or undetectable directly in fresh uncultured PBMCs of HAM/TSP patients, it has been reported that HTLV-1 tax mRNA was detected in cells of spinal cord and cerebellar sections, and HTLV-1 Tax protein could be also detected in CSF cells of HAM/TSP patients   . CD8+ T subjects . HoweverLV-1 PVL . A recenpatients . Moreove the CNS .While virus-specific antibodies play an important role in the control of viral infections in the CNS, intrathecal antibody synthesis appear to be associated with both protective and pathogenic function in chronic infection and immune-mediated diseases of the CNS. Intrathecal antibody synthesis against HTLV-1 has been also confirmed by the presence of HTLV-1-specific antibodies and oligoclonal IgG band in CSF of HAM/TSP patients . Intrath+ T cells are detected in AC and HAM/TSP patients, and HBZ-specific CTL clones were able to lyse naturally infected cells isolated from AC and HAM/TSP patients, but not ATL patients (+ T cells was very low in peripheral blood (HBZ is also an immunogenic protein recognized by HBZ-specific CTL clones . HBZ-spepatients . Howeveral blood .Antibody response against HBZ was detected in serum/plasma of HTLV-1-infected subjects, but the frequency of the subjects with anti-HBZ antibody was about 10–16% in ACs, ATL, and HAM/TSP patients and did not discriminate between clinical status . In addiHBZ gene is constantly expressed while the tax gene is sporadically transcribed. Thus, HTLV-1-infected cells might be able to escape from host immune responses for long periods and result in persistence of HTLV-1 in infected individuals.It has been suggested that the low frequency and affinity of HBZ-specific immune responses may be consequence of the low expression and antigenicity of HBZ in infected cells. Unlike Tax-specific immune responses, it might be difficult to induce effective HBZ-specific immune responses in HAM/TSP patients although the Both Tax and HBZ play critical roles in immune dysregulation in HAM/TSP. However, it is still unknown how expression of Tax and HBZ are regulated and how Tax and HBZ cooperate in naturally infected cells. Therapies that control the expression of HTLV-1 gene products might be effective in preventing the development of HAM/TSP. In addition, effective regulation or induction of HTLV-1 specific immune responses might improve the prognosis of patients with this disorder. Further studies will be necessary to identify the mechanism by which HTLV-1 gene products or other factors including HTLV-1-specific immune responses contribute to the pathogenesis of HAM/TSP.YE-A contributed to paper writing. AV contributed to discussion. SJ supervised and contributed to discussion and writing.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.