Genome-wide association study meta-analysis identifies three novel loci for circulating anti-Müllerian hormone levels in women.

STUDY QUESTION: Can additional genetic variants for circulating anti-Müllerian hormone (AMH) levels be identified through a genome-wide association study (GWAS) meta-analysis including a large sample of premenopausal women? SUMMARY ANSWER: We identified four loci associated with AMH levels at P < 5 × 10-8: the previously reported MCM8 locus and three novel signals in or near AMH, TEX41 and CDCA7. WHAT IS KNOWN ALREADY: AMH is expressed by antral stage ovarian follicles in women, and variation in age-specific circulating AMH levels has been associated with disease outcomes. However, the physiological mechanisms underlying these AMH-disease associations are largely unknown. STUDY DESIGN, SIZE, DURATION: We performed a GWAS meta-analysis in which we combined summary statistics of a previous AMH GWAS with GWAS data from 3705 additional women from three different cohorts. PARTICIPANTS/MATERIALS, SETTING, METHODS: In total, we included data from 7049 premenopausal female participants of European ancestry. The median age of study participants ranged from 15.3 to 48 years across cohorts. Circulating AMH levels were measured in either serum or plasma samples using different ELISA assays. Study-specific analyses were adjusted for age at blood collection and population stratification, and summary statistics were meta-analysed using a standard error-weighted approach. Subsequently, we functionally annotated GWAS variants that reached genome-wide significance (P < 5 × 10-8). We also performed a gene-based GWAS, pathway analysis and linkage disequilibrium score regression and Mendelian randomization (MR) analyses. MAIN RESULTS AND THE ROLE OF CHANCE: We identified four loci associated with AMH levels at P < 5 × 10-8: the previously reported MCM8 locus and three novel signals in or near AMH, TEX41 and CDCA7. The strongest signal was a missense variant in the AMH gene (rs10417628). Most prioritized genes at the other three identified loci were involved in cell cycle regulation. Genetic correlation analyses indicated a strong positive correlation among single nucleotide polymorphisms for AMH levels and for age at menopause (rg = 0.82, FDR = 0.003). Exploratory two-sample MR analyses did not support causal effects of AMH on breast cancer or polycystic ovary syndrome risk, but should be interpreted with caution as they may be underpowered and the validity of genetic instruments could not be extensively explored. LARGE SCALE DATA: The full AMH GWAS summary statistics will made available after publication through the GWAS catalog (https://www.ebi.ac.uk/gwas/). LIMITATIONS, REASONS FOR CAUTION: Whilst this study doubled the sample size of the most recent GWAS, the statistical power is still relatively low. As a result, we may still lack power to identify more genetic variants for AMH and to determine causal effects of AMH on, for example, breast cancer. Also, follow-up studies are needed to investigate whether the signal for the AMH gene is caused by reduced AMH detection by certain assays instead of actual lower circulating AMH levels. WIDER IMPLICATIONS OF THE FINDINGS: Genes mapped to the MCM8, TEX41 and CDCA7 loci are involved in the cell cycle and processes such as DNA replication and apoptosis. The mechanism underlying their associations with AMH may affect the size of the ovarian follicle pool. Altogether, our results provide more insight into the biology of AMH and, accordingly, the biological processes involved in ovarian ageing. STUDY FUNDING/COMPETING INTEREST(S): Nurses' Health Study and Nurses' Health Study II were supported by research grants from the National Institutes of Health (CA172726, CA186107, CA50385, CA87969, CA49449, CA67262, CA178949). The UK Medical Research Council and Wellcome (217065/Z/19/Z) and the University of Bristol provide core support for ALSPAC. This publication is the work of the listed authors, who will serve as guarantors for the contents of this article. A comprehensive list of grants funding is available on the ALSPAC website (http://www.bristol.ac.uk/alspac/external/documents/grant-acknowledgements.pdf). Funding for the collection of genotype and phenotype data used here was provided by the British Heart Foundation (SP/07/008/24066), Wellcome (WT092830M and WT08806) and UK Medical Research Council (G1001357). M.C.B., A.L.G.S. and D.A.L. work in a unit that is funded by the University of Bristol and UK Medical Research Council (MC_UU_00011/6). M.C.B.'s contribution to this work was funded by a UK Medical Research Council Skills Development Fellowship (MR/P014054/1) and D.A.L. is a National Institute of Health Research Senior Investigator (NF-0616-10102). A.L.G.S. was supported by the study of Dynamic longitudinal exposome trajectories in cardiovascular and metabolic non-communicable diseases (H2020-SC1-2019-Single-Stage-RTD, project ID 874739). The Doetinchem Cohort Study was financially supported by the Ministry of Health, Welfare and Sports of the Netherlands. The funder had no role in study design, data collection and analysis, decision to publish or preparation of the manuscript. Ansh Labs performed the AMH measurements for the Doetinchem Cohort Study free of charge. Ansh Labs was not involved in the data analysis, interpretation or reporting, nor was it financially involved in any aspect of the study. R.M.G.V. was funded by the Honours Track of MSc Epidemiology, University Medical Center Utrecht with a grant from the Netherlands Organization for Scientific Research (NWO) (022.005.021). The Study of Women's Health Across the Nation (SWAN) has grant support from the National Institutes of Health (NIH), DHHS, through the National Institute on Aging (NIA), the National Institute of Nursing Research (NINR) and the NIH Office of Research on Women's Health (ORWH) (U01NR004061; U01AG012505, U01AG012535, U01AG012531, U01AG012539, U01AG012546, U01AG012553, U01AG012554, U01AG012495). The SWAN Genomic Analyses and SWAN Legacy have grant support from the NIA (U01AG017719). The Generations Study was funded by Breast Cancer Now and the Institute of Cancer Research (ICR). The ICR acknowledges NHS funding to the NIHR Biomedical Research Centre. The content of this manuscript is solely the responsibility of the authors and does not necessarily represent official views of the funders. The Sister Study was funded by the Intramural Research Program of the National Institutes of Health (NIH), National Institute of Environmental Health Sciences (Z01-ES044005 to D.P.S.); the AMH assays were supported by the Avon Foundation (02-2012-065 to H.B. Nichols and D.P.S.). The breast cancer genome-wide association analyses were supported by the Government of Canada through Genome Canada and the Canadian Institutes of Health Research, the 'Ministère de l'Économie, de la Science et de l'Innovation du Québec' through Genome Québec and grant PSR-SIIRI-701, The National Institutes of Health (U19 CA148065, X01HG007492), Cancer Research UK (C1287/A10118, C1287/A16563, C1287/A10710) and The European Union (HEALTH-F2-2009-223175 and H2020 633784 and 634935). All studies and funders are listed in Michailidou et al. (Nature, 2017). F.J.M.B. has received fees and grant support from Merck Serono and Ferring BV. D.A.L. has received financial support from several national and international government and charitable funders as well as from Medtronic Ltd and Roche Diagnostics for research that is unrelated to this study. N.S. is scientific consultant for Ansh Laboratories. The other authors declare no competing interests.

