Material properties are sensitive to atomistic structure defects such as vacancies or impurities, and it is therefore important to determine not only the local atomic configuration but also their chemical bonding state. Annular dark-field scanning transmission electron microscopy (STEM) combined with electron energy-loss spectroscopy has been utilized to investigate the local electronic structures of such defects down to the level of single atoms. However, it is still challenging to two-dimensionally map the local bonding states, because the electronic fine-structure signal from a single atom is extremely weak. Here, we show that atomic-resolution differential phase-contrast STEM imaging can directly visualize the anisotropy of single Si atomic electric fields in monolayer graphene. We also visualize the atomic electric fields of Stone–Wales defects and nanopores in graphene. Our results open the way to directly examine the local chemistry of the defective structures in materials at atomistic dimensions. Imaging chemical bonding states in defective graphene is important to determine its functional properties. Here, the authors report triangular and rectangular atomic electric fields in monolayer graphene induced by silicon as imaged by differential phase contrast STEM. Pure graphene consists of sp2 hybridized carbon atoms arranged in a honeycomb lattice, but synthesis inevitably introduces a wide variety of atomistic structure defects including impurity dopants, vacancies, non-hexagonal polygons, edges, and nanopores5. Although these structural defects often adversely affect the mechanical or electronic transport properties, graphene edges and nanopores have potential for applications in chemical separation, water-splitting, DNA sequencing, and catalysis3.Graphene is an atomically thin two-dimensional (2D) material, exhibiting a vast range of prominent physical and chemical properties such as mechanical, electric, magnetic, and catalytic properties6. Moreover, the electronic structure can be probed at the atomic level by incorporating electron energy-loss spectroscopy (EELS) in STEM9. The energy-loss near-edge structure (ELNES) is correlated to the local chemical bonding states. However, ELNES signals are extremely weak and it is still challenging to obtain atomic-resolution 2D ELNES maps, especially at defective regions, because the specimen is unstable under the beam irradiation and mechanical instability of the microscope hinders using very long acquisition times. Apart from spectroscopic analysis, differential phase-contrast (DPC) STEM imaging has recently been shown to be capable of visualizing the electric field in atoms12. According to the literature by Linus Pauling13, a chemical bond can be interpreted as the forces acting between atoms, and therefore the direct imaging of electric fields (force fields for electric charges) with DPC–STEM may experimentally probe the local chemical bonding states in two dimensions.Low voltage (< 80 kV) annular dark-field scanning transmission electron microscopy (ADF-STEM) imaging has greatly contributed to the direct identification of individual elements and their locations inside defects in 2D materials14, operating the microscope at 80 kV 15. We note that the present electric field imaging requires a relatively higher spatial resolution than the ADF-STEM image, because the chemical bonding information is present in the signal shape in the space between the atom positions. Therefore, we operated the microscope at 80 kV, which is smaller than the knock-on threshold of 86 kV for carbon atoms. By properly choosing the electron microscope operating condition and the detector geometry, we elucidate the anisotropy of single Si atomic electric fields located in the coordination of threefold and fourfold symmetry in graphene. Furthermore, we also identify the local enhancement or suppression of electric fields of topological defects such as Stone–Wales defects and nanopores in graphene. This result opens a new capability to investigate the local chemistry of atomistic defects in materials and might shed light on our understanding of material properties at the level of single atoms.Here, we show atomic-resolution electric field imaging at dopants and topological defects in graphene by using DPC–STEM recorded with a second-generation segmented annular all-field detector 16. However, it is the coherently scattered electrons in the bright-field region that are expected to contain rich information on the local chemical bonding states. In DPC–STEM, the segmented detectors are set in the bright-field region12 and thus permit direct visualization of the local electric fields at atomic dimensions, where the electric field is detected as the variation of the center of mass (CoM) in the diffraction pattern for each probe position17. A major issue for reliable atomic electric field imaging in DPC–STEM is dynamical diffraction: when a specimen has heavy elements or several-nanometer thickness18, the electric field estimated from the measured CoM in the diffraction plane may deviate significantly from the actual electric field. As graphene is composed of light carbon atoms with single-atom thickness, dynamic diffraction is negligible, and so graphene is an ideal specimen for quantitative electric field imaging in DPC–STEM. However, this requires a trade-off between the experimental signal and specimen damage: a higher electron dose is required to reliably detect the very tiny CoM signal from the weakly scattering carbon atoms in graphene. Even at accelerating voltages lower than the knock-on threshold (< 86 kV), dopants or topological defects in graphene seldom survive high electron dose or prolonged beam irradiation19. To overcome the specimen damage while maintaining a good signal-to-noise ratio (SNR), we performed sequential fast-scanning DPC–STEM imaging with the scintillator-type segmented detector, which suppresses both the electron dose rate limited damage and severe specimen drift20.When an atomically focused electron probe is traveling through the atoms in materials, a fraction of the incident electrons is incoherently scattered into high angles. This imaging mode is known as ADF-STEM, which can be used to directly determine the location of the atoms and is also sensitive to the chemical type −1 by dividing the specimen thickness). Figure 12. For an ionic crystal, the electric field can be approximated to be the superposition of the atomic electric fields of the isolated atoms, and the electric-field sharp minima appear only at the nuclear positions. However, the observed electric field strength map of graphene has a rather complicated intensity distribution: in addition to the nuclear positions (T-site: top site), several electric field minima appear in-between atoms as dark dot contrast. The minimum electric fields appear at the high symmetry points of the carbon hexagon: the hexagon center (H6-site: hollow site) and the midpoints of the hexagon ridges (B-site: bridge site). Owing to the strong covalent bonding between carbon atoms, the interatomic distance is as small as 142 pm, with the consequence that the atomic electric fields of adjacent carbon atoms partially overlap, leading to the emergence of the electric field minima at the high symmetry points. For example, at the B-site, the electric fields of the two neighboring carbon atoms have exactly the same strength but in the opposite direction, and consequently the net projected electric field cancels out . Similarly, the net projected electric field at the H6-site is also a minimum as a result of the superposition of the electric fields of the surrounding six carbon atoms. Figure Figure 13, acting as cations and anions, respectively. We then approximate these single anions or cations to be infinitesimal positive or negative charges, and therefore the observed graphene electric field can be considered as a force field for the single adatom at each position (Hellman–Feynman theorem21). On the basis of the observed graphene electric field image, the stable adsorption sites for the adatom are expected to be the positions of the minimum electric field or dark contrast in Fig. 22, which strongly supports our prediction. Graphene is a relatively simple system and DFT calculation can precisely predict the stable adatom site. However, DFT calculations may be difficult to perform in a large, defective, or complex materials system, and therefore the present view of experimental force field imaging could be useful for the further prediction of a local chemical reaction . From these considerations, DPC–STEM images can be considered as a force field for electric charges and hence this imaging mode may be said to enable visualization of the anisotropic force field between atoms, i.e., chemical bonding.To demonstrate the advantages of atomic electric field imaging, we consider the problem of finding the stable adsorption sites for adatoms on graphene. Since the forces of an adatom on graphene are minimized at a stable absorption site, a 2D force map may be used to find minimum force positions for the adatom. Usually, the single atoms of metallic elements lose their outer electrons easily, while the atoms of nonmetallic elements obtain additional electrons3, substituting for a single carbon atom) and fourfold-  coordinated Si dopants. As per previous EELS experiments9, the single Si dopant in threefold or fourfold coordination forms a different chemical bonding state of sp3 or sp2d, respectively, suggesting that the Si atomic electric field also depends on these defect configurations. Figure 24, and it is therefore necessary to quickly acquire images before defect-diffusion or defect-assisted graphene etching occurs. A series of fast-scanning DPC–STEM images (20 µs/pixel) were recorded  and then averaged after image alignment via cross-correlation20. Since Si is much heavier than C, the strong electric fields of single Si atoms can be recognized as a donut-like ring of bright contrast in Fig. 3 and Si–C4 configurations, respectively. The atomic electric fields of Si and C have an opposite sign along the covalent bonding direction and therefore the Si atomic electric field is suppressed along the C bonding directions , in agreement with our description. The bond length between C atoms in a five-membered ring is slightly shorter than that of a seven-membered ring and we may need to consider the effect of contrast transfer function (CTF) of DPC–STEM imaging for the contrast change26. However, the estimated contrast reduction in a five-membered ring is only a few percent and therefore the effect of CTF is negligibly small. For further confirmation of the validity of our interpretation, we performed quantitative intensity analysis and image simulation with the DFT-derived structure model of Stone–Wales defect . These results well reproduced the experimental electric field strength. The larger electric field within the seven-membered ring could easily collect cations at the hollow site, and therefore the Stone–Wales defect can be considered a chemically active nano-field, especially for a catalytic reaction with metal ions. Here, we note that some atomic reconfigurations were observed at the region indicated by a white circle in Fig. To demonstrate the validity of our description for electric field images, we observed various topological defects in graphene. Figure 27. Figure 28. The strong electric field was observed at the facet graphene edge in Fig. 29. If we can completely decorate the graphene edge by cations30, the local electric field should be much enhanced, supplying an intriguing chemically active nano-field. Thus, direct