Microbial Contribution to Proteolytic Homeostasis in the Gut

Abstract

Proteases mediate a vast array of physiological functions, and their enzymatic activity must be tightly regulated by protease inhibitors. Proteolytic homeostasis is crucial in the gastrointestinal tract, and disturbances are associated with several gastrointestinal disorders including irritable bowel syndrome (IBS), inflammatory bowel disease (IBD), colorectal cancer (CRC), and celiac disease (CeD). Perturbed proteolytic balance is hypothesized to contribute to disease pathophysiology by inducing structural and functional changes in the intestinal barrier and promoting inflammation, and modulation of this proteolytic imbalance through delivery of protease inhibitors represents a therapeutic target. Studies addressing proteolytic homeostasis in the gut have focused on proteases and protease inhibitors produced by the host, while largely ignoring the potential contribution of the microbiota. This is a major oversight given the wide repertoire of proteases and protease inhibitors produced by the intestinal microbiota, as well as the strong association between alterations in microbiota and several chronic intestinal disorders. We hypothesized that increased microbial proteolytic activity can elicit deleterious effects on the host, while microbial protease inhibitors such as those produced by probiotic strains may have a therapeutic role in gut inflammatory disorders. The aims of this thesis were: (1) to investigate the contribution of microbial proteolytic activity to the processes underlying disease pathogenesis, and, (2) to test the therapeutic potential of bacterial probiotic strains naturally expressing protease inhibitor genes. We first employed a reductionist model wherein gnotobiotic mice were colonized with microbial communities expressing a specific proteolytic functional phenotype in vitro. Colonization of germ-free mice with microbiota with high proteolytic activity resulted in increased systemic translocation of live bacteria and low-grade inflammation. Some of these effects were mitigated after treatment with the probiotic B. longum strain expressing its serine protease inhibitor (Srp). Moreover, treatment with this Srp-expressing B. longum strain prevented immunopathology in a mouse model of gluten sensitivity. Together, these results demonstrate the relevance of microbiota-derived proteases and protease inhibitors in the modulation of host responses and gut homeostasis.