Identification of arginine-vasopressin receptor 1a (Avpr1a/AVPR1A) as a novel candidate gene for chronic visceral pain sheds light on the potential role of enteric neurons in the development of visceral hypersensitivity.

Chronic abdominal pain in the absence of ongoing disease is the hallmark of disorders of gut-brain interaction (DGBIs), including irritable bowel syndrome (IBS). While the etiology of DGBIs remains poorly understood, there is evidence that both genetic and environmental factors play a role. In this study, we report the identification and validation of Avpr1a as a novel candidate gene for visceral hypersensitivity (VH), a primary peripheral mechanism underlying abdominal pain in DGBI/IBS. Comparing two C57BL/6 (BL/6) substrains (C57BL/6NTac and C57BL/6J) revealed differential susceptibility to the development of chronic VH following intrarectal zymosan (ZYM) instillation, a validated preclinical model for post-inflammatory IBS. Using whole genome sequencing, we identified a SNP differentiating the two strains in the 5' intergenic region upstream of Avpr1a, encoding the protein arginine-vasopressin receptor 1A (AVPR1A). We used behavioral, histological, and molecular approaches to identify distal colon-specific gene expression and neuronal hyperresponsiveness covarying with Avpr1a genotype and VH susceptibility. While the two BL/6 substrains did not differ across other gastrointestinal (GI) phenotypes (e.g., fecal water retention), VH-susceptible BL/6NTac mice had higher colonic Avpr1a mRNA and protein expression. These results parallel findings that patients' colonic Avpr1a mRNA expression corresponded to higher pain ratings. Moreover, neurons of the enteric nervous system were hyperresponsive to the AVPR1A agonist AVP, suggesting a role for enteric neurons in the pathology underlying VH. Taken together, these findings implicate differential regulation of Avpr1a as a novel mechanism of VH-susceptibility as well as a potential therapeutic target specific to VH. PERSPECTIVE: This article presents evidence of Avpr1a as a novel candidate gene for visceral hypersensitivity in a mouse model of irritable bowel syndrome. Avpr1a genotype and/or tissue-specific expression represents a potential biomarker for chronic abdominal pain susceptibility.

Identification of arginine-vasopressin receptor 1a (Avpr1a/AVPR1A) as a novel candidate gene for chronic visceral pain.

Chronic abdominal pain in the absence of ongoing disease is the hallmark of disorders of gut-brain interaction (DGBIs), including irritable bowel syndrome (IBS). While the etiology of DGBIs remains poorly understood, there is evidence that both genetic and environmental factors play a role. In this study, we report the identification and validation of Avpr1a as a novel candidate gene for visceral hypersensitivity (VH), a primary peripheral mechanism underlying abdominal pain in DGBI/IBS. Comparing two C57BL/6 (BL/6) substrains (C57BL/6NTac and C57BL/6J) revealed differential susceptibility to the development of chronic VH following intrarectal zymosan (ZYM) instillation, a validated preclinical model for post-inflammatory IBS. Using whole genome sequencing, we identified a SNP differentiating the two strains in the 5' intergenic region upstream of Avpr1a, encoding the protein arginine-vasopressin receptor 1A (AVPR1A). We used behavioral, histological, and molecular approaches to identify distal colon-specific gene expression differences and neuronal hyperresponsiveness covarying with Avpr1a genotype and VH susceptibility. While the two BL/6 substrains did not differ across other gastrointestinal (GI) phenotypes (e.g., GI motility), VH-susceptible BL/6NTac mice had higher colonic Avpr1a mRNA and protein expression. Moreover, neurons of the enteric nervous system were hyperresponsive to the AVPR1A agonist AVP, suggesting a role for enteric neurons in the pathology underlying VH. These results parallel our findings that patients' colonic Avpr1a mRNA expression was higher in patients with higher pain ratings. Taken together, these findings implicate differential regulation of Avpr1a as a novel mechanism of VH-susceptibility as well as a potential therapeutic target specific to VH.

Validation of Polygenic Scores for QT Interval in Clinical Populations.