electric field imaging of graphene defects should be a very powerful method to experimentally understand their chemical stabilities and activities from the atomic scale.During the observations, the high chemical reactivity of nanopores with cations was readily realized by the fast growth of nanopores. As we have noted, Stone–Wales defects or larger membered rings have a strong capability to capture single cations which then act as a catalyst to the etching and growth of nanoporesWe have demonstrated the electric field mapping of atomistic defects in graphene, including single Si dopants in threefold and fourfold configurations, Stone–Wales defects, and nanopores by atomic-resolution DPC–STEM imaging. The electric field images can be considered as showing a force field formed by constituent atoms, which makes it possible to understand the local chemistry such as bonding orientation and the local chemical activity. Our findings show how electric field imaging can be used to investigate stable adatom sites and the growth of nanopores. Since the resultant signals in DPC–STEM imaging are mostly very small, we must carefully select the optimal experimental conditions such as accelerating voltage, beam current, and dwell time. The present method of multiple-frame averaging with a relatively low-dose condition could be helpful to suppress the electron beam damage. By properly choosing the experimental condition, the present electric field imaging is applicable to many other systems, and may also be helpful to identify the ion diffusion pathways in ionic conductive materials (the minimum electric field path) by experiments.12. For the construction of the atomic electric field, we approximate the center of mass based on the signals in the 16 detector segments17.We used a commercially available monolayer graphene , grown by chemical vapor deposition and then transferred onto a TEM grid. In this growth process, the sample was exposed in some Si sources, which provides Si dopants in graphene. To remove hydrocarbon contamination, the TEM grid was annealed at 400 °C for 3 h in a high vacuum TEM via in situ TEM heating holder , and then allowed to cool down to room temperature. Atomic-resolution ADF and DPC STEM images were acquired with a JEM 300CF installed in the University of Tokyo, equipped with a JEOL ETA corrector, cold field emission gun, and a second generation of SAAF detector (DPC STEM) operated at 80 kV. The used probe current was ~23 pA, as measured by a Faraday cup. The probe-forming aperture was 27 mrad in semi-angle , the ADF detector spanned 42–170 mrad, and the bright-field disk edge was set in the middle of the third ring of the SAAF detector3 and Si–C4 configurations, we performed DFT calculations using the VASP code31. The image simulations were performed using an absorptive model for thermal scattering, and imaging parameters consistent with the instrument configuration used: a 80-kV probe, a 27-mrad probe forming an aperture semi-angle, a chromatic aberration coefficient of 0.89 mm, and a Gaussian energy spread with a full-width-half-maximum of 0.45 eV. Effective source size was incorporated via convolution with a 2D pseudo-Voigt-like function, comprising a 20% weighting of a Gaussian with half-width-half-maximum of 0.39 Å and an 80% weighting of a Lorentzian with half-width-half-maximum of 0.47 Å as determined by fitting to the experimental ADF-STEM images.To obtain the stable defect structure models of Si–CSupplementary Information


Psychiatric diagnostic manuals recognise the importance of local expressions of distress in culturally diverse settings [i.e. idioms/cultural concepts of distress (CCDs)], yet there is a lack of consensus on how these should be incorporated into mental health related research.To perform a narrative synthesis and critical review of research exploring how idioms/CCDs have been integrated into assessment measures and interventions.A systematic review was conducted in accordance with PRISMA guidelines. An adapted version of the COSMIN checklist was used to assess the quality of the linguistic translation of the idioms/CCDs.Twenty-nine papers were included in the final review. Primary qualitative research was the most common method of gathering information about idioms/CCDs. The majority of studies described integrating idioms/CCDs into assessment measures as opposed to interventions. Some studies used information relating to idioms/CCDs to develop novel assessment measures, while others adapted pre-existing assessment measures. The measures generated moderate to high levels of validity. Information relating to the linguistic translation conducted in the completion of the studies tended to be inadequately reported.Integrating information about idioms/CCDs into assessment measures can enhance the validity of these assessments. Allocating greater research attention to idioms/CCDs can also promote more equitable exchanges of knowledge about mental health and wellbeing between the Global North and the Global South. Many of these documented terms can serve as some or all of these three subgroups of ‘CCDs’. For example ‘khyal cap’ is both an explanatory model – in that it may be perceived as the cause of a set of symptoms – and an idiom of distress – as it is a linguistic term used among particular groups to talk about suffering or distress. A review of epidemiological studies of CCDs by Kohrt et al.  movement has been engaged in concerted efforts to promote equitable access to mental health services for people worldwide. Commentators, such as Chowdhary et al. , have re et al., . Nichter et al., , 2010 in et al.,  uses thet et al.  found thet al. ?What were the psychometric properties of the assessment measures that were developed?As a multitude of terms are used to denote idioms/CCDs, for this review they were defined as any distress concept  described in the terms used by the DSM  or a distress concept that is described as being culturally influenced.The bulk of GMH-related research continues to be published in English-language journals. As such, research into idioms/CCDs frequently requires processes of translation and/or interpretation but also the negotiation of local and global epistemologies. To date, there has not been a systematic effort to identify and review the existing body of research that has sought to integrate information about idioms/CCDs into mental health assessment measures and/or interventions. The current systematic review sought to remedy this by conducting a narrative synthesis of relevant research studies. In addition, an adapted version of the COSMIN  checklist  Statement  database (registration number: CRD42017069244).Systematic review methods and inclusion criteria were documented in a protocol consistent with the PRISMA-P guidelines, as recommended by Shamseer et al. . After iContained primary data relating to a mental health assessment or intervention that incorporated information about an idiom/CCDThe idioms/CCDs addressed were linguistic (i.e. expressed in the form of language rather than behaviour)Were available in English, French or SpanishDiscussed how idioms/CCDs were incorporated into a psychosocial assessment measure or interventionStudies were deemed eligible if they:The databases Scopus, PsychInfo, MEDLINE, Web of Science, Sociological Abstracts, CINAHL and ASSIA were searched with no time period limits. The databases were last searched on 19 October 2018. Databases were searched using terms relating to: idioms/CCDs such as ‘local concepts’; intervention and assessment such as ‘measurement’ and ‘treatment’; the CCDs listed by DSM-5 . In order to establish coherence of understanding of the inclusion criteria, an initial sample of 10 studies was selected at random for the researchers to screen. They compared their results before carrying out independent screening of all articles. The researchers scanned the abstracts and titles before examining the resulting full texts for eligibility based on the pre-determined criteria. Any disparities between the researchers were resolved by thorough discussion of the inclusion/exclusion criteria.Aims of the studyPopulation and locationMethods of collection of information on idioms/CCDsLanguage the research was carried out inTerminology used by researcher(s) to refer to idioms/CCDsMethods/frameworks used to develop the assessment measure, or to integrate idioms/CCDs into existing assessment measures and/or psychosocial/psychological interventionsTherapeutic models  that guided the content of the interventionsHow translation was conducted (if applicable)Whether idioms/CCDs were compared with DSM/ICD categoriesResults of piloting/psychometric property testingAn extraction form was developed based on the research questions. Information was extracted by one researcher (C.C.) on the following aspects of each study:et al., et al., http://www.cosmin.nl).The ‘cross-cultural validity evaluation’ section of the COSMIN checklist    inventory   of distress’ . Most discussed developing novel assessments for the same cultural group from whom the idiom/CCDs data were gathered  a concept closely related to Hwa-Byung  framework  and (B), the expertise of the translators was adequately described. In papers in category (C), all except for one  demonstrated inadequate levels of reliability and validity; in Bass et al. ; in Rasmussen et al. , primary qualitative research was the most commonly used method. The justification for the choice of study language varied across the studies. In two studies, research was carried out in the preferred language of participants , new assessment measures were developed by creating items based on signs and symptoms identified in qualitative interviews. For studies that created local assessment measures for constructs that had been identified elsewhere like depression or anger attacks, they either created new scales  created a symptom scale of locally identified idioms of distress and other ‘universal terms’ that correlated with the diagnosis of depression in the DSM. They concluded that, ‘only certain idioms of distress are useful in detecting pathological states such as depression’ (p. 147). In such circumstances, it is important to be aware of the risk of category fallacy , some papers did not sufficiently discuss translation, with six studies failing to detail translation processes of idiom/CCD terms  may be occurring in the Global North. This is consistent with the previously noted observation that none of the CCDs listed in the glossary of the DSM-5 are English language terms, and none are deemed to originate in North America or Europe. These descriptions of idioms/CCDs do not form part of the taxonomy of mental illness in the main text but are instead restricted to the appendix  (Applied Mental Health Research Group, et al. (The current review had a number of limitations. First, the terminology used to refer to idioms/CCDs varied greatly between papers and included many different terms. This is similar to what Kohrt et al.  found inThe authors recognise the important role of ethnographic research methods for examining the utility and validity of idioms/CCDs in assessments and interventions. However it was beyond the scope of the current review to conduct a thorough examination of the ethnographic methods employed across the studies, which is a potential limitation of this review. A further potential limitation of the current review is that it included only published articles and the included papers were selected according to the parameters of our inclusion criteria. Thus, many studies relevant to this topic of idioms/CCDs were not included.A final limitation relates to the fact that the scope of the current review was specifically limited to the linguistic translation of narrative descriptions of idioms/CCDs. A key focus was the attention allocated to processes of translation when integrating idioms/CCDs into assessment measures and interventions, however it should be noted that cross cultural mental health related work may well involve working with idioms that are ‘untranslatable’ (Kirmayer & Swartz, et al., emic/bottom-up development of assessment measures developed in local contexts of the Global South. This is in keeping with calls for more equitable exchanges of knowledge about mental health and wellbeing between the Global North and the Global South (White et al., We advocate for the integration of idioms/CCDs into assessment measures and interventions to increase understanding of forms of suffering, improve clinical communication and treatment outcomes, and reduce stigma (Kaiser


