Toxin A released by Clostridium difficile interacts with the single layer of intestinal epithelial cells that lines the host's intestinal tract and leads to mucosal damage and inflammation that manifests clinically as antibiotic-associated diarrhea and pseudomembranous colitis. Activation of the transcription factor NF-kappaB in intestinal epithelial cells is important for regulating the expression of epithelial cell proinflammatory genes and cell survival. However, the role of NF-kappaB activation in the pathogenesis of C. difficile toxin A-induced colitis is unknown. To determine the functional importance in vivo of NF-kappaB activation in intestinal epithelium in the pathogenesis of C. difficile-induced colitis, we used mutant mice that do not activate the classical NF-kappaB signaling pathway in intestinal epithelial cells due to a conditional deficiency in those cells of the IkappaB-kinase beta (IKKbeta) subunit of IKK. C. difficile toxin A challenge of intestinal loops in intestinal epithelial cell IKKbeta-deficient mice induced a rapid and significant increase in intestinal epithelial apoptosis compared with littermate controls. This was accompanied by a significant increase in acute mucosal inflammation, mucosal injury, luminal fluid secretion, and bacterial translocation. We conclude that activation of intestinal epithelial cell NF-kappaB by toxin A plays an important host mucosal protective role after C. difficile toxin A exposure that is mediated, at least in part, through promoting epithelial cell survival by abrogating epithelial cell apoptosis.