Development of Folate-Group Impedimetric Biosensor Based on Polypyrrole Nanotubes Decorated with Gold Nanoparticles.

In this study, polypyrrole nanotubes (PPy-NT) and gold nanoparticles (AuNPs) were electrochemically synthesized to form a hybrid material and used as an electroactive layer for the attachment of proteins for the construction of a high-performance biosensor. Besides the enhancement of intrinsic conductivity of the PPy-NT, the AuNPs act as an anchor group for the formation of self-assembly monolayers (SAMs) from the gold-sulfur covalent interaction between gold and Mercaptopropionic acid (MPA). This material was used to evaluate the viability and performance of the platform developed for biosensing, and three different biological approaches were tested: first, the Avidin-HRP/Biotin couple and characterizations were made by using cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS), wherein we detected Biotin in a linear range of 100-900 fmol L-1. The studies continued with folate group biomolecules, using the folate receptor α (FR-α) as a bioreceptor. Tests with anti-FR antibody detection were performed, and the results obtained indicate a linear range of detection from 0.001 to 6.70 pmol L-1. The same FR-α receptor was used for Folic Acid detection, and the results showed a limit of detection of 0.030 nmol L-1 and a limit of quantification of 90 pmol L-1. The results indicate that the proposed biosensor is sensitive and capable of operating in a range of clinical interests.

Remodeling of the collagen fiber architecture due to compaction in small vessels under tissue engineered conditions.