Expression profiles for upregulated and downregulated mRNAs in RAS-STCs were functionally interpreted by gene ontology analysis. STC activation was assessed by counting the total number of STCs in pig kidney sections using flow cytometry, whereas cell proliferation was assessed in vitro. Results: Of all expressed genes, 1430 genes were upregulated and 315 downregulated in RAS- versus Normal-STCs. Expression of selected candidate genes followed the same fold change directions as the mRNAseq findings. Genes upregulated in RAS-STCs were involved in cell adhesion, extracellular matrix remodeling, and kidney development, whereas those downregulated in RAS-STCs are related to cell cycle and cytoskeleton. The percentage of STCs from dissociated kidney cells was higher in RAS versus Normal pigs, but their proliferation rate was blunted. Conclusions: Renal ischemia and hypertension in swine induce changes in the mRNA profile of STCs, associated with increased STC activation and impaired proliferation. These observations suggest that RAS may alter the reparative capacity of STCs.Background: Scattered tubular-like cells (STCs) proliferate and differentiate to support neighboring injured renal tubular cells during recovery from insults. Renal artery stenosis (RAS) induces renal ischemia and hypertension and leads to loss of kidney function, but whether RAS alters renal endogenous repair mechanisms, such as STCs, remains unknown. We hypothesize that RAS in swine modifies the messenger RNA (mRNA) profile of STCs, blunting their in vitro reparative capacity. Methods: CD24+/CD133+ STCs were isolated from pig kidneys after 10-weeks of RAS or sham ( Renal tubules have a limited endogenous repair capacity that is circumscribed to the replacement of cells that are lost during injury. Adult differentiated tubular epithelial cells that survive episodes of acute kidney injury (AKI) undergo a process characterized by upregulation of dedifferentiation and injury markers to become scattered tubular-like cells (STCs) ,2. PreviExogenous delivery of STCs can regenerate tubular structures of different portions of the nephron and ameliorate renal dysfunction in animal models of acute and chronic renal damage. For example, administration of STCs ameliorated morphologic kidney damage in murine models of AKI  and promRenal artery stenosis (RAS) is an important cause of secondary hypertension and renal dysfunction that produces a reduction in renal blood flow (RBF) and chronic ischemia ,9. RAS aDomestic pigs underwent unilateral RAS or sham and were studied in vivo 10 weeks later. STCs were isolated from fresh pig kidneys and characterized, as previously shown ,12. PorcThe mRNAseq data of each individual sample generated in this study are available as a mRNAseq analysis identified a total of 1745 genes dysregulated in RAS-STCs, among which 1,430 were upregulated and 315 downregulated compared to Normal-STCs A.p < 0.05) have an important cellular and organelle component  have also primarily cellular and organelle components . Expression of selected candidate genes followed the same patterns as the mRNAseq findings, where Dachsous Cadherin-Related 1 (DCHS1), Olfactomedin 4 (OLFM4), and Pleckstrin Homology Like Domain Family A Member 3 (PHLDA3) were upregulated and Centromere Protein F (CENPF), Claspin (CLSPN), and RNA Binding Motif Protein 41 (RBM41) were downregulated in RAS-STCs B.p < 0.05). Immunohistological staining, with antibodies against CD133 and CD24 and the tubular cell marker phytohemagglutinin (PHA)-E, was performed to identify STCs in frozen pig kidney sections. CD133+/CD24+ cells were detected in proximal tubules (PHA-E) in situ A. STC acIn the current study, we used a swine model that recapitulates human RAS to test the hypothesis that chronic renal ischemia and hypertension alter the gene expression profile of STCs. RAS is characterized by repeated episodes of insults mimicking AKI ,14, and COL6A2, COL18A1). STCs display a strong expression of structural proteins like collagen, which serve as an attachment system that anchors the basal membrane of the epithelium to the underlying extracellular matrix [Also of note is the up-regulation of mRNAs implicated in extracellular matrix remodeling, like several genes encoding for collagen  genes. Previous studies have shown that in response to acute hypoxia or toxic injury, there is a rapid and robust proliferation of surviving epithelial cells, which is accompanied by the expression of SOX genes [Renal ischemia also upregulated STC genes involved in kidney development, such as sex-determining region Y-box  and those with helicase-like activity . Chromosomes undergo continuous morphological changes as cells advance through the cell cycle, which are in part orchestrated by SMC genes [Contrarily, genes downregulated in RAS-STCs are related to cell cycle, including genes implicated in the structural maintenance of chromosomes  and xylazine (2 mg/kg), and anesthesia was then maintained with intravenous ketamine (0.2 mg/kg/min) and xylazine (0.03 mg/kg/min). Unilateral RAS was induced in 3 animals (RAS group) by placing a local irritant coil in the main renal artery using fluoroscopy , which leads to the gradual development of unilateral RAS over a 7–10-day period, as reported previously [All animal procedures were approved by the Institutional Animal Care and Use Committee . Six female domestic pigs  were studied for 10 weeks. We opted for using female pigs to test whether the effects on hypertension and renal ischemia on STCs outweigh any gender-specific protection. At baseline, animals were anesthetized with intramuscular tiletamine hydrochloride/zolazepam hydrochloride , as previously described ,33. FollA few days after MDCT studies, animals were euthanized with sodium pentobarbital . The kidneys were removed using a retroperitoneal incision, immediately dissected. Kidney cells were then dissociated and the number of STCs quantified by flow cytometry. STCs were isolated from fresh pig kidneys, cultured, characterized, and RNA isolated for mRNA sequencing analysis.STCs were isolated from fresh pig kidneys, as previously shown ,12. Brie2, Illumina cBot and cBot Paired-end cluster kit). STCs were sequenced on an Illumina HiSeq 2000 using TruSeq SBS kit version-3 and HCS v2.0.12 data collection software [mRNA sequencing analysis was performed as previously described . Sequencsoftware ,38,39.p-value smaller than 0.05 . Genes up-regulated and down-regulated in RAS-STCs were classified by their cellular component and molecular function using Protein Analysis Through Evolutionary Relationships (PANTHER) [http://david.abcc.ncifcrf.gov/) [http://bioinfogp.cnb.csic.es/tools/venny/) to visualize the total number genes upregulated or downregulated in the top functional category, whereas Search Tool for the Retrieval of Interacting Genes (STRING) version 9.1 (http://string-db.org/) was used to predict associations between these genes.Raw Fastq files were processed through Mayo’s internal MAP-RSeq pipeline  (VersionPANTHER) . The Datrf.gov/) ,46 was uDCHS1, OLFM4, PHLDA3, CENPF, CLSPN, and RBM41  in Normal- and RAS-STCs. Gene expression was normalized to GAPDH.RNA isolation, cDNA synthesis, and qPCR using the ΔΔCt method were used to randomly evaluate gene expression levels of 2 area) using bright field area vs. aspect ratio features. CD133 and CD24 positive events were quantified using flow-gaiting strategy: number of double-positive CD133 and CD24 events/number of singlet events between 10–20 µm diameter and expressed as a percentage. To detect STCs in frozen pig kidney sections, we performed double-staining of CD133  and CD24 , and the tubular marker phaseolus vulgaris erythroagglutinin  [STC activation was assessed by counting the total number of STCs in pig kidney sections using flow cytometry  with CD133  CD24  antibodies [To assess cell proliferation, STCs were harvested in M199 media with 3% FBS and 2500 cells/well seeded onto a 96-well tissue-culture plate. Cell confluence was imaged in real-time by phase-contrast using the IncuCyte Zoom  system30, 31. Frames were captured at 2-h intervals from 60 h, analyzed using the integrated confluence algorithm, and proliferation growth curves constructed using IncuCyte™ Zoom software.t-test. Statistical significance for all tests was accepted for p < 0.05. Repeated-measures ANOVA, which takes measurements on the same experimental unit over time, taking into account the probability that measurements for a given experimental unit will be correlated in some way [Statistical analysis was performed using the JMP Pro 14.0 software . Results are expressed as means ± SD. Comparisons among groups were performed using Student’s some way , was use


Twins with discordant growth have increased risks of perinatal mortality and morbidity. Previous studies have identified a number of risk factors for inter-twin birth weight discordance, yet no study has examined the effect of maternal hepatitis C infection.We used the twin birth records extracted from the 2011 to 2015 United States birth records created by the Centers for Disease Control and Prevention. The outcome variable of this study was inter-twin birth weight discordance, defined as [ / birth weight of larger twin]. The independent association of hepatitis C infection with birth weight discordance was examined using the gamma regression or log binomial regression, adjusted by potential confounders.P<0.05).Of the 270,256 twin pairs included in the final analysis, 850 (0.31%) had positive hepatitis C. Compared to mothers without hepatitis C, mothers with hepatitis C positive tended to have higher risk of birth weight discordance, but with no statistical significance. After adjustment for potential confounding factors, hepatitis C positive became a significant risk factor for birth weight discordance >25% . Sensitivity analyses (by treating birth weight discordance as a continuous outcome or dichotomizing into by different cutoffs) yielded similar results, with relative risks ranging from 1.07 to 1.12 (all Maternal hepatitis C positive is associated with inter-twin birth weight discordance, an important adverse infant outcome in twin pregnancies, although the effect size is small. Approximately 16% of twin pregnancies have birth weight discordance of at least 20%  × 100%. The confounding factors considered in this study included maternal age, race, educational level, the Special Supplemental Nutrition Program for Women, Infants, and Children (WIC), parity, assisted reproductive technology (ART), diseases prior to or developed during pregnancy , body mass index  or relative risk (RR) were used to represent the effect size of HCV infection.All potential confounding factors associated with HCV