Neutrophil influx into the intestinal lumen is a critical response to infectious agents, but is also associated with severe intestinal damage observed in idiopathic inflammatory bowel disease. The chemoattractant hepoxilin A3, an eicosanoid secreted from intestinal epithelial cells by the apically restricted efflux pump multidrug resistance protein 2 (MRP2), mediates this neutrophil influx. Information about a possible counterbalance pathway that could signal the lack of or resolution of an apical inflammatory signal, however, has yet to be described. We now report a system with such hallmarks. Specifically, we identify endocannabinoids as the first known endogenous substrates of the apically restricted multidrug resistance transporter P-glycoprotein (P-gp) and reveal a mechanism, which we believe is novel, for endocannabinoid secretion into the intestinal lumen. Knockdown or inhibition of P-gp reduced luminal secretion levels of N-acyl ethanolamine-type endocannabinoids, which correlated with increased neutrophil transmigration in vitro and in vivo. Additionally, loss of CB2, the peripheral cannabinoid receptor, led to increased pathology and neutrophil influx in models of acute intestinal inflammation. These results define a key role for epithelial cells in balancing the constitutive secretion of antiinflammatory lipids with the stimulated secretion of proinflammatory lipids via surface efflux pumps in order to control neutrophil infiltration into the intestinal lumen and maintain homeostasis in the healthy intestine.

Enteroaggregative Escherichia coli (EAEC) causes diarrhea and intestinal inflammation worldwide. EAEC strains are characterized by the presence of aggregative adherence fimbriae (AAF), which play a key role in pathogenesis by mediating attachment to the intestinal mucosa and by triggering host inflammatory responses. Here, we identify the epithelial transmembrane mucin MUC1 as an intestinal host cell receptor for EAEC, demonstrating that AAF-mediated interactions between EAEC and MUC1 facilitate enhanced bacterial adhesion. We further demonstrate that EAEC infection also causes elevated expression of MUC1 in inflamed human intestinal tissues. Moreover, we find that MUC1 facilitates AAF-dependent migration of neutrophils across the epithelium in response to EAEC infection. Thus, we show for the first time a proinflammatory role for MUC1 in the host response to an intestinal pathogen.

Enterohemorrhagic Escherichia coli (EHEC) adheres to intestinal epithelial cells, then stimulates the actin nucleation promoting factor N-WASP to induce localized actin assembly resulting in an actin “pedestal”, the function of which is poorly understood. EHEC also produces Shiga toxin (Stx), which penetrates the intestinal epithelium to cause a life-threatening renal and systemic disease. To assess the role of pedestal formation in colonization and disease, we utilized the murine pathogen Citrobacter rodentium, which also forms actin pedestals, and the genetically engineered C. rodentium (Φstx2dact), which additionally triggers Stx-mediated systemic disease. We found that an intestine-specific N-WASP-deficient (iNWKO) mouse suffered dramatically less colonization and disease than N-WASP-proficient littermate controls when infected with either strain. In addition, upon infection of wild type mice, mutants of C. rodentium or C. rodentium (Φstx2dact) that are specifically defective in pedestal formation demonstrated a relatively modest defect in cecal colonization and fecal shedding, but a more severe defect in colonization of the colonic mucosa. The C. rodentium (Φstx2dact) pedestal-defective mutant did not cause renal disease and, after normalizing for fecal bacterial load, was associated with a 16-fold lower risk of lethality. These findings suggest that the ability of an attaching and effacing pathogen to promote localized actin assembly contributes to virulence by promoting mucosal attachment.