On the contribution of mucosal mast cells to the regulation of mouse intestinal barrier function

Abstract

The primary functions of the small intestine are the digestion and transport of luminal content and the absorption of nutrients. During these processes the intestinal mucosa is exposed to various ingested and resident pathogens. The ability of the intestinal wall to prevent transmucosal passage of toxins or of harmful micro-organisms and their products is defined as the intestinal barrier function. Defective intestinal barrier function plays a role in a number of disorders such as inflammatory bowel disease, acute pancreatitis (AP), parasite infections and chronic stress. The mechanisms underlying the regulation of the intestinal barrier under (patho)physiological conditions are not well known. The principal goal of this thesis was to investigate the contribution of mucosal mast cells (MMC) to the regulation of the mouse intestinal barrier function. The first chapter of this thesis focuses on the effects of probiotics on the impairment of the mucosal barrier during acute pancreatitis (AP). During clinical AP, a disturbed intestinal barrier function favors the passage of enteric bacteria and toxins resulting in infection. In a mouse model of AP we demonstrate that application of probiotics caused a complete recovery of the damaged epithelial integrity but only when applied before onset of AP. The second chapter of this thesis focuses on the intestinal mucosal barrier during a parasitic infection (intestinal schistosomiasis). Previously, it was shown that MMC can release large quantities of the mouse chymase mMCP-1 or the rat homologue (RMCP-2), which increases the intestinal permeability thereby facilitating the expulsion intestinal parasites. We investigated whether mMCP-1 induces changes in the mucosal barrier and facilitates passage of schistosome eggs across the intestinal wall. The intestinal barrier function of the ileal mucosa was investigated by measuring the electrical resistance of the tissue, the transmucosal passage of a marker molecule, and the secretory response in Ussing chambers. Structural integrity was assessed from the distribution of Occludin, Claudin-3 and ZO-1. We concluded that the impairment of the mucosal barrier function during schistosomiasis is not caused by mMCP-1. The results also indicate that schistosoma egg passage is facilitated by mechanisms which affect the epithelial cells rather than tight junctions. The remaining chapters of this thesis focus on the activation of MMC by the neuropeptide calcitonin gene-related peptide (CGRP). This activation is thought to contribute to the process of neurogenic inflammation via release of mast cell mediators which can regulate the intestinal barrier function. Using pharmacological and immunohistochemical techniques, the presence and functionality of the CGRP1 receptor was demonstrated on MMC. CGRP was found to stimulate a selective release of mMCP-1 from MMC without degranulation. Electron microscopy demonstrated a piecemeal mode of release upon CGRP stimulation of MMC. These findings support the hypothesis that the CGRP signaling from afferent nerves to MMC in the gastrointestinal wall is receptor-mediated. In summary, this thesis presents new mechanisms involved in the activation of intestinal mucosal mast cells. This activation is important for the maintenance of homeostasis and integrity of the mucosal barrier and also provides an appropriate response to injury.