infection and birth weight discordance were included in the adjusted models. We used generalized linear models to examine the independent association of HCV infection with birth weight discordance. For dichotomized discordance, log binomial regression was used; while for continuous discordance, Gamma regression was used. General linear model was not used in our study because of the poor model fit attributed to the skewed distribution of birth weight discordance. Adjusted regression coefficient (2), normal (18.5−24.9 kg/m2), overweight (25.0−29.9 kg/m2), and obese (≥30.0 kg/m2); weight gain during pregnant was divided by tertile: 1st tertile (<30 pounds), 2nd tertile (30−42 pounds), and 3rd tertile (≥43 pounds). Some of the mother’s education and race groups in the original files were combined in order to enhance the power and robustness of estimation. All reported congenital anomalies  were combined as a single variable. Twin match and data analyses were performed using the software of SAS version 9.4 .In the adjusted models, confounding factors were categorized as the followings: maternal age<25, 25−34, and ≥35, BMI underweight  were successfully matched. After excluding records with missing information on outcome and critical co-variables, 540,512 twin records  were included in the final analysis . The meaOf the 270,256 twin pairs, 850 mothers (0.31%) had positive HCV infection. Compared to mothers without HCV, mothers with HCV infection tended to be younger, more likely being white, with lower education, more likely being WIC recipients, less likely to use ART to conceive, more likely being multiparous, with lower BMI and gained less weight during pregnancy, with STDs and hepatitis B infections, shorter gestational age, lower total birth weight (combined birth weight of the twins), and higher congenital anomaly rate . On the P values <0.05).After adjustment for potential confounding factors, as shown in Our study based on a large cohort of twins in the United States demonstrated that twins born to mothers with HCV infection had significantly higher risk of developing inter-twin birth weight discordance than twins born to mothers without HCV infection, after adjusting for a number of known risk factors of birth weight discordance. This association remained essentially the same using different cut-off points or treating birth weight discordance as a continuous outcome variable. To our knowledge, this is the first study that has examined the association between maternal HCV infection and inter-twin birth weight discordance. The large sample of our study allowed a stable estimation and adjustment for a number of risk factors simultaneously. The association between maternal HCV infection and inter-twin birth weight discordance became stronger and statistically significant after adjusting for a number of confounding factors, including maternal demographic factors. We acknowledged that the effect size of HCV infection on birthweight discordance was not very large for clinical importance. However, it is still a problem, from scientific point of view, that may still worth exploration.A few studies have assessed the association between maternal HCV and infant outcomes other than inter-twin birth weight discordance –18. EarlOur study has limitations. Because universal HCV testing in pregnancy is not mandatory in the United States, mothers recorded with HCV positive may have increased risks for care providers to initiate test, which could introduce ascertainment bias. However, there is no data reporting the rate of antenatal HCV test. According to a previous report, the rate of HCV screening was 4.31% in population aged above 8 years old . During HCV infection in pregnancy represents a non-negligible health problem for unborn fetus, and our study adds to the literature by highlighting its potential impact on inter-twin birth weight discordance, an important adverse outcome to be prevented in twin pregnancies. Regardless the mechanisms, the increased risk of inter-twin birth weight discordance for twins born to these mothers suggests that extra attention should be paid in the care of them. To prevent transmission of HCV, it is important that health care providers adhere to standard precautions, follow fundamental infection-control principles, and use appropriate procedural techniques. Because no safe and effective method exists to reduce vertical transmission of HCV once a woman becomes pregnant, it is important to identify and treat HCV before pregnancy , 24. In S1 File(PDF)Click here for additional data file.


The retroperitoneal Müllerian cysts are extremely rare lesions seen mostly in female patients but their occurrence in males is quite possible. Their exact etiology stills unknown, its histological and immunohistochemical characteristics may serve as an evidence that reflects its origin.We present a case of a resected retroperitoneal Müllerian cyst that reoccurred severely after 3 months of the first laparotomy in a male patient with a significant history of testicular teratocarcinoma. Due to the cyst’s localization and its proximity to vital retroperitoneal structures, total surgical resection couldn’t be made and the recurrence was minimized by post-operative oral chemotherapy.The retroperitoneal Müllerian cyst is a benign cystadenoma that could be confused with other retroperitoneal lesions, which makes the pathological examination with the immunohistochemical study of the cyst’s wall essential to make the diagnosis. The immunopositivity to Cytokeratin 7 and the immunonegativity to Cytokeratin 20 is a key feature that confirm the diagnosis whenever the retroperitoneal Müllerian cyst is suspected. Due to its high vascularity, we highly recommend the administration of chemotherapy which targets the proliferative cyst’s cells.The RMCs are rare benign lesions that tend to reoccur if total surgical resection isn’t made, when the total resection couldn’t be achieved, the-unspecific anti-mitotic drugs may help in minimizing the recurrence and improve the life quality of the patient. His past medical history was significant for testicular teratocarcinoma which was treated by orchiectomy followed by a course of eight shots of chemotherapy. A computed tomography scan (CT scan) revealed a large multilocular cystic formation arising from the retroperitoneum, occupying the entire abdomen, displacing the abdominal organs to the right. In addition, a left hydronephrosis was noticed due to compression of the left ureter .Fig. 1ThAs the cyst’s nature was not clear and several aspiration attempts failed to relief the symptoms due to the cystic septations, an emergent debulking surgery was performed. A midline incision was made, but due to the large size of the cyst and its close proximity to the retroperitoneal viscera and vasculature, an “en-bloc” resection could not be performed, so aspirating the cystic fluid followed by resecting the cystic septations were the only way to achieve the removal of the cyst from the retroperitoneum. At the end of the operation, 20 liters of serous fluid were aspirated and the majority of the cyst’s septa were resected. Macroscopic examination revealed a multilocular cystic mass composed of smooth surfaces without any papillae or vegetations. Microscopic examination revealed that the cyst was lined with a single layer of columnar epithelium with normal nuclei; no atypia was noticed, beneath the epithelium, a fibrous stroma, smooth muscle bundles, and extensive vasculature were found .Fig. 2ThImmunohistochemically, the cyst’s epithelium was immunoreactive to cytokeratin 7 (CK-7), epithelial membrane antigen (EMA) and Wilms tumor 1 (WT1), and it was immunonegative for the progesterone receptor (PR) and cytokeratin 20 (CK-20), while only the stromal cells were positive for estrogen receptors (ER), therefore, a retroperitoneal Müllerian cyst was diagnosed.After the surgery the patient was only stable for 3 months before the cyst reoccurred severely to occupy the entire abdomen again. The cyst’s recurrence was confirmed using the CT scan , in addiAs the benign nature of the cyst was confirmed pathologically, another laparotomy was performed with wider resection field. The inferior vena cava, the abdominal aorta, the kidney and the ureter were heavily isolated and dissected from the cyst’s septa .Fig. 4. After 6 months of follow up the patient was stable but a minimal cystic recurrence was detected by the CT scan .Fig. 5Th3Retroperitoneal Müllerian cysts (RMCs) are extremely rare lesions of the retroperitoneum which occur mainly in female patients , but as In the human embryo, the Müllerian ducts are a pair of tubes that are formed at the lateral aspect of the urogenital ridge during pregnancy. In male embryos, they undergo an incomplete regression under the influence of the Anti-Müllerian hormone secreted by the Sertoli cells of the developing testis, leaving the prostatic utricle and the appendix of testicles as embryological residues, whereas in female embryos, they continue their development to form the major parts of the female reproductive tract.One hypothesis suggests that incomplete regression of these ducts can leave an asymptomatic remnant which have the capacity of growing under certain hormonal estrogenic stimulations, causing this cystic formation , as the Whatever the origin of these cysts is, it doesn’t change the fact that the pre-operative diagnosis is quiet hard because the clinical examination lacks of information and the radiologic findings are unspecific.To the best of our knowledge, the only way to confirm the diagnosis of RMC is examining the pathological and immunohistochemical features of the cyst, some authors considered that the multiple layers of the cyst’s wall in addition to the presence of smooth muscle bundles is a proof that RMCs emerged directly from the Müllerian apparatus . AlthougCurrently, the only curative treatment that has been applied is the total surgical resection, other treatments such as percutaneous aspiration, resulted in the recurrence of the cyst ,4. Even 4The RMC is a rare lesion that can occur in male patients. The final diagnosis is made after examining the cyst's wall microscopically and studying its immunohistochemical profile. The cyst's immunopositivity to CK-7 and immunonegativity to CK-20 have an essential role in differentiating it from other retroperitoneal cystic lesions. Some types of the RMCs may have a good vascular supply which encourage the administration of the anti-mitotic drugs to prevent further growth and recurrence.No funding sources.No ethical approval was needed as we didn’t use any experimental drug or new surgical techniques to treat the patient. We combined multiple classical methods of treatment to achieve the best outcome and prevent further recurrence as long as possible.Written patient consent was taken before reporting his case and sharing his pre-surgical, surgical images and the CT scan radiologic images.Antoine Naem: wrote the abstract, introduction, discussion, conclusion, provided the language-editing services and all the captions of the figures.Ammar Dlewati: wrote the presentation of the case, reviewed the literature and designed all the figures.Maryam Alhimyar: supervised the literature review and data extraction process.Mohamad Ali Ousta: participated in the surgical treatment and collecting the patient’s data.Bayan Alsaid: participated in the surgical treatment and supervised the scientific and academic aspects of the manuscript preparation and submission.N/A.A guarantor of data is the corresponding author, Prof. Bayan Alsaid.Editorially reviewed, not externally peer-reviewed.No conflict of interests.