Mechanical loading protocols in tissue engineering (TE) aim to improve the deposition of a properly organized collagen fiber network. In addition to collagen remodeling, these conditioning protocols can result in tissue compaction. Tissue compaction is beneficial to tissue collagen alignment, yet it may lead to a loss of functionality of the TE construct due to changes in geometry after culture. Here, a mathematical model is presented to relate the changes in collagen architecture to the local compaction within a TE small blood vessel, assuming that under static conditions, compaction is the main factor responsible for collagen fiber organization. An existing structurally based model is extended to incorporate volumetric tissue compaction. Subsequently, the model is applied to describe the collagen architecture of TE constructs under either strain based or stress based stimulus functions. Our computations indicate that stress based simulations result in a helical collagen fiber distribution along the vessel wall. The helix pitch angle increases from a circumferential direction in the inner wall, over about 45 deg in the middle vessel layer, to a longitudinal direction in the outer wall. These results are consistent with experimental data from TE small diameter blood vessels. In addition, our results suggest a stress dependent remodeling of the collagen, suggesting that cell traction is responsible for collagen orientation. These findings may be of value to design improved mechanical conditioning protocols to optimize the collagen architecture in engineered tissues.

Fabry disease caused by the GLA p.Phe113Leu (p.F113L) variant: Natural history in males.

BACKGROUND, AIMS AND METHODS: The α-galactosidase gene (GLA) c.337T>C/p.Phe113Leu variant was originally described in patients with late-onset cardiac forms of Fabry disease (FD), who had residual α-galactosidase activity. It has since emerged as the most commonly reported GLA variant in Portuguese subjects diagnosed with FD but is also prevalent in the Italian population, where two boys carrying the GLA Leu113 allele were identified in a large-scale newborn screening program, the variant allele segregating in both cases with the same surrounding haplotype. To further delineate the genotype-phenotype correlations of this GLA variant, we have reviewed the natural history and clinical phenotypes of 11 symptomatic Portuguese males, from 10 unrelated families originating from several different areas in mainland Portugal and Madeira Island, who were diagnosed with FD associated with the GLA Leu113 allele in a diversity of clinical and screening settings. Nine of the patients were the probands of their respective families. To test whether the GLA Leu113 allele inherited by the 10 Portuguese and the two Italian families resulted from independent mutational events, we have additionally performed a haplotype analysis with 5 highly polymorphic, closely linked microsatellite markers surrounding the GLA gene. RESULTS AND CONCLUSIONS: Hemizygosity for the GLA Leu113 variant allele is associated with a late-onset form of FD, invariably presenting with severe cardiac involvement. Clinically relevant cerebrovascular and kidney involvement may also occur in some patients but the pathogenic relationship between the incomplete α-galactosidase deficiency and the risks of stroke and of chronic kidney disease is not straightforward. The observation that the Leu113 allele segregated within the same GLA microsatellite haplotype in both the Portuguese and Italian families suggests its inheritance from a common ancestor.

Genotype-Specific Growth and Proteomic Responses of Maize Toward Salt Stress.

Salt stress in plants triggers complex physiological responses that are genotype specific. Many of these responses are either not yet described or not fully understood or both. In this work, we phenotyped three maize genotypes of the CIMMYT gene bank alongside the reference B73 genotype (NCRPIS - United States) under both control and salt-stressed conditions. We have ranked their growth potential and we observed significant differences in Na+ and Cl- ion accumulation. Genotype CML421 showed the slowest growth, while CML451 had the lowest accumulation of ions in its leaves. The phenotyping defined the right timing for the proteomics analysis, allowing us to compare the contrasting genotypes. In general 1,747 proteins were identified, of which 209 were significantly more abundant in response to salt stress. The five most significantly enriched annotations that positively correlated with stress were oxidation reduction, catabolic process, response to chemical stimulus, translational elongation and response to water. We observed a higher abundance of proteins involved in reactions to oxidative stress, dehydration, respiration, and translation. The five most significantly enriched annotations negatively correlated with stress were nucleosome organization, chromatin assembly, protein-DNA complex assembly, DNA packaging and nucleosome assembly. The genotypic analysis revealed 52 proteins that were correlated to the slow-growing genotype CML421. Their annotations point toward cellular dehydration and oxidative stress. Three root proteins correlated to the CML451 genotype were annotated to protein synthesis and ion compartmentalization. In conclusion, our results highlight the importance of the anti-oxidative system for acclimatization to salt stress and identify potential genotypic marker proteins involved in salt-stress responses.

