High-Dimensional Clustering of 4000 Irritable Bowel Syndrome Patients Reveals Seven Distinct Disease Subsets.

BACKGROUND AND AIMS: Irritable bowel syndrome (IBS) is a pain disorder classified by bowel habits, disregarding other factors that may influence the clinical course. The aim of this study was to determine if IBS patients can be clustered based on clinical, dietary, lifestyle, and psychosocial factors. METHODS: Between 2013 and 2020, the Mayo Clinic Biobank surveyed and received 40,291 responses to a questionnaire incorporating Rome III criteria. Factors associated with IBS were determined and latent class analysis, a model-based clustering, was performed on IBS cases. RESULTS: We identified 4021 IBS patients (mean 64 years; 75% women) and 12,063 controls. Using 26 variables separating cases from controls, the optimal clustering revealed 7 latent clusters. These were characterized by perceived health impairment (moderate or severe), psychoneurological factors, and bowel dysfunction (diarrhea or constipation predominance). Health impairment clusters demonstrated more pain, with the severe cluster also having more psychiatric comorbidities. The next 3 clusters had unique enrichment of psychiatric, neurological, or both comorbidities. The bowel dysfunction clusters demonstrated less abdominal pain, with diarrhea cluster most likely to report pain improvement with defecation. The constipation cluster had the highest exercise score and consumption of fruits, vegetables, and alcohol. The distribution of clusters remained similar when Rome IV criteria were applied. Physiologic tests were available on a limited subset (6%), and there were no significant differences between clusters. CONCLUSIONS: In this cohort of older IBS patients, 7 distinct clusters were identified demonstrating varying degrees of gastrointestinal symptoms, comorbidities, dietary, and lifestyle factors. Further research is required to assess whether these unique clusters could be used to direct clinical trials and individualize patient management.

Gut microbial ß-glucuronidases regulate host luminal proteases and are depleted in irritable bowel syndrome.

Intestinal proteases mediate digestion and immune signalling, while increased gut proteolytic activity disrupts the intestinal barrier and generates visceral hypersensitivity, which is common in irritable bowel syndrome (IBS). However, the mechanisms controlling protease function are unclear. Here we show that members of the gut microbiota suppress intestinal proteolytic activity through production of unconjugated bilirubin. This occurs via microbial ß-glucuronidase-mediated conversion of bilirubin conjugates. Metagenomic analysis of faecal samples from patients with post-infection IBS (n = 52) revealed an altered gut microbiota composition, in particular a reduction in Alistipes taxa, and high gut proteolytic activity driven by specific host serine proteases compared with controls. Germ-free mice showed 10-fold higher proteolytic activity compared with conventional mice. Colonization with microbiota samples from high proteolytic activity IBS patients failed to suppress proteolytic activity in germ-free mice, but suppression of proteolytic activity was achieved with colonization using microbiota from healthy donors. High proteolytic activity mice had higher intestinal permeability, a higher relative abundance of Bacteroides and a reduction in Alistipes taxa compared with low proteolytic activity mice. High proteolytic activity IBS patients had lower fecal ß-glucuronidase activity and end-products of bilirubin deconjugation. Mice treated with unconjugated bilirubin and ß-glucuronidase-overexpressing E. coli significantly reduced proteolytic activity, while inhibitors of microbial ß-glucuronidases increased proteolytic activity. Together, these data define a disease-relevant mechanism of host-microbial interaction that maintains protease homoeostasis in the gut.

Post-infection Irritable Bowel Syndrome.

Epidemiologic data support that acute gastrointestinal infection is one of the strongest risk factors for development of irritable bowel syndrome (IBS). Risk of post-infection IBS (PI-IBS) seems to be greater with bacterial and protozoal than viral enterocolitis. Younger individuals, women, and those with severe enterocolitis are more likely to develop PI-IBS. Disease mechanisms in animal models and humans involve chronic perturbation of intestinal microbiome, epithelial and neuronal remodeling, and immune activation. These mechanisms can lead to luminal (increased proteolytic activity, altered bile acid composition) and physiologic (increased permeability, transit changes, and visceral hypersensitivity) alterations that can mediate PI-IBS symptoms.

Intestinal barrier dysfunction in irritable bowel syndrome: a systematic review.