We analyzed the association between birthweight, nutritional status and transverse maxillary growth in 7- to 9-year-old schoolchildren. We undertook a cross-sectional survey nested in a population-based cohort study of 158 schoolchildren. The participants lived in the urban area of a small town within the Western Brazilian Amazon. The outcome was represented by the upper intermolar distance given in millimeters (mm), as an indicator of the degree of maxillary bone growth in its transverse dimension. The exposures were sex, birthweight, the bottle-feeding pattern operationalized by a scale corresponding to the age of introduction of the bottle and Body Mass Index-for-age z-score (BAZ) at 4 to 6 ys. Path analysis was employed to estimate standardized direct, indirect and total effects of exposures on the outcome using structural equations model (SEM) supported by Mplus 7 program. The values of standardized coefficients (SC) showed significant direct positive effects of sex , birth weight  and BAZ  on transverse maxillary growth. The indirect effects  and the total effect  of sex on the outcome were statistically significant. The indirect effects of birth weight on the outcome were not significant , however, the total effect was significant . In conclusion, sex, birthweight, bottle beginning age and BAZ showed association with the transverse growth of the maxillary bone. In addition to contributing to an adequate birth weight of the child, policies and programs that favor prenatal care and conditions to guarantee a full-term birth can positively affect transverse growth of the maxilla. From a Public Health Surveillance point of view, children with reduced birthweight, inadequate breastfeeding pattern and nutritional deficit for age may be more likely to develop atrophy of the jaws which, depending on the severity, may result in malocclusion with an important impact on quality of life. Malocclusion is a multifactorial alteration of the normal growth and development with effects on muscles and facial bones during childhood and adolescence; it is a collection of situations, any of which can aggravate due to genetic predisposition or external factors. It can affect the oral functions, such as mastication, swallowing and speech; it implicates greater susceptibility to trauma and periodontal disease; and it can also have an important psychosocial repercussion due to impaired dentofacial esthetics.Although much epidemiological information is the result of the analysis of cross-sectional studies, including the possible bias inherent in such research, it is recognized that the prevalence and severity of malocclusions have increased in the last 200 years, especially dental crowding, which is the most prevalent type of malocclusion , 2. MaloThe maxilla is a complex anatomical structure whose growth process is given by multiple factors, such as sutural growth until 7 years old. After this age it gets slower and is more characterized by areas of apposition and resorption, simultaneously accompanied by displacement forward and downward. It is accompanied by a calcification process, tooth eruption and the intervention of muscular forces that harmoniously promote proper development. The eruption of the teeth and the consequent growth of the alveolar process will increase the vertical dimension of the maxilla. With the eruption of the first permanent molars begins the mixed dentition, which extends from 6 to 12 years, and it is a period of particular importance in the etiology of malocclusion since during these years a series of interconnected processes lead to changes of temporary by permanent dentition; and definitive occlusion is established. At 6 years old, the upper first permanent molars erupt behind the second temporary molar. During the time of temporary dentition the width of the dental arch increases slightly. The main increase of the arch occurs by growth in a posterior direction as the teeth erupt, this expansion is detected in the same way in permanent dentition. The expansion of the upper dental arch width is greater than in the lower one .McNamara reported normal maxillary width of 35mm in mixed dentition measured from the lingual surfaces. Values from 36 to 39 mm could accommodate a dentition of average size without crowding or spacing, whereas maxillary arches less than 31 mm in width might be crowded and thus in need of orthopedic or surgically assisted expansion [nature and nurture [Potential factors associated with changes in maxillary growth include genetic traits , mouth breathing, decreased oral functions, premature loss of temporary teeth, infantile facial paralysis, delayed onset of complementary feeding, muscular hyper or hypotonicity (in the orbicular muscles of the mouth), non-nutritive sucking habits, atypical phonation and swallowing, and abnormal tongue position or other environmental factors, thus demonstrating an interaction  nurture , 8. ThesPrevious studies conducted on this subject are limited, therefore epidemiological information is scarce and mostly generated from cross-sectional designs preventing a view on the role of certain characteristics of the child's life course, a perspective that has become important with the emergence of theories about the early life influence in the health status in later years and in the rest of life. It has been hypothesized that aspects of the perinatal period and childhood could promote a "biological programming" associated with development of chronic diseases years later. David Barker, one of the pioneers in this subject, proposed that the first stages of early life were a critical period that permanently programs body metabolism and growth, and thus determines pathologies in adulthood, in addition to being related to future socioeconomic conditions. Several relationships between circumstances of early life and consequences at different times throughout life have been studied , 10.The aim of the study was to investigate the factors associated with transverse growth of maxillary bone, expressed by the upper intermolar distance, from a point of view related to the course of life.The study was approved by the Institutional Review Board of the School of Public Health, University of Sao Paulo, Brazil (297/2009). Written informed consent was obtained from the parents or guardians of children who participated in the study before enrollment. Data were analyzed anonymously.It links to a larger research project on health and nutrition conditions at Acre state developed by Sao Paulo University and Acre Federal University, whose field activities began with population-based cross-sectional surveys in 2003 in the urban area of a small town within the Western Brazilian Amazon . The chaThe reference population described elsewhere  compriseSchoolchildren of both sexes, with no extensive occlusal caries in upper permanent molars; no history of orthodontic or major surgical treatment; no congenitally missing teeth; no signs or symptoms of temporomandibular joint disorder, no mouth breathing, no diseases such as systemic and/or neurological ones and those whose responsible person accepted the dental examination, were eligible for the study.The dependent variable was represented by the upper intermolar distance given in millimeters, measured between the center point of the occlusal surface of the right and left first permanent molars (mm), assuming that it can operate as an indicator of the degree of maxillary bone growth in its transverse dimension , 5. In 2The independent variables were related to the characteristics of the child. Sex, birth weight and the bottle-feeding pattern operationalized by a scale corresponding to the age of introduction of the bottle, were obtained in a population-based cross-sectional study involving children aged 0.1–5.5 years at 2003 . The botThe descriptive analysis of the data was performed with the program Stata/SE 13.1. It was tested adherence to the normal curve of the dependent variable through the Kolmogorov-Smirnov test. The t-test was performed to compare the mean values of the intermolar distance between sexes, as well as differences in birth weight and BMI-for-age z-score between sexes.Statistical analyzes of the data were carried out using the path analysis approach in the framework of a structural equations model (SEM) with aid of the Mplus 7 program . The RobThis modelling allows to verify a theoretical model, in which the hypothesized relationships among variables are tested in a linear equation system. A variable can be defined as an outcome variable in one relation and as an explanatory variable in another relation. The study of these multiple relations allows testing several hypotheses where an association of these variables with the growth of the maxillary bone, would be present. Firstly, as in previous studies, boys are expected to have a higher birth weight than girls . ChildreThe hypothesis supporting an association between bottle beginning age and transverse maxillary growth was based on studies that have produced evidence that bottle-feeding is detrimental to the development and function of oral maxillofacial structures –28. In aThe adjustment of the structural equations model was performed using the goodness of fit indicators: Comparative Fit index (CFI) with reference value higher than 0.95; Tucker Lewis Index (TLI) with values above 0.95; and the root mean square errors of approximation (RMSEA) expecting values less than 0.05.As no previous information is available, the study was considered exploratory. No a priori hypothesis was admitted about presumed associations and no study power calculation was performed hoping that subsequent research can be designed with greater precision knowledge.The study sample selected from the reference population consisted of 158 children between 7 and 9 years of age, of whom 73 were male (53.8%) and 85 were female (46.2%). Thirty-seven were no eligible and twenty-three had missing data on specific variables. The age distribution in the sample was given by 48 children aged 7 years (30.4%), 62 children aged 8 years (39.2%) and 48 children aged 9 years (30.4%). A proportion of 8.9% (n = 14) of children with low birth weight  was found. Of these, 10 (71.4%) were female.The upper intermolar distance showed adherence to the normal distribution (p = 0.625). When comparing the mean values of the upper intermolar distance between the two sexes, a statistical difference was found (p = 0.001); with the mean in boys being 46.25 mm and in girls being 44.63 mm. Comparing the differences between the mean values of the upper intermolar distance between the children's ages , there were non-significant statistical differences.Several relations between the