The serotonin receptor 3E variant is a risk factor for female IBS-D

Abstract

Irritable bowel syndrome (IBS) is a gut-brain disorder of multifactorial origin. Evidence of disturbed serotonergic function in IBS accumulated for the 5-HT₃ receptor family. 5-HT₃Rs are encoded by HTR3 genes and control GI function, and peristalsis and secretion, in particular. Moreover, 5-HT₃R antagonists are beneficial in the treatment of diarrhea predominant IBS (IBS-D). We previously reported on functionally relevant SNPs in HTR3A c.-42C > T (rs1062613), HTR3C p.N163K (rs6766410), and HTR3E c.*76G > A (rs56109847 = rs62625044) being associated with IBS-D, and the HTR3B variant p.Y129S (rs1176744) was also described within the context of IBS. We performed a multi-center study to validate previous results and provide further evidence for the relevance of HTR3 genes in IBS pathogenesis. Therefore, genotype data of 2682 IBS patients and 9650 controls from 14 cohorts (Chile, Germany (2), Greece, Ireland, Spain, Sweden (2), the UK (3), and the USA (3)) were taken into account. Subsequent meta-analysis confirmed HTR3E c.*76G > A (rs56109847 = rs62625044) to be associated with female IBS-D (OR = 1.58; 95% CI (1.18, 2.12)). Complementary expression studies of four GI regions (jejunum, ileum, colon, sigmoid colon) of 66 IBS patients and 42 controls revealed only HTR3E to be robustly expressed. On top, HTR3E transcript levels were significantly reduced in the sigma of IBS patients (p = 0.0187); more specifically, in those diagnosed with IBS-D (p = 0.0145). In conclusion, meta-analysis confirmed rs56109847 = rs62625044 as a risk factor for female IBS-D. Expression analysis revealed reduced HTR3E levels in the sigmoid colon of IBS-D patients, which underlines the relevance of HTR3E in the pathogenesis of IBS-D.

Keywords: Females; IBS-D; Irritable bowel syndrome; Serotonin type 3 receptor.

Fig. 1

Schematic of the HTR3 genes indicating the location and type of the analyzed SNPs. The gray box indicates the coding sequence of a HTR3 gene. The upstream region of the start codon AUG (5′untranslated region [5′UTR]) is indicated by the thin line on the left, whereas the respective 3′ untranslated region resides downstream of the stop codon, illustrated by an asterisk on the right. CDS, coding sequence (protein coding portion)

Fig. 2

Forest-plots illustrating genotype relative risks of IBS phenotypes were quantified by odds ratios with corresponding 95% confidence intervals (indicated by lower and upper limit) based on a logistic regression model under dominant genetic penetrance for all but HTR3C. We assumed that the identified studies were random samples from a general population, and therefore used a random effects model to summarize odds ratio estimates in the meta-analyses of the respective gene SNP. Confidence intervals for each individual study were indicated by horizontal lines, single ORs by squares that reflected study sizes, and summary estimates by diamonds with horizontal limits at confidence limits and width inversely proportional to the standard error

Fig. 3

qPCR analysis of total RNA isolated from various GI regions including A jejunum, B ileum, C colon, and D, E sigmoid colon. Samples were run in triplicate and data analyzed by the 2-∆∆Ct method with correction for primer efficiency. Two-tailed parametric tests were used as appropriate (unpaired t test, one-way ANOVA followed by Bonferroni correction post-hoc test) using GraphPad Prism 5.0 software