BACKGROUND: Polygenic risk scores (PGS) enable rapid estimation of genome-wide susceptibility for traits, which may be useful in clinical settings, such as prediction of QT interval. In this study, we sought to validate PGS for QT interval in 2 real-world cohorts of European ancestry (EA) and African ancestry (AA). METHODS AND RESULTS: Two thousand nine hundred and fifteen participants of EA and 366 of AA in the MGH CAMP study (Cardiology and Metabolic Patient) were genotyped on a genome-wide array and imputed to the 1000 Genomes reference panel. An additional 820 EA and 57 AA participants in the Partners Biobank were genotyped and used for validation. PGS were created for each individual using effect estimates from association tests with QT interval obtained from prior genome-wide association studies, with variants selected based from multiple significance thresholds in the original study. In regression models, clinical variables explained ≈9% to 10% of total variation in resting QTc in EA individuals and ≈12% to 18% in AA individuals. The PGS significantly increased variation explained at most significance thresholds (P<0.001), with a trend toward increased variation explained at more stringent P value cut points in the CAMP EA cohort (P<0.05). In AA individuals, PGS provided no improvement in variation explained at any significance threshold. CONCLUSIONS: For individuals of European descent, PGS provided a significant increase in variation in QT interval explained compared with a model with only nongenetic factors at nearly every significance level. There was no apparent benefit gained by relaxing the significance threshold from conventional genome-wide significance (P<5×10-8).

Pathogenic Anti-Müllerian Hormone Variants in Polycystic Ovary Syndrome.

Context: Polycystic ovary syndrome (PCOS), a common endocrine condition, is the leading cause of anovulatory infertility. Objective: Given that common disease-susceptibility variants account for only a small percentage of the estimated PCOS heritability, we tested the hypothesis that rare variants contribute to this deficit in heritability. Design, Setting, and Participants: Unbiased whole-genome sequencing (WGS) of 80 patients with PCOS and 24 reproductively normal control subjects identified potentially deleterious variants in AMH, the gene encoding anti-Müllerian hormone (AMH). Targeted sequencing of AMH of 643 patients with PCOS and 153 control patients was used to replicate WGS findings. Main Outcome Measures: Dual luciferase reporter assays measured the impact of the variants on downstream AMH signaling. Results: We found 24 rare (minor allele frequency < 0.01) AMH variants in patients with PCOS and control subjects; 18 variants were specific to women with PCOS. Seventeen of 18 (94%) PCOS-specific variants had significantly reduced AMH signaling, whereas none of 6 variants observed in control subjects showed significant defects in signaling. Thus, we identified rare AMH coding variants that reduced AMH-mediated signaling in a subset of patients with PCOS. Conclusion: To our knowledge, this study is the first to identify rare genetic variants associated with a common PCOS phenotype. Our findings suggest decreased AMH signaling as a mechanism for the pathogenesis of PCOS. AMH decreases androgen biosynthesis by inhibiting CYP17 activity; a potential mechanism of action for AMH variants in PCOS, therefore, is to increase androgen biosynthesis due to decreased AMH-mediated inhibition of CYP17 activity.

Common Genetic Variant Risk Score Is Associated With Drug-Induced QT Prolongation and Torsade de Pointes Risk: A Pilot Study.

BACKGROUND: Drug-induced QT interval prolongation, a risk factor for life-threatening ventricular arrhythmias, is a potential side effect of many marketed and withdrawn medications. The contribution of common genetic variants previously associated with baseline QT interval to drug-induced QT prolongation and arrhythmias is not known. METHODS: We tested the hypothesis that a weighted combination of common genetic variants contributing to QT interval at baseline, identified through genome-wide association studies, can predict individual response to multiple QT-prolonging drugs. Genetic analysis of 22 subjects was performed in a secondary analysis of a randomized, double-blind, placebo-controlled, crossover trial of 3 QT-prolonging drugs with 15 time-matched QT and plasma drug concentration measurements. Subjects received single doses of dofetilide, quinidine, ranolazine, and placebo. The outcome was the correlation between a genetic QT score comprising 61 common genetic variants and the slope of an individual subject's drug-induced increase in heart rate-corrected QT (QTc) versus drug concentration. RESULTS: The genetic QT score was correlated with drug-induced QTc prolongation. Among white subjects, genetic QT score explained 30% of the variability in response to dofetilide (r=0.55; 95% confidence interval, 0.09-0.81; P=0.02), 23% in response to quinidine (r=0.48; 95% confidence interval, -0.03 to 0.79; P=0.06), and 27% in response to ranolazine (r=0.52; 95% confidence interval, 0.05-0.80; P=0.03). Furthermore, the genetic QT score was a significant predictor of drug-induced torsade de pointes in an independent sample of 216 cases compared with 771 controls (r2=12%, P=1×10-7). CONCLUSIONS: We demonstrate that a genetic QT score comprising 61 common genetic variants explains a significant proportion of the variability in drug-induced QT prolongation and is a significant predictor of drug-induced torsade de pointes. These findings highlight an opportunity for recent genetic discoveries to improve individualized risk-benefit assessment for pharmacological therapies. Replication of these findings in larger samples is needed to more precisely estimate variance explained and to establish the individual variants that drive these effects. CLINICAL TRIAL REGISTRATION: URL: http://www.clinicaltrials.gov. Unique identifier: NCT01873950.

