Rome III Criteria Capture Higher Irritable Bowel Syndrome SNP-Heritability and Highlight a Novel Genetic Link With Cardiovascular Traits.

BACKGROUND & AIMS: Irritable bowel syndrome (IBS) shows genetic predisposition, and large-scale genome-wide association studies (GWAS) are emerging, based on heterogeneous disease definitions. We investigated the genetic architecture of IBS defined according to gold standard Rome Criteria. METHODS: We conducted GWAS meta-analyses of Rome III IBS and its subtypes in 24,735 IBS cases and 77,149 asymptomatic control subjects from 2 independent European cohorts (UK Biobank and Lifelines). Single-nucleotide polymorphism (SNP)-based heritability (h2SNP) and genetic correlations (rg) with other traits were calculated. IBS risk loci were functionally annotated to identify candidate genes. Sensitivity and conditional analyses were conducted to assess impact of confounders. Polygenic risk scores were computed and tested in independent datasets. RESULTS: Rome III IBS showed significant SNP-heritability (up to 13%) and similar genetic architecture across subtypes, including those with manifestations at the opposite ends of the symptom spectrum (rg = 0.48 between IBS-D and IBS-C). Genetic correlations with other traits highlighted commonalities with family history of heart disease and hypertension, coronary artery disease, and angina pectoris (rg = 0.20-0.45), among others. Four independent GWAS signals (P < 5×10-8) were detected, including 2 novel loci for IBS (rs2035380) and IBS-mixed (rs2048419) that had been previously associated with hypertension and coronary artery disease. Functional annotation of GWAS risk loci revealed genes implicated in circadian rhythm (BMAL1), intestinal barrier (CLDN23), immunomodulation (MFHAS1), and the cyclic adenosine monophosphate pathway (ADCY2). Polygenic risk scores allowed the identification of individuals at increased risk of IBS (odds ratio, 1.34; P = 1.1×10-3). CONCLUSIONS: Rome III Criteria capture higher SNP-heritability than previously estimated for IBS. The identified link between IBS and cardiovascular traits may contribute to the delineation of alternative therapeutic strategies, warranting further investigation.

Genome-wide analysis of 944 133 individuals provides insights into the etiology of haemorrhoidal disease.

OBJECTIVE: Haemorrhoidal disease (HEM) affects a large and silently suffering fraction of the population but its aetiology, including suspected genetic predisposition, is poorly understood. We report the first genome-wide association study (GWAS) meta-analysis to identify genetic risk factors for HEM to date. DESIGN: We conducted a GWAS meta-analysis of 218 920 patients with HEM and 725 213 controls of European ancestry. Using GWAS summary statistics, we performed multiple genetic correlation analyses between HEM and other traits as well as calculated HEM polygenic risk scores (PRS) and evaluated their translational potential in independent datasets. Using functional annotation of GWAS results, we identified HEM candidate genes, which differential expression and coexpression in HEM tissues were evaluated employing RNA-seq analyses. The localisation of expressed proteins at selected loci was investigated by immunohistochemistry. RESULTS: We demonstrate modest heritability and genetic correlation of HEM with several other diseases from the GI, neuroaffective and cardiovascular domains. HEM PRS validated in 180 435 individuals from independent datasets allowed the identification of those at risk and correlated with younger age of onset and recurrent surgery. We identified 102 independent HEM risk loci harbouring genes whose expression is enriched in blood vessels and GI tissues, and in pathways associated with smooth muscles, epithelial and endothelial development and morphogenesis. Network transcriptomic analyses highlighted HEM gene coexpression modules that are relevant to the development and integrity of the musculoskeletal and epidermal systems, and the organisation of the extracellular matrix. CONCLUSION: HEM has a genetic component that predisposes to smooth muscle, epithelial and connective tissue dysfunction.

Longitudinal Multi-omics Reveals Subset-Specific Mechanisms Underlying Irritable Bowel Syndrome.

The gut microbiome has been implicated in multiple human chronic gastrointestinal (GI) disorders. Determining its mechanistic role in disease has been difficult due to apparent disconnects between animal and human studies and lack of an integrated multi-omics view of disease-specific physiological changes. We integrated longitudinal multi-omics data from the gut microbiome, metabolome, host epigenome, and transcriptome in the context of irritable bowel syndrome (IBS) host physiology. We identified IBS subtype-specific and symptom-related variation in microbial composition and function. A subset of identified changes in microbial metabolites correspond to host physiological mechanisms that are relevant to IBS. By integrating multiple data layers, we identified purine metabolism as a novel host-microbial metabolic pathway in IBS with translational potential. Our study highlights the importance of longitudinal sampling and integrating complementary multi-omics data to identify functional mechanisms that can serve as therapeutic targets in a comprehensive treatment strategy for chronic GI diseases. VIDEO ABSTRACT.

Direct repression of anoctamin 1 (ANO1) gene transcription by Gli proteins.

