A50 MICROBIOTA-MAST CELL INTERACTIONS IN A HUMANIZED MOUSE MODEL OF IBS

Accumulating evidence suggests that mast cells play an important role in the pathogenesis of Irritable Bowel Syndrome (IBS). Colonic mast cells are increased in patients with IBS, and their products, including histamine and tryptase are putative mediators of visceral hypersensitivity in IBS. To investigate the effect of gut microbiota on mast cells in a humanized mouse model of IBS. Germ-free NIH Swiss mice were colonized with fecal microbiota from two IBS patients (non-constipation) (n=24 mice/patient), and one healthy control (HC) (n=8). Fecal bacteria and their supernatants were collected from each mouse. Intestinal tissues were taken for immunohistochemistry and RNA scope assay. Mouse bone marrow derived mast cells (BMMC) or peritoneal mast cells (PMC) were obtained from additional mice and co-cultured with 10% bacteria or 10% bacterial supernatant for 4 hours. We studied mast cell degranulation (β-hexosaminidase assay), chemotaxis, adherence, histamine levels (ELISA), and expression of histamine receptors (H1R, H2R, H4R) and chemokines (CXCL12) using real time PCR. Tryptase positive mast cells and tryptase positive area were higher in the colon of IBS mice compared to HC mice. RNA scope demonstrated direct interactions between colonic mast cells and bacteria in mice with IBS but not HC microbiota. Microbial supernatant from IBS mice did not alter β-hexosaminidase and histamine release from BMMC. However, microbiota from IBS mice induced BMMC degranulation compared to microbiota from HC mice. Furthermore, bacterial supernatant from IBS mice increased BMMC chemotaxis and adherence. Expression of H2R, H4R and CXCL12 mRNA was higher in PMC stimulated by bacterial supernatant from IBS mice, compared to those from HC mice. Microbiota and microbial products from mice with IBS microbiota induce mast cell degranulation, enhance mast cell chemotaxis and adherence, and increase expression of H2R, H4R and CXCL12. Our results suggest that IBS microbiota and their products may induce mast cell infiltration and activation, likely resulting in gut dysfunction and symptom generation. CIHRNIH