Meta-analysis identifies common and rare variants influencing blood pressure and overlapping with metabolic trait loci.

Meta-analyses of association results for blood pressure using exome-centric single-variant and gene-based tests identified 31 new loci in a discovery stage among 146,562 individuals, with follow-up and meta-analysis in 180,726 additional individuals (total n = 327,288). These blood pressure-associated loci are enriched for known variants for cardiometabolic traits. Associations were also observed for the aggregation of rare and low-frequency missense variants in three genes, NPR1, DBH, and PTPMT1. In addition, blood pressure associations at 39 previously reported loci were confirmed. The identified variants implicate biological pathways related to cardiometabolic traits, vascular function, and development. Several new variants are inferred to have roles in transcription or as hubs in protein-protein interaction networks. Genetic risk scores constructed from the identified variants were strongly associated with coronary disease and myocardial infarction. This large collection of blood pressure-associated loci suggests new therapeutic strategies for hypertension, emphasizing a link with cardiometabolic risk.

Expression Quantitative Trait Locus Mapping Studies in Mid-secretory Phase Endometrial Cells Identifies HLA-F and TAP2 as Fecundability-Associated Genes.

Fertility traits in humans are heritable, however, little is known about the genes that influence reproductive outcomes or the genetic variants that contribute to differences in these traits between individuals, particularly women. To address this gap in knowledge, we performed an unbiased genome-wide expression quantitative trait locus (eQTL) mapping study to identify common regulatory (expression) single nucleotide polymorphisms (eSNPs) in mid-secretory endometrium. We identified 423 cis-eQTLs for 132 genes that were significant at a false discovery rate (FDR) of 1%. After pruning for strong LD (r2 >0.95), we tested for associations between eSNPs and fecundability (the ability to get pregnant), measured as the length of the interval to pregnancy, in 117 women. Two eSNPs were associated with fecundability at a FDR of 5%; both were in the HLA region and were eQTLs for the TAP2 gene (P = 1.3x10-4) and the HLA-F gene (P = 4.0x10-4), respectively. The effects of these SNPs on fecundability were replicated in an independent sample. The two eSNPs reside within or near regulatory elements in decidualized human endometrial stromal cells. Our study integrating eQTL mapping in a primary tissue with association studies of a related phenotype revealed novel genes and associated alleles with independent effects on fecundability, and identified a central role for two HLA region genes in human implantation success.

Evolutionary forward genomics reveals novel insights into the genes and pathways dysregulated in recurrent early pregnancy loss.

