Introduction

Irritable bowel syndrome (IBS) is a common disorder rooted in a disruption of the gut-brain-axis, characterized by abdominal pain and altered bowel habits. Altered gut microbiota is reported in IBS, with microbial metabolites, including bile acids, recognized to be potentially involved in the generation of IBS symptoms.

Aims & Methods

This study aims to clarify the potential link between the gut microbiota composition, fecal bile acid levels, colonic gene expression, and IBS symptoms. In patients with IBS (n = 152), and healthy controls without gastrointestinal symptoms (n = 32), we measured fecal bile acids using targeted metabolomics. We profiled the fecal microbiota by whole-genome metagenomics, and analyzed colonic gene expression patterns by RNA sequencing transcriptomics of mucosal biopsies. All individuals completed self-reported questionnaires on GI symptom severity and patterns (GSRS-IBS, Bristol Stool Form Scale-based stool diary). Spearman’s correlation with Benjamini-Hochberg multiple testing adjustment was used for correlation analyses, permutational ANOVA for analysis of gut microbial composition on Bray-Curtis dissimilarity, and the Wald test for analysis of differences in colonic gene expression. Differences in bile acid levels were tested with Kruskal-Wallis and post hoc Conover-Iman tests, with Benjamini-Hochberg adjustment.

Results

The levels of the hepatically synthesized primary bile acids, chenodeoxycholic acid and cholic acid, were lower in IBS-C (constipation-predominant IBS) compared to IBS-D (diarrhea-predominant IBS, p = 2.31e^-7), IBS-M (mixed type IBS, p = 0.002) and healthy controls (p = 2.57e^-6, Table 1). In IBS patients, levels of primary bile acids correlated with perceived symptom severity of constipation (negatively, rho = -0.3, p = 0.0002) and diarrhea (positively, rho = 0.26, p = 0.00106). When dividing IBS patients into quartiles by primary bile acid levels, 95 genes in the lowest quartile showed altered colonic expression. Gene-set enrichment analysis revealed enrichment of immune response-related genes, including those involved in chemokine and cytokine activity. Permutational ANOVA tests revealed altered gut microbiota composition in the two lowest quartiles of primary bile acid levels compared to healthy controls (p = 0.0001, p = 0.0003).

Conclusion

Alterations in fecal primary bile acid levels were associated with stool consistency in IBS, and lower levels of primary bile acids were linked with altered gut microbiota composition and expression of colonic immune-related genes, suggesting involvement of gut microbiota-bile acid-host crosstalk in symptom generation in IBS.

Introduction

While the mechanistic background to functional bowel disorders, including irritable bowel syndrome (IBS) is insufficiently understood, there is growing evidence that cytokine imbalance and inflammation may contribute to IBS pathogenesis.

Aims & Methods

We undertook a comprehensive investigation of serum cytokine profiles in individuals with functional bowel disorders (IBS n=164, functional constipation n=6, functional diarrhea n=3) and healthy controls (n=31), mapping their associations with clinical symptoms (GSRS-IBS, HAD), the gut microbiome (whole-metagenome sequencing of fecal samples), colonic mucosal responses (bulk RNA sequencing of sigmoid biopsies), intestinal physiological parameters, and the serum metabolome (liquid chromatography-mass spectrometry). Serum levels of IFN-γ, IL-1β, IL-6, IL-8, IL-10, IL-12p70, IL-13, TNF-α, IL-17A and CXCL10 were measured with Meso Scale Discovery (MSD) multiplex immunoassay kits.

Results

While cytokine levels did not differ significantly between persons with functional bowel disorders and healthy individuals, CXCL10 was reduced in participants with predominant diarrhea (IBS-D and functional diarrhea, p=0.01), whereas IL-12p70 was increased in predominant constipation (IBS-C and functional constipation, p=0.03), compared to healthy controls. Within both the full study cohort and participants with functional bowel disorders, IL-12p70 levels correlated with oroanal transit time (rho 0.20 and 0.19, p=0.007 and 0.01, respectively), and were inversely associated with fecal short-chain fatty acids (p=0.006, 0.01 and 0.03 for acetate, propionate and butyrate, respectively), as well as with the abundance of several butyrate-producing species. By contrast, IL-12p70 levels were positively associated with mucus-resident species (e.g., from the Akkermansia and Victivallis genera), with metabolites potentially derived from the degradation of mucosal carbohydrates, and with a distinct T helper 1 type response in colonic biopsies. Moreover, IL-12p70 levels correlated with anxiety (rho 0.17, p=0.03) in participants with functional bowel disorders.

