Microbial-derived purines in intestinal homeostasis

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PROJECT SUMMARY/ABSTRACT In a healthy gastrointestinal (GI) tract, intestinal epithelial cells (IECs) form a dynamic physical and biochemical barrier that maintains homeostasis by isolating the host immune system from an external environment of pathogenic and commensal microorganisms. Disruption of this barrier increases bacterial translocation, triggering inappropriate immune responses and unresolving inflammation. Dysregulation of IEC barrier in inflammatory bowel disease (IBD) and dysbiosis of the intestinal microbiota coincides with profound shifts in metabolic energy stores, especially in the colon, which exists in an energetically-vulnerable state of physiologic hypoxia. Substantial interest lies in understanding immunometabolism as a window to the molecular mechanisms of inflammatory progression or resolution, such as how metabolically active microbiota-derived small molecules promote intestinal homeostasis. Our ongoing work has identified purines as significant products of the intestinal microbiota. We identified a strong capacity for IECs to salvage hypoxanthine (Hpx) for ATP production, coinciding with improved barrier and wound healing capabilities. Furthermore, an unbiased microarray analysis of Hpx-treated T84 IECs identified TP53-inducible glycolysis and apoptosis regulator (TIGAR) as an induced target. We postulate that TIGAR induction by Hpx is central to mediating metabolism in response to purine availability or need for salvage due to stress-induced energetic deficits. In ongoing studies, we found that germ-free (GF) mice monocolonized with purine-producing bacteria have significantly increased colon tissue Hpx, with concomitant TIGAR induction. Further studies showed that the monocolonized mice were protected from dextran sulfate sodium (DSS)-induced colitis. Based on these results, we hypothesize that microbial-derived purines significantly contribute to intestinal homeostasis through TIGAR-mediated metabolic shifts. Aim 1 will define the mechanism(s) of Hpx-dependent TIGAR induction and the impact of that induction on purine metabolism. Aim 2 will elucidate the contribution of microbial-derived purines to intestinal homeostasis. Aim 3 will utilize TIGAR knockout (-/-) mice to assess the baseline contribution of TIGAR to intestinal homeostasis and inflammation resolution. This work lays a foundation for an IBD therapeutic in which patients are treated with nonpathogenic microbes specifically modified to produce small molecules (e.g. purines) that drive wound healing and inflammatory resolution. This proposal will provide outstanding mentorship and training under the guidance of an experienced sponsor in the ideal environment of a rigorous basic science lab that is well-integrated clinically, while having the necessary resources and mentorship to complete each aspect of this project, including a mentorship team within the Mucosal Inflammation Program. This training will foster the applicant’s independent science research skills in cross-cutting work that advances therapeutics for tissue damage repair and novel disease target identification.