STUDY QUESTION: Are the genes that gained novel expression in the endometria of Eutherian (placental) mammals more likely to be dysregulated in patients with endometrial-associated recurrent early pregnancy loss (REPL)? SUMMARY ANSWER: There was a significant enrichment of genes dysregulated in REPL patients among the Eutherian-specific endometrial genes. WHAT IS KNOWN ALREADY: Pregnancy loss is the most common complication of human pregnancy. REPL has multiple etiologies, including dysregulation of endometrial function, leading to 'suboptimal' implantation. Although the implantation process is tightly regulated in Eutherian (placental) mammals, the molecular factors contributing to dysregulated endometrial gene expression patterns in women with REPL are largely unknown. STUDY DESIGN, SIZE, DURATION: Endometrial biopsies were obtained from 32 REPL patients during the mid-luteal phase, and evaluated for glandular development arrest based on elevated nuclear cyclin E levels in gland cells, and for out-of-phase endometrial development based on histology. Gene expression levels were measured using Illumina Human HT-12v4 BeadChip arrays. PARTICIPANTS/MATERIALS, SETTING, METHODS: Differentially expressed genes were identified between patients with (i) out-of-phase (n = 10) versus normal (n = 22) histological dating and (ii) abnormally elevated (n = 9) versus normal (n = 23) cyclin E levels in the nuclei of endometrial glands, using a likelihood ratio test. Enrichment of dysregulated genes in REPL endometria among Eutherian-specific genes was tested by permutation. Gene ontology and pathway enrichment analyses were carried out for the dysregulated genes. MAIN RESULTS AND THE ROLE OF CHANCE: Fifty-eight and eighty-one genes were identified as differentially expressed at P < 0.001 in women with out-of-phase histological dating and abnormally elevated glandular cyclin E levels, respectively. Genes that were recruited into endometrial expression during the evolution of pregnancy in Eutherian mammals were significantly enriched for dysregulated genes (P = 0.002 for histology, P = 0.021 for cyclin E), as well as for genes involved in immune response and signaling pathways with essential roles in implantation and endometrial biology. LIMITATIONS, REASONS FOR CAUTION: Small sample size limits the statistical power to detect dysregulated genes, and the lack of non-REPL control women does not allow us to test for the contribution of these genes to overall risk of REPL. WIDER IMPLICATIONS OF THE FINDINGS: Enrichment of functional gene categories, as well as genes gained expression in the Eutherian endometria, help to identify molecular etiologies that contribute to normal functioning of the endometrium. These pathways are also strong candidates for successful pregnancy outcomes. Using the evolutionary history of mammalian gene expression in the endometrial tissue may be a promising approach to discover genes involved in female reproductive disorders. STUDY FUNDING/COMPETING INTERESTS: This work is supported by National Institutes of Health (NIH) grant R01 HD21244 to C.O. Authors declare no competing interests.

Variants in DPF3 and DSCAML1 are associated with sperm morphology.

PURPOSE: Abnormalities in semen parameters are often associated with reduced fertility in males, and may, in part, be attributed to genetic variation. Aim of this study is to determine if genetic variants that were previously shown to be predictors of family size and birth rate in healthy men are also associated with sperm morphology in men recruited from an infertility laboratory. METHODS: Genetic associations with sperm morphology phenotypes in 126 ethnically diverse men from Chicago at 41 independent loci, previously shown to be predictors of family size and birth rate in healthy men, were tested. RESULTS: Two intronic SNPs, rs680730 (in DSCAML1) and rs10129954 (in DPF3), were associated with the percent of normal sperm morphology in Chicago men (P = 0.017 and 0.023, respectively). Furthermore, both loci were associated with increased occurrence of sperm head defects. CONCLUSIONS: SNPs in two genes, both of which have roles in nervous system development, were associated with poor sperm morphology. These results may be helpful in identification of other novel genes and biological pathways whose proper functioning is crucial for sperm production and male reproductive processes.

Genetics of the polycystic ovary syndrome.

Polycystic ovary syndrome (PCOS) is a highly complex endocrine disorder, characterized by hyperandrogenemia, menstrual irregularities and polycystic ovaries. A strong genetic component to the etiology of PCOS is evident. However, due to the genetic and phenotypic heterogeneity of PCOS and the lack of insufficiently large cohorts, studies to identify specific contributing genes to date have yielded only few conclusive results. In this review we discuss the current status of the genetic analysis of PCOS including the results of numerous association studies with candidate genes involved in TGF-ß and insulin signaling, type 2 diabetes mellitus and obesity susceptibility. Furthermore, we address current challenges in genetic studies of PCOS, and the promise of new approaches, including genome-wide association studies and next-generation sequencing.

Genome-wide association study identifies candidate genes for male fertility traits in humans.

