Depletion of AhR Ligands in Diet Worsens Inflammatory Conditions in Mice with Chronic DSS‐Induced Colitis

The aryl hydrocarbon receptor (AhR) is an important transcription factor that regulates xenobiotic metabolism and is vital for maintaining immune cell populations, proliferation, and motility in the gastrointestinal (GI) tract. Recent studies have shown that deletion of AhR from mouse intestinal epithelial cells worsens experimental colitis, while activation of AhR by its agonists protects against acuteinflammation. AhR is activated by a variety of ligands including indole‐3‐carbinole (I3C) derived from cruciferous vegetables, such as broccoli and brussels sprouts. Whether AhR ligands derived from the diet impact chronic intestinal inflammation and/or cause gut microbiota alterations has not yet been investigated. Methods C57BL/6 mice (7‐9 wk, M) were fed either standard chow diet or purified diet (depleted of AhR ligands, referred to as ‐I3C diet) or the purified diet supplemented with 200 ppm of I3C (+I3C diet). Along with the diet, the mice were given either water or treated with 5 cycles of 2.5% dextran sulfate sodium (DSS) alternating weekly with water to induce chronic colitis. Total RNA was extracted from mouse colonic mucosal scrapings and amplified using Real Time qRT‐PCR. Myeloperoxidase (MPO) assay and histological analysis was used to assess inflammation. 16S sequencing was performed on DNA extracted from cecal contents for microbiota analysis (Zymo). Results : Depletion of AhR ligands in the diet increased severity of DSS induced colitis as evidenced by: i) a significant decrease in the colon length and an increase in colon weight to length ratio in chronic DSS mice fed the purified ‐I3C diet vs +I3C diet; ii) an increase in myeloperoxidase activity and increased histological scores in chronic DSS mice fed the ‐I3C diet vs DSS mice fed +I3C diet; iii) a significant decrease in mRNA expression of pro‐inflammatory cytokines, TNF‐α, IL‐1β, CCl20, and CXCl2 in the colon of DSS mice fed +I3C diet compared with ‐I3C diet; iv) a significant decrease in tight junction (occludin and ZO‐1) and adherens junction (E‐cadherin) protein levels in chronic DSS mice fed ‐I3C diet and this decrease was attenuated by supplementation of I3C. The different diets modulated gut microbiota under basal and in chronic colitis conditions. ‐I3C diet alone led to the emergence of the pathobiont, Parvibacter caecicola (phylum Actinobacterium) which was not detected in either the chow or +I3C diet. Chronic DSS and ‐I3C diet induced major alterations in phylum Firmicutes (with less abundance of Clostridia and an increase of Erysipeltrichia), and an increase in abundance of Bacteroides thetaoitaomicron (phylumActinobacterium). Interestingly, these changes in microbiota composition in DSS groups were reversed with I3C supplementation to diet. Conclusions Depletion of AhR ligands in the diet worsens colitis and induces dysbiosis. These results highlight the importance of diet in shaping gut microbiota and underscore the significance of potential nutritional strategies to alleviate colitis.