variables, both direct and indirect, were observed in the SEM . The modThe four independent variables considered in the study  had a direct and significant association with upper intermolar distance. The overall effect of sex on the outcome was significant . The direct positive effect of sex on the intermolar distance reflects that the boys presented higher values of this measure indicating, therefore, a greater transverse growth of the maxillary bone in this group . The t-test showed that for boys the average intermolar distance was 1.627 mm higher than for girls (p = 0.001).The positive direct effect of birth weight on the outcome indicates that the higher the birth weight of the child, the higher the intermolar distance . Likewise, the higher the BMI-for-age z-score, the higher the intermolar distance .The bottle-feeding pattern scale showed a negative direct effect on the outcome. Given that this variable assigns a value from 1 to 10 (where 1 is for those who never used a bottle and 10 for who started to use a bottle between 0 and 14 days of age) as seen in The indirect effects produced by the variables on the outcome can be seen through several paths among the independent variables. Starting with sex, which is the most distal variable with respect to intermolar distance, we observed the direct effect and five ways that contribute to the indirect effects of this variable on the object of study. As shown previously, the direct effect was significant . The set of the five indirect effects was also significant . However, when analyzing each indirect effect separately, it is observed that only one of them of them was significant .Regarding the effects of birth weight, besides the direct effect, two paths were found through which indirect effects were observed. As shown previously, the direct effect of birth weight on the upper intermolar distance was statistically significant . The indirect effects together  or each path separately did not show statistical significance , but theBottle feeding pattern showed an indirect effect way with no statistical significance . The total effect that summarizes direct and indirect effects was significant .Sex, birth weight, bottle-feeding pattern and BAZ were associated with upper intermolar distance. The effect of each variable on the outcome was expressive. In addition, indirect effects of these variables on the upper intermolar distance were identified.As strengths of the work, the characteristic of being a cross-sectional study nested in a cohort and the utilizing of structural equation models could be quoted, since this combination allows to determine the temporal sequence and to study several regression models simultaneously for establishing direct and indirect relationships among the considered variables.Although the study has yielded new insights into the growth of the maxilla associated with nutritional and health determinants in early childhood, and may be a useful tool for further research to be developed in populations with other characteristics, some limitations should be highlighted. Socioeconomic variables, skeletal pattern, altered jaws and teeth sizes, type of respiration and swallowing, complementary feeding and non-nutritive sucking habits were not investigated at first stages of children’s development; and it would be quite important to study these aspects. Excepting non-nutritive sucking habits, there are scarce information about their impact on the outcome. Impaired oral functions could be etiological factors of the malocclusion as pressures and forces exerted during these functions could influence the growth of the jaws and tooth eruption . When atThe effect of sex on upper intermolar distance had a significant overall effect with higher values for boys. These results are consistent with those obtained by Aznar et al. from a cross-sectional study in a sample of 1,297 children aged 3 to 6 years in Spain. Their results pointed to canine and intermolar distances—both in the maxilla and mandible—significantly higher in boys. However, the authors did not raised hypotheses to explain it . PreviouIn addition to the direct effect of sex on the upper intermolar distance, five indirect effects were found in the structural model, which together showed statistical significance. These findings demonstrate that maxillary growth depends on multiple factors, in addition to the genetic influence that is always present. The results regarding the way in which birth weight influences transverse maxillary growth, indicate that the manifestations on the outcome depend on a combined action due to the duration of breastfeeding and physical characteristic of the child measured by BMI-for-age z-score.Monitoring birth weight is important to avoid risks related to neonatal and infant morbidity and mortality. However, the relationship of birth weight and transverse maxillary growth had not yet been explored. Regarding the effects of birth weight on the outcome, it was observed that the birth weight showed significance by itself and also in the presence of other variables, suggesting that the birth weight influences the transverse growth of the maxilla. Although the indirect effects of birth weight on the upper intermolar distance were not significant, the total effect was.As expected, boys presented a higher birth weight than girls. Infants with a higher birth weight could have higher values of BMI during childhood compared to those with reduced birth weight. Children with low birth weight have smaller dental crowns and poor-quality enamel –24. ThisAssociation between the infant feeding pattern and the BMI z-score has been previously studied which varies as the duration and exclusivity of breastfeeding changes and the introduction of other types of foods occur. The association changes in magnitude as the feeding pattern changes as well . In thisThere is evidence of the detrimental effect of bottle feeding on the development and function of oral maxillofacial structures –28. It cIn conclusion, girls and boys with reduced birth weight, very early bottle beginning age and diminished BAZ are more likely to have decreased transverse maxillary growth. The results suggest that policies and programs of child health promotion can have a significant effect on transverse maxillary growth contributing to the control of malocclusions such as posterior crossbite, dental crowding and facial asymmetries.S1 Data set(XLSX)Click here for additional data file.


Background: Preterm birth is strongly associated with increasing mortality, incidence of disability, intensity of neonatal care required, and consequent costs. We examined the clinical utility of the potential preterm birth risk factors from admitted pregnant women with symptomatic preterm labor and developed prediction models to obtain information for prolonging pregnancies.Methods: This retrospective study included pregnant women registered with the KOrean Preterm collaboratE Network (KOPEN) who had symptomatic preterm labor, between 16 and 34 gestational weeks, in a tertiary care center from March to November 2016. Demographics, obstetric and medical histories, and basic laboratory test results obtained at admission were evaluated. The preterm birth probability was assessed using a nomogram and decision tree according to birth gestational age: early preterm, before 32 weeks; late preterm, between 32 and 37 weeks; and term, after 37 weeks.Results: Of 879 registered pregnant women, 727 who gave birth at a designated institute were analyzed. The rates of early preterm, late preterm, and term births were 18.16%, 44.02%, and 37.83%, respectively. With the developed nomogram, the concordance index for early and late preterm births was 0.824 (95% CI: 0.785-0.864) and 0.717 (95% CI: 0.675-0.759) respectively. Preterm birth was significantly more likely among women with multiple pregnancy and had water leakage due to premature rupture of membrane. The prediction rate for preterm birth based on decision tree analysis was 86.9% for early preterm and 73.9% for late preterm; the most important nodes are watery leakage for early preterm birth and multiple pregnancy for late preterm birth.Conclusion: This study aims to develop an individual overall probability of preterm birth based on specific risk factors at critical gestational times of preterm birth using a range of clinical variables recorded at the initial hospital admission. Therefore, these models may be useful for clinicians and patients in clinical decision-making and for hospitalization or lifestyle coaching in an outpatient setting. The overall spontaneous and iatrogenic preterm birth rates showed clinically varied country-specific rates between 5% to 13% per year over the past few decades Identifying women at risk of preterm birth is an important task for clinical care providers. However, there are only a few methods available for reliably predicting actual preterm labor in women who present with symptoms of labor. Currently, pregnant women with symptomatic preterm labor undergo a transvaginal ultrasound examination and cervicovaginal fetal fibronectin test, but these yield a very high false-positive rate which leads to increased unnecessary hospitalizations and administration of tocolytics and glucocorticoids As a part of the preterm birth management entry process, electronic systems, such as the clinical decision support systems, help determine the risk for a range of medical conditions, directly affecting the decision making and the individual patient-specific assessment and counseling. The use of these systems is effective and has a significant impact on the improvement of clinical practice In this context, we examine the clinical utility of the potential preterm birth risk factors from admitted pregnant women with symptomatic preterm labor. We developed prediction models for preterm birth and described the information obtained from the prediction models to serve as a useful guideline for prolonging pregnancies.National obstetric specialists and researchers from 20 tertiary hospitals were included in the KOrean Preterm collaboratE Network (KOPEN) registry  using the internet-based clinical research and trial management system at each tertiary hospital.Collected data included demographics, obstetric and medical histories, and basic laboratory test results, including blood tests and vaginal discharge findings for clinical and basic characteristics. In the questionnaire, sleep quality was evaluated using the Pittsburgh Sleep Quality Index. A pelvic examination was performed to assess the cervical condition, such as presence of bleeding, ripening, opening, and water leakage. A speculum examination was used for fetal fibronectin and vaginal swab