Conclusion

Systemic cytokine profiles did not discriminate persons with functional bowel disorders from healthy individuals, but participants with predominant constipation and diarrhea exhibited distinct cytokine profiles. Alteration of systemic cytokines, notably IL-12p70, may result from disrupted mucosal barrier integrity and host-microbe homeostasis, and contribute to extraintestinal symptoms in a subset of individuals with functional bowel disorders.

Introduction

The pathophysiology of irritable bowel syndrome (IBS) includes alterations in gut-brain communication, gastrointestinal (GI) physiology and gut microbiota. The essential amino acid tryptophan (Trp) is metabolized in the gut to different classes of metabolites, including serotonin, kynurenine and indoles which have been implicated in the modulation of intestinal motility, immune function and pain sensation. Balanced Trp metabolism requires a complex host-microbiota interplay, which might be dysregulated in IBS. However, Trp metabolism is incompletely understood, and studies are needed to clarify its link with IBS and gastrointestinal physiology.

Aims & Methods

Aim: To profile fecal and circulating levels of Trp metabolites in IBS and explore relationships with oro-anal transit time (OATT) and symptoms.
Methods: We combined targeted and untargeted metabolomics to determine fecal and serum Trp metabolites in a cohort of healthy controls (HC; n=29) and IBS patients with predominant constipation (IBS-C; n=59), diarrhea (IBS-D; n=53) or mixed bowel habits (IBS-M; n=55). OATT was measured using radiopaque markers and participants completed questionnaires to assess GI symptom severity (GSRS-IBS), stool frequency and consistency (Bristol Stool Form Scale), non-GI somatic symptoms (PHQ-12), and symptoms of anxiety and depression (HAD). For correlation analyses, Spearman’s rho (r) was used.

Results

Levels of Trp did not differ significantly between HC and IBS subtypes. The Trp metabolite kynurenine, the microbially produced indole propionic acid (IPA) and the serotonin degradation metabolite 5-hydroxyindoleacetic acid (5-HIAA) were lower in IBS-C compared to HC and IBS-D. There were inverse correlations between OATT and both fecal (p<0.0001; r= -0.40) and circulating (p= 0.021; r= -0.17) Trp levels. Similarly, stool kynurenine and indoles were negatively correlated with OATT. Fecal but not circulating serotonin was positively correlated with OATT (p= 0.041; r= 0.14) while 5-HIAA showed a negative correlation (p<0.0001; r= -0.29), suggesting potential serotonin accumulation due to deficient degradation. In IBS patients, fecal Trp, 5-HIAA, IPA and kynurenine, correlated negatively with the proportion of hard stools (BSF1-2) and positively with the proportion of loose stools (BSF6-7). Finally, serotonin catabolites and IPA were negatively correlated with the severity of constipation and positively correlated with diarrhea. No statistically significant correlations were noted between these metabolites and abdominal pain, abdominal distension, early satiety, or postprandial fullness while bloating was positively correlated with circulating IPA (p= 0.018; r= 0.18). Additionally, we report a negative correlation between circulating serotonin with symptoms of depression and anxiety, which was not found for fecal serotonin. No significant correlations were noted with non-GI somatic symptoms.

Conclusion

Our results suggest that slower transit time may favour increased intestinal catabolism of Trp, directly or indirectly through its effects on the gut environment and the microbiota. In addition, we report an association of fecal Trp metabolites with bowel habits, in line with their relations to OATT. On the other hand, these same fecal metabolites did not correlate with other GI (i.e., pain and bloating) or non- GI symptoms. This study underscores the link between intestinal transit time and the regulation of host and microbial metabolites important for intestinal homeostasis.

Introduction

Population-based surveys have demonstrated substantial overlap of DGBI across multiple gastrointestinal (GI) regions and that this overlap is associated with increased health impairment. Comprehensive assessments of this overlap in patients with IBS are lacking, and the link with pathophysiology is not understood. In addition, IBS patients also frequently report overlapping non-GI comorbidities (somatic symptoms disorder , depression and anxiety) .The interaction between overlapping DGBI and non-GI comorbidities in IBS, and the impact of this interaction needs to be further investigated.

