Enterohemorrhagic Escherichia coli (EHEC) O157:H7 intimately attaches to intestinal epithelial monolayers and produces attaching and effacing (A/E) lesions. In addition, EHEC infection causes disruptions of intercellular tight junctions, leading to clinical sequelae that include acute diarrhea, hemorrhagic colitis, and the hemolytic-uremic syndrome. Current therapy remains supportive since antibiotic therapy increases the risk of systemic complications. This study focused on the potential therapeutic effect of an alternative form of therapy, probiotic Lactobacillus rhamnosus strain GG, to attenuate EHEC-induced changes in paracellular permeability in polarized MDCK-I and T84 epithelial cell monolayers. Changes in epithelial cell morphology, electrical resistance, dextran permeability, and distribution and expression of claudin-1 and ZO-1 were assessed using phase-contrast, immunofluorescence, and transmission electron microscopy and macromolecular flux. This study demonstrated that pretreatment of polarized MDCK-I and T84 cells with the probiotic L. rhamnosus GG reduced morphological changes and diminished the number of A/E lesions induced in response to EHEC O157:H7 infection. With probiotic pretreatment there was corresponding attenuation of the EHEC-induced drop in electrical resistance and the increase in barrier permeability assays. In addition, L. rhamnosus GG protected epithelial monolayers against EHEC-induced redistribution of the claudin-1 and ZO-1 tight junction proteins. In contrast to the effects seen with the live probiotic, heat-inactivated L. rhamnosus GG had no effect on EHEC binding and A/E lesion formation or on disruption of the barrier function. Collectively, these findings provide in vitro evidence that treatment with the probiotic L. rhamnosus strain GG could prove to be an effective management treatment for preventing injury of the epithelial cell barrier induced by A/E bacterial enteropathogens.

Background & Aims RAC1 is a GTPase that has an evolutionarily conserved role in coordinating immune defenses, from plants to mammals. Chronic inflammatory bowel diseases (IBD) are associated with dysregulation of immune defenses. We studied the role of RAC1 in IBD using human genetic and functional studies and animal models of colitis. Methods We used a candidate gene approach to HapMap-Tag single nucleotide polymorphisms (SNPs) in a discovery cohort; findings were confirmed in 2 additional cohorts. RAC1 mRNA expression was examined from peripheral blood cells of patients. Colitis was induced in mice with conditional disruption of Rac1 in phagocytes by administration of dextran sulphate sodium (DSS). Results We observed a genetic association between RAC1 with ulcerative colitis (UC) in a discovery cohort, 2 independent replication cohorts, and in combined analysis for the SNPs rs10951982 (Pcombined UC = 3.3 × 10–8, odds ratio [OR]=1.43 [1.26–1.63]) and rs4720672 (Pcombined UC=4.7 × 10–6, OR=1.36 [1.19–1.58]). Patients with IBD who had the rs10951982 risk allele had increased expression of RAC1, compared to those without this allele. Conditional disruption of Rac1 in macrophage and neutrophils of mice protected them against DSS-induced colitis. Conclusion Studies of human tissue samples and knockout mice demonstrated a role for the GTPase RAC1 in the development of UC; increased expression of RAC1 was associated with susceptibility to colitis.