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Abstract: The murine model of Citrobacter rodentium infection has been used to complement in vitro studies of enteropathogenic Escherichia coli (EPEC) infections of human intestinal epithelial cells (IECs). However, the differences in epithelial cell responses between these two models are not fully understood. We used an in vitro model of C. rodentium infection to examine important, yet incompletely understood, cellular responses of murine IECs to this pathogen. C. rodentium attached to CMT-93 cells and disrupted their tight junctional expression of claudins-4 and -5. This was associated with a loss of barrier function that required live bacteria and was partially prevented by the inhibition of Rho kinase. Furthermore, C. rodentium caused an upregulation of IEC apoptosis that was associated with the cytoplasmic accumulation of apoptosis-inducing factor, but not with the activation of caspase-3. These studies demonstrate for the first time that C. rodentium affects murine IECs in ways that may be similar, but distinct, to the effects of EPEC on human IECs. [Copyright &y& Elsevier]

Abstract: Enteropathogenic Escherichia coli (EPEC) is an important cause of diarrhea, particularly among infants in developing countries. An increase in intestinal permeability due to EPEC infection has been suggested as a factor in the development of diarrhea. Abnormally high levels of programmed cell death (apoptosis) of intestinal epithelial cells can lead to increased intestinal permeability. The effects of EPEC on cell apoptosis remain incompletely understood. This study characterized the mechanisms of EPEC-induced epithelial apoptosis and examined whether this effect contributes to heightened permeability in an in vitro model of infection. We report that EPEC-induced apoptosis in T84 intestinal epithelial cells via a mechanism involving caspases-3, -6, -8, and -9, the cleavage of PARP, and oligonucleosome formation. In addition, EPEC time-dependently increased paracellular permeability as assessed by transepithelial resistance and the apical-to-basolateral movement of 3000MW dextran. Furthermore, EPEC infection led to the cleavage and mislocalization of tight junctional ZO-1 and occludin. However, pharmacological inhibition of caspases did not prevent the EPEC-induced disruptions in epithelial barrier structure and function. Taken together, these results suggest that a caspase-dependent upregulation in epithelial cell apoptosis during EPEC infection occurs independent of impaired intestinal barrier function. [Copyright &y& Elsevier]