Despite the fact that hundreds of genes are known to affect fertility in animal models, relatively little is known about genes that influence natural fertility in humans. To broadly survey genes contributing to variation in male fertility, we conducted a genome-wide association study (GWAS) of two fertility traits (family size and birth rate) in 269 married men who are members of a founder population of European descent that proscribes contraception and has large family sizes. Associations between ∼250,000 autosomal SNPs and the fertility traits were examined. A total of 41 SNPs with p ≤ 1 × 10(-4) for either trait were taken forward to a validation study of 123 ethnically diverse men from Chicago who had previously undergone semen analyses. Nine (22%) of the SNPs associated with reduced fertility in the GWAS were also associated with one or more of the ten measures of reduced sperm quantity and/or function, yielding 27 associations with p values < 0.05 and seven with p values < 0.01 in the validation study. On the basis of 5,000 permutations of our data, the probabilities of observing this many or more small p values were 0.0014 and 5.6 × 10(-4), respectively. Among the nine associated loci, outstanding candidates for male fertility genes include USP8, an essential deubiquitinating enzyme that has a role in acrosome assembly; UBD and EPSTI1, which have potential roles in innate immunity; and LRRC32, which encodes a latent transforming growth factor ß (TGF-ß) receptor on regulatory T cells. We suggest that mutations in these genes that are more severe may account for some of the unexplained infertility (or subfertility) in the general population.

The CFTR Met 470 allele is associated with lower birth rates in fertile men from a population isolate.

Although little is known about the role of the cystic fibrosis transmembrane regulator (CFTR) gene in reproductive physiology, numerous variants in this gene have been implicated in etiology of male infertility due to congenital bilateral absence of the vas deferens (CBAVD). Here, we studied the fertility effects of three CBAVD-associated CFTR polymorphisms, the (TG)m and polyT repeat polymorphisms in intron 8 and Met470Val in exon 10, in healthy men of European descent. Homozygosity for the Met470 allele was associated with lower birth rates, defined as the number of births per year of marriage (P = 0.0029). The Met470Val locus explained 4.36% of the phenotypic variance in birth rate, and men homozygous for the Met470 allele had 0.56 fewer children on average compared to Val470 carrier men. The derived Val470 allele occurs at high frequencies in non-African populations (allele frequency = 0.51 in HapMap CEU), whereas it is very rare in African population (Fst = 0.43 between HapMap CEU and YRI). In addition, haplotypes bearing Val470 show a lack of genetic diversity and are thus longer than haplotypes bearing Met470 (measured by an integrated haplotype score [iHS] of -1.93 in HapMap CEU). The fraction of SNPs in the HapMap Phase2 data set with more extreme Fst and iHS measures is 0.003, consistent with a selective sweep outside of Africa. The fertility advantage conferred by Val470 relative to Met470 may provide a selective mechanism for these population genetic observations.

Colloquium papers: Heritability of reproductive fitness traits in a human population.

The genetic basis of fitness traits has been studied widely in animals, yet the contribution of genetic variation to these traits in humans is controversial. In particular, it is difficult to disentangle genetic versus environmental effects on fertility, because of within-family correlations of sociocultural, economic, and other nongenetic factors that influence family sizes. In this study, we investigated the genetic architecture of reproductive fitness traits in a fertile human population whose communal lifestyle assures uniform and equal access to resources. Our study revealed significant heritabilities for reproductive traits in both men and women, after accounting for common household effects shared among siblings and demographic changes in reproductive practices. Furthermore, our results indicate that both autosomal and X-linked additive and dominance variances contribute to these traits. We therefore propose that reproductive traits should be amenable to genetic mapping studies, and the results we present here will facilitate the search for the novel genes influencing natural fertility in humans.

Variation in ITGB3 is associated with whole-blood serotonin level and autism susceptibility.

Autism is a pervasive developmental disorder affecting more males than females. Heritability estimates for autism can rise above 90%, and genes influencing the serotonin system are strong candidates for autism susceptibility genes, as drugs selectively acting on the serotonin system are some of the most effective treatments for maladaptive behaviors seen in autism. ITGB3 was recently identified as a male quantitative trait locus (QTL) for whole-blood serotonin levels in the Hutterites (P = 0.0003). Here, we demonstrate associations between variation in ITGB3 and serotonin levels in two outbred samples (P = 0.010 and 0.015). Lastly, we show that a coding variant of ITGB3 is associated with autism susceptibility in a large multiplex sample (P = 0.00082), and that this variation has different effects in males and females (P = 0.0018).