Clostridium difficile toxins A and B can alter epithelial permeability andpromote bacterial paracellular migration through HT-29 enterocytes

Clostridium difficile toxins A and B are the widely recognized etiologic ag ents of antibiotic-associated diseases ranging from diarrhea to pseudomembr anous colitis. We hypothesized that C. difficile toxins may alter intestina l epithelial permeability and facilitate bacterial penetration of the intes tinal epithelial barrier. Experiments were designed to clarify the effects of C. difficile toxins A and B on the flux of inert particles across HT-29 enterocyte monolayers, and to correlate these results with bacteria-enteroc yte interactions. in all experiments, mature, confluent HT-29 cultures were preincubated 16 h with toxin A or B (1-100 ng/mL). To study alterations in epithelial permeability, toxin-treated enterocytes were incubated with 5 p M solutions of 10- and 40-kD inert dextran particles. Toxin A, but not toxi n B, was associated with increased dextran flux through enterocyte monolaye rs. To study bacteria-enterocyte interactions, toxin-treated enterocytes we re incubated with 10(8) Salmonella typhimurium, Proteus mirabilis, or Esche richia coli. Although numbers of internalized bacteria were generally unaff ected, both toxins were associated with increased bacterial adherence, as w ell as increased bacterial transmigration through enterocyte monolayers. Ba cterial transmigration was significantly greater using toxin A- compared to toxin B-treated enterocytes, consistent with the observation that dextran flux was significantly greater using toxin A- compared to toxin B-treated e nterocytes. Thus intestinal colonization with toxigenic C. difficile may fa cilitate bacterial penetration of the intestinal epithelium by a mechanism involving increased permeability of the intestinal epithelial barrier.