BACKGROUND AND AIM: Irritable bowel syndrome (IBS) is a complex and heterogeneous disorder. Sensory, motor and barrier dysfunctions are the key physiological endophenotypes of IBS. Our aim is to review studies evaluating barrier dysfunction in adults and children with IBS, as well as to link those changes with IBS symptomatology and quality of life. METHODS: A comprehensive and systematic review of multiple databases was performed up to March 2020 to identify studies comparing intestinal permeability in IBS patients with healthy controls. Both in vivo and in vitro studies were considered. RESULTS: We identified 66 studies, of which 27 used intestinal probes to quantify barrier function. The prevalence of barrier dysfunction differed between PI-IBS (17-50%), IBS-D (37-62%) and IBS-C (4-25%). At a group level, permeability was increased compared with healthy controls in IBS-D (9/13 studies) and PI-IBS (4/4 studies), but only a minority of IBS-C (2/7 studies) and not in the only IBS-M study. All four studies in children with IBS demonstrated loss of barrier function. A heterogeneous set of tight junction genes were found to be altered in small and large intestines of adults with IBS, but these have not been evaluated in children. Positive associations were identified between barrier dysfunction and bowel disturbances (6/9 studies), abdominal pain (9/13 studies), overall symptom severity (1/6 studies), depression and anxiety (1/1 study) and quality of life (1/4 studies). Fecal slurry or supernatants of IBS patients were found to induce barrier disruption in animal models (5/6 studies). CONCLUSIONS: Barrier dysfunction is present in a significant proportion of adult and all pediatric IBS studies, especially in the IBS-D and PI-IBS subtype. The majority of studies indicated a positive association between loss of barrier function and symptoms such as abdominal pain and changes in the bowel function.

Serine proteases as luminal mediators of intestinal barrier dysfunction and symptom severity in IBS.

OBJECTIVE: The intestinal lumen contains several proteases. Our aim was to determine the role of faecal proteases in mediating barrier dysfunction and symptoms in IBS. DESIGN: 39 patients with IBS and 25 healthy volunteers completed questionnaires, assessments of in vivo permeability, ex vivo colonic barrier function in Ussing chambers, tight junction (TJ) proteins, ultrastructural morphology and 16 s sequencing of faecal microbiota rRNA. A casein-based assay was used to measure proteolytic activity (PA) in faecal supernatants (FSNs). Colonic barrier function was determined in mice (ex-germ free) humanised with microbial communities associated with different human PA states. RESULTS: Patients with IBS had higher faecal PA than healthy volunteers. 8/20 postinfection IBS (PI-IBS) and 3/19 constipation- predominant IBS had high PA (>95th percentile). High-PA patients had more and looser bowel movements, greater symptom severity and higher in vivo and ex vivo colonic permeability. High-PA FSNs increased paracellular permeability, decreased occludin and increased phosphorylated myosin light chain (pMLC) expression. Serine but not cysteine protease inhibitor significantly blocked high-PA FSN effects on barrier. The effects on barrier were diminished by pharmacological or siRNA inhibition of protease activated receptor-2 (PAR-2). Patients with high-PA IBS had lower occludin expression, wider TJs on biopsies and reduced microbial diversity than patients with low PA. Mice humanised with high-PA IBS microbiota had greater in vivo permeability than those with low-PA microbiota. CONCLUSION: A subset of patients with IBS, especially in PI-IBS, has substantially high faecal PA, greater symptoms, impaired barrier and reduced microbial diversity. Commensal microbiota affects luminal PA that can influence host barrier function.

Measurement of novel intestinal secretory and barrier pathways and effects of proteases.

The epithelial lining of the gastrointestinal (GI) tract in conjunction with the enteric nervous system (ENS) plays an important role in mediating solute absorption and secretion. A dysregulated ionic movement across the epithelium can result in GI diseases that manifest as either watery diarrhea or constipation. Hirschsprung disease is an example of an ENS disorder characterized by absence of enteric ganglia in distal gut resulting in obstructive phenotype. Receptor rearranged during transfection (RET) gene variants are the most commonly recognized genetic associations with Hirschsprung disease. In this issue of Neurogastroenterology and Motility, Russell et al demonstrate that RET mediates colonic ion transport through modulation of cholinergic nerves. They go on to show inhibition of RET can attenuate accelerated transit in a rat model. Normalizing secretory and absorptive defects has been an attractive therapeutic strategy. In addition to the intrinsic regulation of secretory processes, luminal mediators like bile acids, short-chain fatty acids, and proteases can affect both secretion and barrier function of the intestinal epithelium. Elevated levels of proteases have been identified in a wide range of GI diseases including irritable bowel syndrome. Proteases are known to cause visceral hypersensitivity and barrier disruption in vitro and in animal models. The goals of this review are to describe fundamental concepts related to intestinal epithelial secretion, the utility of Ussing chambers to measure ionic mechanisms and to discuss examples of novel signaling pathways; namely the RET signaling cascade in secretomotor neurons and effects of luminal proteases on barrier and ionic secretion.