Aims & Methods

We included IBS patients with completed ROME II, III or IV questionnaires to assess overlapping DGBI in other anatomical GI regions (esophageal, gastroduodenal, anorectal). Patients completed validated questionnaires to measure the severity of GI (Gastrointestinal Symptom Rating Scale for IBS (GSRS-IBS)) and psychological (Hospital Anxiety and Depression Scale (HADS)) symptoms, IBS-specific quality of life (QOL) (IBS QOL), GI symptom-specific anxiety (Visceral Sensitivity Index (VSI)), and somatic symptoms (Patient Health Questionnaire-15 (PHQ-15) and Symptom Checklist-90 (SCL90)). Overlap of non-GI comorbidites (somatic symptom disorder, depression and anxiety) were assessed using suggested cut-off levels on PHQ, SCL-90 and HADS. Oro-anal transit time (OATT) was measured with radiopaque markers, and rectal sensitivity with balloon distensions using a barostat. GI symptoms following a Lactulose Nutrient Challenge Test (LNCT) were also recorded as a measurement of sensitivity to microbial fermentation and nutrients.

Results

We included 688 patients with IBS (70.5% female, mean age 37.3 ± 12.5 years; IBS-C 29.7 %, IBS-D 34.6 %, IBS-M 24.7%, and IBS-U 11.0%.). Overall, 68.3% of IBS patients had overlapping DGBI in other anatomical GI regions; one region: 39.4%, two regions: 23.3%, three regions: 5.7%. An overlapping esophageal DGBI was seen in 26.2%, a gastroduodenal DGBI in 31.6%, and anorectal DGBI in 38.1%. The prevalence of overlapping somatic symptom disorder in IBS was 53.8%, while depression and anxiety overlap was seen in 32.2% and 50.9% of the IBS patients. A higher number of non-GI comorbidities was associated with a greater number of overlapping DGBI in IBS (p<0.01).
With increasing number of overlapping DGBI diagnoses and non-GI comorbidities in IBS, there was a gradual increase in the severity of GI and psychological symptoms, as well as a gradual decrease in QOL (Table 1). In addition, with increasing number of overlapping diagnoses, a gradual increase in visceral sensitivity (reduction in rectal discomfort threshold and increased discomfort during LNCT) was seen, but no links with abnormal OATT, age or body mass index were noted.

Conclusion

Overlapping DGBI diagnoses and non-GI comorbidities are common among IBS patients. This overlap has an impact on both GI and non-GI symptoms, as well as a link with GI sensitivity. Assessment of overlapping DGBI and non-GI comorbidites seems to be of importance to optimize clinical management in patients with IBS.

Introduction

Food is often reported as a trigger for symptoms in patients with irritable bowel syndrome (IBS). One of the proposed pathophysiological mechanisms is a local immune response to food antigens, triggered by a preceding infection¹.

Aims & Methods

This study aimed to investigate rectal mucosal reactions to food in patients with IBS. Therefore, patients with IBS and food-related symptoms were included. Allergies to the food items that were tested during the colonoscopic antigen provocation (COLAP) were excluded by an allergologist. The COLAP was started in the absence of macroscopic abnormalities during sigmoidoscopy. A 200 μL solution of each food antigen (wheat, gluten, soy, and milk; administered in a randomized order and blinded), a negative control (NaCl 0.9%), and a positive control (histamine) were injected in the rectal mucosa, with a positive reaction defined as mucosal edema 12 minutes after injection and judged based on a previously used semiquantitative scale ². The reaction was independently judged by two endoscopists, resulting in a positive or negative reaction when they agreed and doubtful when they did not.

Results

Data from ten patients were available for this interim analysis. Nine were female, with a median age of 45 [range 24-58] years, and a median IBS symptom severity score of 356 [range 197-416], i.e. in the moderate to severe range. Six patients had predominant constipation, one predominant diarrhea, and three mixed bowel habits. COLAP was positive in seven patients, with a positive reaction for one food antigen in three patients and for two food antigens in four patients. In the other three patients, the test was only doubtful for one of the food antigens (n=2), or negative for all (n=1). A positive reaction to gluten was observed in five patients, to soy in three, to wheat in two, and to milk in one (Table). The reaction to histamine was positive in eight patients, with no positive reaction after injection of the negative control in all.

Conclusion

A local intestinal reaction to food antigens was seen in a subset of patients with IBS. Further research will focus on underlying pathophysiological mechanisms and clinical relevance of these findings.

References

1. Aguilera-Lizarraga J, Florens MV, Viola MF, Jain P, Decraecker L, Appeltans I, et al. Local immune response to food antigens drives meal-induced abdominal pain. Nature. 2021;590(7844):151-6.
2. Bischoff SC, Mayer J, Wedemeyer J, Meier PN, Zeck-Kapp G, Wedi B, et al. Colonoscopic allergen provocation (COLAP): a new diagnostic approach for gastrointestinal food allergy. Gut. 1997;40(6):745-53.