Role of Microbiota-Derived Bile Acids in Enteric Infections

Bile acids are abundant in the mammalian gut where they undergo bacteria-mediated transformation, generating a large pool of bioactive molecules. Although bile acids are known to affect host metabolism, cancer progression and innate immunity, it is unknown whether they affect adaptive immune cells such as T helper cells expressing IL-17a (Th17 cells) and regulatory T cells (Tregs). By screening a library of bile acid metabolites, we identified two distinct derivatives of lithocholic acid (LCA), 3-oxoLCA and isoalloLCA, as T cell regulators. 3-oxoLCA inhibited Th17 cell differentiation by directly binding to its key transcription factor RORγt (retinoid-related orphan receptor γt) and isoalloLCA enhanced Treg differentiation through the production of mitochondrial reactive oxygen species (mitoROS), leading to increased FoxP3 expression. IsoalloLCA-mediated Treg enhancement required an intronic FoxP3 enhancer, the conserved noncoding sequence 3 (CNS3), a distinct mode of action from previously-identified Treg enhancing metabolites that require CNS1. Administration of 3-oxoLCA and isoalloLCA to mice reduced Th17 and increased Treg cell differentiation in the intestinal lamina propria. Our data suggest novel mechanisms by which bile acid metabolites control host immune responses by directly modulating the Th17 and Treg balance.

The gastrointestinal tracts of mammals are colonized by hundreds of microbial species that contribute to health, including colonization resistance against intestinal pathogens 1 . Many antibiotics destroy intestinal microbial communities and increase susceptibility to intestinal pathogens 2 . Among these, Clostridium difficile, a major cause of antibiotic-induced diarrhea, greatly increases morbidity and mortality in hospitalized patients 3 . Which intestinal bacteria provide resistance to C. difficile infection and their in vivo inhibitory mechanisms remain unclear. By treating mice with different antibiotics that result in distinct microbiota changes and lead to varied susceptibility to C. difficile, we correlated loss of specific bacterial taxa with development of infection. Mathematical modeling augmented by microbiota analyses of hospitalized patients identified resistance-associated bacteria common to mice and humans. Using these platforms, we determined that Clostridium scindens, a bile acid 7-dehydroxylating intestinal bacterium, is associated with resistance to C. difficile infection and, upon administration, enhances resistance to infection in a secondary bile acid-dependent fashion. Using a workflow involving mouse models, clinical studies, metagenomic analyses and mathematical modeling, we identified a probiotic candidate that corrects a clinically relevant microbiome deficiency. These findings have implications for rational design of targeted antimicrobials as well as microbiome-based diagnostics and therapeutics for individuals at risk for C. difficile infection.

GPBAR1 (TGR5 or M-BAR) is a G protein-coupled receptor for secondary bile acids that is highly expressed in monocytes/macrophages. In this study, we aimed to determine the role of GPBAR1 in mediating leukocyte trafficking in chemically induced models of colitis and investigate the therapeutic potential of BAR501, a small molecule agonist for GPBAR1. These studies demonstrated that GPBAR1 gene ablation enhanced the recruitment of classically activated macrophages in the colonic lamina propria and worsened the severity of inflammation. In contrast, GPBAR1 activation by BAR501 reversed intestinal inflammation in the trinitrobenzenesulfonic acid and oxazolone models by reducing the trafficking of Ly6C+ monocytes from blood to intestinal mucosa. Exposure to BAR501 shifted intestinal macrophages from a classically activated (CD11b+, CCR7+, F4/80-) to an alternatively activated (CD11b+, CCR7-, F4/80+) phenotype, reduced the expression of inflammatory genes (TNF-α, IFN-γ, IL-1β, IL-6, and CCL2 mRNAs), and attenuated the wasting syndrome and severity of colitis (≈70% reduction in the Colitis Disease Activity Index). The protective effect was lost in Gpbar1-/- mice. Exposure to BAR501 increased the colonic expression of IL-10 and TGF-β mRNAs and the percentage of CD4+/Foxp3+ cells. The beneficial effects of BAR501 were lost in Il-10-/- mice. In a macrophage cell line, regulation of IL-10 by BAR501 was GPBAR1 dependent and was mediated by the recruitment of CREB to its responsive element in the IL-10 promoter. In conclusion, GPBAR1 is expressed in circulating monocytes and colonic macrophages, and its activation promotes a IL-10-dependent shift toward an alternatively activated phenotype. The targeting of GPBAR1 may offer therapeutic options in inflammatory bowel diseases.