culture of microorganisms, if possible. An ultrasound examination was conducted to assess the cervical length and shape, fetal gestational age and weight, presence of anomaly, presentation, amniotic fluid volume, and presence of maternal anatomical abnormalities. Blood serum samples were obtained for assessing the blood count and C-reactive protein (CRP) level.Preterm labor was defined by uterine contractions lasting 40 to 120 seconds more than two or three times per 20 minutes, or eight times within 60 minutes during electro-fetal monitoring with or without cervical dilatation. Gestational age was assigned based on the last menstrual period and confirmed in the first- and early second-trimester ultrasound examinations. Premature rupture of membranes is defined as the rupture of the fetal membranes before the onset of labor. Cervical incompetence is defined as the inability of the uterine cervix to retain a pregnancy in the second trimester without clinical contractions and/or labor We categorized three gestational age terms as follows: early preterm birth (before 32 weeks), late preterm birth (32-37 weeks), and term birth (after 37 weeks), based on the WHO preterm birth subgroup categories R language version 3.3.3 , T&F program version 2.6 , and IBM SPSS Statistics version 22  were used for statistical analyses. Data were expressed as mean ± standard deviation for continuous variables. When variables were normally distributed, the difference between the means of two sample groups, defined by the gestational age at birth, were tested using the Student's t-test or Welch's t-test as appropriate. For non-normally distributed variables, the Mann-Whitney U test was used. For categorical variables, data were expressed as numbers and percentages, n (%). The chi-squared test or Fisher's exact test was performed to test the association between the gestational age subgroups at birth and other categorical variables as appropriate using a contingency table.We developed preterm birth prediction models, devised nomograms, and evaluated the discriminatory power of the prediction models using an internal validation procedure.p-values were computed using the null hypothesis of AUC = 0.5. A p-value cutoff of 0.1 was applied to select potential variables that were used in the construction of the prediction model for preterm birth. The cutoff values for the potential variables were selected to maximize the sum of sensitivity and specificity, which were used to transform the variables to binary predictors of preterm birth.Receiver operating characteristic (ROC) curve analysis was performed to select potential variables that predict preterm birth defined by gestational age at birth. The discrimination performance of the variables was estimated as the area under the curve (AUC), and p-value < 0.2 in the univariate analysis. The discriminatory power of the constructed models was estimated using the AUC with leave-one-out cross-validation (LOOCV) performed to estimate the reliability of the constructed model through an internal validation procedure.Binary logistic regression analysis was performed to analyze the effect of each potential predictor, selected from basic statistics and ROC curve analysis, of preterm birth. Univariate analysis was performed to investigate the association between outcomes and clinical variables or questionnaire variables. To construct the best-fit prediction model for preterm birth, multivariable logistic regression analysis was performed using a backward variable selection method to determine independent covariates. The criterion for initial input variables was a To facilitate the practical application of the prediction model in the clinical field, a nomogram was developed. Significant factors from the multivariable logistic regression model were incorporated using a weighted-point system to create a clinical prediction algorithm in a nomogram format. A computer-based application program was developed to facilitate the use of individual probability of preterm birth.For practical application of the prediction model in the clinical field, a Classification and Regression Tree (CART) analysis was performed to determine the complex interactions among the candidate predictors in the final tree to build the classification trees.This study was approved by the institutional review board at Ewha Womans University Medical Center  (IRB No. 2016-04-021), and informed consent was obtained from all participants before enrollment in the study.In total, 879 pregnant women in preterm labor were registered at the 20 participating tertiary perinatal centers between March 2016 and November 2016  and objective signs  at admission, early preterm birth was associated with fewer subjective symptoms and more objective signs. The measured biologic characteristics, including shorter cervical length, higher white blood cell count, higher CRP level, and presence of ruptured amniotic membranes, were significantly associated with the early preterm birth group Table .We performed a multivariate logistic regression analysis Figure , which iOur objective was to predict the estimated time of delivery between 32 and 37 weeks of gestation Figure , where bAll variables of tables used in the tested models for the decision tree analysis for the three groups are shown in Figure . In case of multiple pregnancy , 86.4% had preterm birth . CART analysis shows that in the case of multiple pregnancy, the nodes of subjective symptoms such as more labor-like pain and feelings of uterine contractions were associated with late preterm birth; then, nodes of objective signs such as water leakage due to membrane rupture, lower hemoglobin (Hb) levels, and having an occupation at admission were related to late preterm birth.The decision tree analysis for late preterm birth showed an overall prediction rate of 73.9% Figure . The mosTo our knowledge, this is the first study where predictive models were developed for clinically assessing preterm birth periods  using information contained in an eCRF and data obtained at admission, especially subjective symptoms.Usually, management of patients by obstetricians is based on risk estimation, patient counseling, and decision making. However, commonly used risk estimation methods apply the same risk level to all patients; this approach does not offer the possibility of individualization.To eliminate this problem and to obtain more accurate predictions, researchers have developed predictive and prognostic tools based on statistical models, which have shown better clinical judgment for predicting probability of outcomes The first attempt to do this in an obstetric setting had low accuracy and could not be individualized In the present study, the nomogram-based prediction model may provide information for a personalized assessment of the likelihood of preterm birth by incorporating general risk factors either before completion of 32 weeks of gestation or between 32 and 37 weeks of gestation. We simply developed the nomogram by automatically calculating the probability for individuals using a Microsoft Excel spreadsheet (Supplementary On the other hand, the proposed decision tree provides a base for developing an antenatal preterm prevention step-by-step guide through the design, implementation, and evaluation of the stages of antenatal lifestyle interventions, such as dietary habits and physical activity levels. In the CART decision tree that we developed, good eating habits, nutrient supplementation and regular physical activity were associated with longer gestational time. Some studies reported that improving diet and physical activity during pregnancy can improve short-term pregnancy outcomes as well as long-term maternal and offspring health To this end, nomogram and CART decision trees may be helpful for obstetricians to prepare adequate advice and educate pregnant women. Nomograms are simple and noninvasive visual instruments with a graphical interface that promotes the use of prediction risk models. CART analysis is another type of predictive model with the capacity to account for complex relationships and is relatively easy to use for the clinician. Accurate estimation of preterm birth risk using prediction models improves patient satisfaction after preterm management. In particular, a small change in gestational time by delaying labor could significantly reduce neonatal morbidity and mortality by allowing for an intervention period to accelerate fetal lung maturation The main strength of our study is providing communication and education as a tool to improve treatment of patients and using currently available preterm birth data and environmental factors involved in a multicenter cohort with prospective recording variables. Our models are based on widely used criteria and a combination of well-known risk factors for preterm birth obtained by using questionnaires on subjective symptoms. Health care providers should evaluate the risks and provide appropriate information for avoiding or managing preterm birth. Our study shows there is a tendency to a prolonged gestational time in patients experiencing subjective symptoms, such as pelvic pain and sense of pelvic prolapse, rather than objective symptoms, such as vaginal bleeding and water leakage.Another strength of this study is that all the variables in our predictive models are based on data available from the clinical obstetric history, allowing for easy assessment of patients. Preterm births between 32 and 37 weeks of gestation have a relatively lower risk of mortality and morbidity than early preterm births, but the impact on healthcare worldwide may be significant due to their higher risks than full term births Many antenatal and postnatal factors modify the risk of preterm birth. Our statistical analyses show there is a selection bias due to fact that the source population consists of a registry of pregnant women with preterm labor symptoms rather than all pregnant women, including those who are asymptomatic Despite these limitations, this study developed personalized prediction models of preterm birth risk and an estimation of the delivery period using a wide range of clinical variables obtained at the initial hospital admission. Therefore, these models may assist clinicians and patients in clinical decision making so that appropriate decisions for hospitalization or lifestyle coaching in the outpatient setting can be made. This may also be useful to counsel and educate patients by calculating the overall probability of preterm birth for the individual patient and considering specific risk factors present during critical time points in the gestational period.Supplementary figures.Click here for additional data file.