The Ca2+-activated Cl- channel, anoctamin 1 (Ano1, also known as transmembrane protein 16A) contributes to intestinal pacemaking, fluid secretion, cellular excitability, and tissue development. The human ANO1 promoter contains binding sites for the glioma-associated oncogene (Gli) proteins. We investigated regulation of ANO1 transcription by Gli. ANO1 promoter activity was determined using a luciferase reporter system. Binding and functional effects of Glis on ANO1 transcription and expression were demonstrated by chromatin immunoprecipitation, small interfering RNA knockdown, PCR, immunolabeling, and recordings of Ca2+-activated Cl- currents in human embryonic kidney 293 (HEK293) cells. Results from previous genome-wide association studies were used to test ANO1 promoter polymorphisms for association with disease. Gli1 and Gli2 bound to the promoter and repressed ANO1 transcription. Repression depended on Gli binding to a site close to the ANO1 transcriptional start site. Mutation of this site prevented Gli binding and transcriptional repression. Knockdown of Gli expression and inhibition of Gli activity increased expression of ANO1 RNA and Ca2+-activated Cl- currents in HEK293 cells. A single-nucleotide polymorphism prevented Gli binding and showed association with irritable bowel syndrome. We conclude that Gli1 and Gli2 repress ANO1 by a novel mechanism that is independent of Gli cleavage and that has a role in gastrointestinal function.-Mazzone, A., Gibbons, S. J., Eisenman, S. T., Strege, P. R., Zheng, T., D'Amato, M., Ordog, T., Fernandez-Zapico, M. E., Farrugia, G. Direct repression of anoctamin 1 (ANO1) gene transcription by Gli proteins.

A preliminary study of plasma cyclase-associated protein 2 as a novel biomarker for early stage and alpha-fetoprotein negative hepatocellular carcinoma patients.

BACKGROUND AND OBJECTIVE: Cyclase-associated protein 2 (CAP2) has recently been suggested to be a candidate biomarker for hepatocellular carcinoma (HCC). We aim to investigate the application of CAP2 as a novel biomarker for HCC patients especially for those at early stage and are AFP-negative. METHODS: The CAP2 and AFP plasma levels were analyzed by enzyme-linked-immunosorbent assay in 86 HCC, 59 cirrhotic patients, and 30 normal individuals. Their correlation with HCC tumor behavior, disease stages, diagnostic sensitivity, specificity and accuracy were analyzed. RESULTS: The results showed that both CAP2 and AFP plasma levels in HCC patients were significantly elevated when compared to cirrhosis and controls. CAP2 levels correlate well with HCC patient's histological grade, clinical stage and tumor size, but not with patient's age, gender, hepatitis B virus infection status and plasma AFP level. CAP2 had better sensitivity as compared to AFP (82.6% vs 59.3%) for general HCC, and early stage of HCC patients (78.6% vs 40.4%). In addition, CAP2 is able to complement AFP to predict 82.9% of HCC in AFP-negative patients. CONCLUSION: We suggest that CAP2 is a novel biomarker for HCC patient, this may be especially useful for detection of early stage HCC and when plasma AFP level is negative.

Plasma minichromosome maintenance complex component 6 is a novel biomarker for hepatocellular carcinoma patients.

AIM: This study aimed to investigate the presence of plasma minichromosome maintenance complex component 6 (MCM6) mRNA and protein levels in hepatocellular carcinoma (HCC) patients and evaluate their diagnostic value for HCC. METHODS: Blood samples were collected from 61 HCC and 29 cirrhotic patients, and 30 healthy individuals. Circulating RNA was extracted from plasma of all samples. The mRNA for MCM6 were amplified and quantified by real-time polymerase chain reaction. Plasma MCM6 and α-fetoprotein (AFP) protein levels were measured by enzyme-linked immunosorbent assay. RESULTS: In HCC patients, MCM6 mRNA and protein levels were significantly increased over the cirrhotic and healthy controls. The levels of MCM6 mRNA and protein in the plasma of HCC patients correlated to vascular invasion (P < 0.01). Higher MCM6 protein levels also correlated with tumor stage progression and lymph node metastasis. The MCM6 protein has sensitivity of 67.2% and specificity of 89.8% in differentiating total HCC from non-HCC individuals. In the AFP negative HCC group, MCM6 mRNA and protein could both detect 76.9% of HCC patients; combining the two of them increased the detection rate to 84.6%. In small HCC patients, MCM6 mRNA and protein could detect 64.3% and 71.4% of patients, respectively; combining AFP, MCM6 mRNA and MCM6 protein could detect 85.7% of small HCC patients. CONCLUSION: Our results suggest that MCM6 mRNA and protein levels in plasma can be promising independent biomarkers for HCC, especially in AFP negative and small HCC patients.

Fluconazole inhibits hERG K(+) channel by direct block and disruption of protein trafficking.

Fluconazole, a commonly used azole antifungal drug, can induce QT prolongation, which may lead to Torsades de Pointes and sudden death. To investigate the arrhythmogenic side effects of fluconazole, we studied the effect of fluconazole on human ether-a-go-go-related gene (hERG) K(+) channels (wild type, Y652A and F656C) expressed in human embryonic kidney (HEK293) cells using a whole-cell patch clamp technique, Western blot analysis and confocal microscopy. Fluconazole inhibited wild type hERG currents in a concentration-dependent manner, with a half-maximum block concentration (IC(50)) of 48.2±9.4µM. Fluconazole did not change other channel kinetics (activation and steady-state inactivation) of hERG channel. Mutations in drug- binding sites (Y652A or F656C) of the hERG channel significantly attenuated the hERG current blockade by fluconazole. In addition, fluconazole inhibited the trafficking of hERG protein by Western blot analysis and confocal microscopy, respectively. These findings indicate that fluconazole may cause acquired long QT syndrome (LQTS) via a direct inhibition of hERG current and by disrupting hERG protein trafficking, and the mutations Y652 and F656 may be obligatory determinants in inhibition of hERG current for fluconazole.