Environmental Strategies of Affect Regulation and Their Associations With Subjective Well-Being.

Environmental strategies of affect regulation refer to the use of natural and urban socio-physical settings in the service of regulation. We investigated the perceived use and efficacy of environmental strategies for regulation of general affect and sadness, considering them in relation to other affect regulation strategies and to subjective well-being. Participants from Australia, Finland, Germany, Great Britain, Italy, India, the Netherlands, Portugal, and Sweden (N = 507) evaluated the frequency of use and perceived efficacy of affect regulation strategies using a modified version of the Measure of Affect Regulation Styles (MARS). The internet survey also included the Satisfaction with Life Scale (SWLS), emotional well-being items from the RAND 36-Item Health Survey, and a single-item measure of perceived general health. Environmental regulation formed a separate factor of affect regulation in the exploratory structural equation models (ESEM). Although no relations of environmental strategies with emotional well-being were found, both the perceived frequency of use and efficacy of environmental strategies were positively related to perceived health. Moreover, the perceived efficacy of environmental strategies was positively related to life satisfaction in regulating sadness. The results encourage more explicit treatment of environmental strategies in research on affect regulation.

Age of sexual initiation and depression in adolescents: Data from the 1993 Pelotas (Brazil) Birth Cohort.

BACKGROUND: Studies have shown that sexual initiation at earlier ages increases the risk of depressive symptoms in adolescents. However, little is known about its association with major depressive episode (MDE). METHODS: The association between age of sexual initiation and MDE at 18 years was assessed in the 1993 Pelotas Birth Cohort using multiple logistic regression. Sexual initiation characteristics (age and type of partner) were assessed at the 15- and 18-years follow-up. The age of sexual initiation was evaluated in categories (11-14, 15-16, 17+ years). The type of partner was categorized into: boyfriend/ girlfriend, casual partner and other. MDE was assessed using the Mini International Neuropsychiatric Interview (MINI). RESULTS: From the 4027 adolescents assessed, the prevalence of MDE was higher in females (10.1%) than in males (3.4%), and 66.7% of the males and 58.6% of the females reported sexual initiation up to 16 years (p < 0.001). Female adolescents who had sexual initiation <17 years had higher odds of MDE (15-16 years: OR 2.29; 11-14 years: OR 2.23), however no association was found for males. The type of partner in the first sexual intercourse was not associated to depression. LIMITATIONS: Possibility of recall bias on the age of sexual initiation, and low statistical power for some analyses. CONCLUSIONS: A positive association between age of sexual initiation and MDE was observed only in females. More investigation is needed to understand the mechanisms through which age of sexual initiation can affect the risk of depression and whether the association persists in adulthood.

Age-Dependent Changes in Geometry, Tissue Composition and Mechanical Properties of Fetal to Adult Cryopreserved Human Heart Valves.

There is limited information about age-specific structural and functional properties of human heart valves, while this information is key to the development and evaluation of living valve replacements for pediatric and adolescent patients. Here, we present an extended data set of structure-function properties of cryopreserved human pulmonary and aortic heart valves, providing age-specific information for living valve replacements. Tissue composition, morphology, mechanical properties, and maturation of leaflets from 16 pairs of structurally unaffected aortic and pulmonary valves of human donors (fetal-53 years) were analyzed. Interestingly, no major differences were observed between the aortic and pulmonary valves. Valve annulus and leaflet dimensions increase throughout life. The typical three-layered leaflet structure is present before birth, but becomes more distinct with age. After birth, cell numbers decrease rapidly, while remaining cells obtain a quiescent phenotype and reside in the ventricularis and spongiosa. With age and maturation-but more pronounced in aortic valves-the matrix shows an increasing amount of collagen and collagen cross-links and a reduction in glycosaminoglycans. These matrix changes correlate with increasing leaflet stiffness with age. Our data provide a new and comprehensive overview of the changes of structure-function properties of fetal to adult human semilunar heart valves that can be used to evaluate and optimize future therapies, such as tissue engineering of heart valves. Changing hemodynamic conditions with age can explain initial changes in matrix composition and consequent mechanical properties, but cannot explain the ongoing changes in valve dimensions and matrix composition at older age.