There is a need to comprehensively investigate the relationship between tooth eruption and infant growth to explain the theory of tooth emergence. This study aimed to investigate the association between infant growth during the first year of life and the emergence of the permanent teeth.A random sample of 668, 12-year-old students was recruited from a birth cohort. Erupted permanent tooth number was recorded. The association of infant growth (growth trajectories and growth rates) and permanent tooth emergence was examined through logistic regression analyses. The regression model was adjusted by potential confounders including gender, gestational age, mode of delivery, type of feeding, parental education, and health status.n = 514). Two hundred and forty-five (47.7%) children had all 28 permanent teeth erupted. Infants who had higher birth weight z-scores and those who had grown slowly during the first three months of life were more likely to have complete permanent teeth emergence at their 12-year-old in both unadjusted (p <  0.01) and adjusted model . However, no significant association was found between the growth trajectories and permanent tooth emergence in either unadjusted or adjusted models (p > 0.05).The response rate was 76.9% (Birth weight and infant growth during the first three months of life might be associated with permanent tooth emergence at their 12 years of age. This association may be applied in the assessment of risk for dental caries or malocclusion. Tooth eruption is the process of a tooth moving from its site developed in alveolar bone to its functional position in occlusion . The eruDisturbances in timing or sequence of eruption may result in a chain of complications such as malocclusion, periodontal disease, and dental caries; and subsequently increase the associated dental and orthodontic treatment needs. Furthermore, the consequence extends beyond simply ‘oral health’ as it also has implications for child development  and general health status , 5. AlthEarly life programming is thought to be of great importance in later life health outcomes. In particular, it is accepted that early childhood events, especially in the first year of life, can have considerable impact not only on childhood survival but also morbidities of later life . MoreoveThe hypothesis of early life programming is supported by epidemiological evidence in humans revealing that rapid infant growth increases the risk of metabolic syndrome , earlierN = 8327) [The oral health survey was of a cross-sectional design which fused into a longitudinal Chinese birth cohort study. The participants were randomly selected from a longitudinal Chinese birth cohort ‘Children of 1997’. Eighty eight percent of all infants that were born between April 1st and May 31st 1997 in Hong Kong were recruited in the cohort ( = 8327) . After 1 = 8327) . It was The clinical examination was conducted in the student’s school within two months before the participant’s 13th birthday. In order to gain adequate visibility during examination, the students were asked to rinse their mouth before lying supine on portable dental chairs, and then their gingivae were cleaned with gauze to eliminate food debris for proper visibility through examination. Prior to the field survey, two dentists who were blind to the students’ information and the objectives of the study were trained and calibrated by an experienced Paedodontist (HMW) in clinical assessment of the emergence of a tooth. In the field study, dental examinations were performed using a plain intra-oral disposable mouth mirror with a built in light-emitting diode (LED) light source and a blunt probe. The probe was used to sense or prove the occurrence of newly erupted teeth. The diagnostic criterion for an emerged tooth was when any portion of the crown has perforated the oral mucosa and was visible through the oral mucosa . RecordsEven though the diagnostic criteria for the emergence of teeth have been clearly stated and discussed prior to the survey, it could still be difficult to determine whether a tooth has emerged or not when it was close to emergence or has just barely emerged. To guarantee the consistence throughout the study, blind duplicate oral exam was conducted among 10% participants who were randomly selected from the study sample to examine the intra- and inter-examiner reliabilities.The participants’ information on their growth and development was retrieved from the well documented medical records at MCHCs. The data of the ‘Children of 1997’ birth cohort included body weights at birth, and 1, 3 and 12 months of age; information on birth characteristics , and type of feeding. Five gender-specific growth trajectories were generated according to body weight at different time points  through latent class analysis in the whole cohort through SAS (version 9.1) . FirstlySocio-demographic status, with regards to the child’s gender and parental educational attainment, was also retrieved from the medical records at MCHCs.Data of tooth emergence status from the oral health survey at 12 years old were combined with data from ‘Children of 1997’ in terms of birth characteristics, body weight during the first year of life, growth trajectories, and socio-demographic status. Descriptive statistics were used to summarise demographic data in terms of birth characteristics, socio-demographics and permanent tooth emergence through t-test or ANOVA for continuous data, and χ2 test for categorical variables. Included participants were those with growth trajectories. Those without data on growth trajectories were excluded. Variance on demographic data between the included and the excluded students were compared.The association of infant growth with permanent tooth emergence at 12 years of age was examined through logistic regression models. The permanent tooth emergence was the dependent variable, with code 1 as the complete emergence group and 0 as the incomplete emergence group. The main independent variables were growth variables- growth trajectories and growth rates . We put these two growth variables in the model separately. The co-variables in the study, obtained from MCHCs were: (a) birth characteristics: gestational age  and mode of delivery ; (b) health status  highest parental educational attainment ; (d) type of feeding , and (e) student’s gender. Furthermore, models were additionally adjusted for birth weight z-score so as to differentiate the growth effect irrespective of the birth weight. The OR was reported with 95% confidence intervals (CI).Further analyses at the tooth level were performed if significant differences were found in logistic regression analyses at the subject level. Statistical analysis was performed using IBM SPSS Statistics 20.0 .p <  0.05). Children included in the study had higher proportion of natural birth and less proportion of caesarean birth (p <  0.05). No significant difference was found in other information in terms of gender, health status at birth, type of feeding, parental education attainment, or oral health data among participants and non-participants (p > 0.05) (Table The response rate was 76.9% as 514 written consents were obtained from 668 parents. Four hundred and eighty-five of those 514 students had the complete profile on birth characteristics (Table 5) Table .Table 1BAmong the 485 students, 232 (47.8%) children had 28 permanent teeth emerged. Second molar had the highest rate of un-emergence (14.2–32.2%), followed by second premolar (5.4–5.6%), canine (0.8–3.3%), first premolar (2.3–2.5%), lateral incisor (0.8–1.4%), central incisor (0–0.8%), and first molar (0.2%), see Table p > 0.05, Tables p > 0.05, Tables Significant association was found between the emergence of permanent tooth and growth rates from birth to 3 months. Unadjusted logistic regression model identified that children with less changes of weight-for-age z-score from birth to 3 months had significantly higher chance of having complete permanent tooth emerged than those with great changes , Table Results from the tooth level in bivariate and multivariate analyses showed that students who had a slow growth rate from birth to 3 months of age had a higher chance having the maxillary second molars erupted at their 12 years of age Tables  and 7.Ta, while two growth parameters were employed to represent the nutritional status during infancy. We provided information and conducted statistical analyses on i) five growth trajectories during the first year of life which was considered as categorical variables; and ii) two growth rates (changes of weight-for-age z-scores from birth to 3 months and from 3 months to 12 months) which were treated as continuous variables.There is a growing interest in seeking specific factors to recognize trends in oral health over the lifespan of individuals and to determine whether such factors can alter subsequent oral health events through life-course studies. The investigation of health status during the life-course helps to understand and evaluate the effects of genetic and environmental factors occurred at different ages of life on the occurrence of several health related events . It is uTo the best of our knowledge, this is the first study to investigate the relationship between infant growth and subsequent emergence of permanent tooth at 12 years of age considering growth trajectories and growth rates in a sub-sample from a Chinese birth cohort. The study comprised of 485 students from a birth cohort with an approximately equal distribution of gender (241 boys and 244 girls). It was our observation that children of a higher birth weight had a significant higher chance of having complete emergence of permanent teeth in the permanent dentition at their 12 years of age; and a slow growth rate from birth to 3 months of age had a significant higher chance of having complete emergence of permanent teeth in permanent dentition at their 12 years of age. The results should be treated with caution because this article described statistical associations between birth weight development and tooth emergence in a cross-sectional sample. Using cross-sectional design with a one-time measurement on a phenomenon like growth and tooth emergence could not yield a cause-effect conclusion. However, it provided hints for further studies, in terms of studies with multiple age groups, longitudinal studies, as well as experimental studies. Moreover, the dichotomous analysis of tooth emergence status limited detailed exploration of individual tooth eruption though it is methodologically very difficult to collect data of the exact emergence time.The finding of association between higher birth weight and complete emergence of permanent teeth appears to be similar to reports by other investigators that subjects with low birth weight and/or preterm birth had a higher chance of delayed tooth eruption compared to those with the normal birth weight and/or full term birth –30. It wInterestingly it was found in this study that slow infant growth in the first three months of life was related to the complete emergence of permanent teeth at 12 years of age. The results cannot be fully explained by current knowledge because no study has sought to investigate the possible relationship between infant growth and emergence of permanent teeth in 12-year-old children; this also precludes the possibility of comparing our findings with other published data. However, other investigations in Medicine showed that rapid infant growth are related to certain health problems , 32. AccWhen the association of birth weight and growth rates during the first three months of life on permanent tooth emergence were further investigated for each tooth type Table , signifiThe lack of an association between growth trajectories and permanent tooth emergence in secondary school children, found in this study, may have been because the sub-sample was drawn from a randomized stratified sampling procedure. Only term births (gestation ≥37 weeks) were included to generate the growth trajectories in the study population because different trajectories have been found for term births and preterm births . This isFurther investigations, especially longitudinal studies to monitor growth changes such as infant’s growth in length, and exact timing of individual tooth emergence, as well as experimental studies on the molecular mechanisms triggering dental development are required to expand our knowledge in tooth eruption/emergence.In summary, the results of the logistic regression analyses indicated that infants in a Chinese birth cohort with a heavy birth weight and slow growth during the first three months of life were more likely to have complete emergence of permanent teeth at 12 years of age. The first three months of life might be a critical period for the permanent tooth emergence later in life, while slow growth during this period of time might have beneficial effects on tooth emergence. These findings may have clinical significance of predicting dental events later in life in terms of the risk for dental caries due to prolonged exposure in the oral cavity, and the probability of malocclusions due to unfavorable eruption sequence. Other applications include medical/legal issues and forensic investigations.