Necrotizing enterocolitis is characterized by the loss of epithelial barrier function, bacterial translocation into the intestinal wall and subsequent intestinal necrosis. Although many risk factors predisposing to the disease and some mediators contributing to the pathogenesis of NEC have been identified, the exact sequence of events leading to intestinal necrosis has not been delineated yet. The goal of the present study was a preliminary characterization of the role of epithelial apoptosis in the pathogenesis of NEC. NEC was induced either by intravenous injection of platelet activating factor (PAF) to juvenile rats or by formula feeding of neonatal rats combined with cold- and asphyxiastressing. Both PAF-induced and neonatal NEC was accompanied by widespread epithelial apoptosis in tissue sections of the small intestine as determined by fluorescence TUNEL staining. In order to delineate the mechanisms of epithelial apoptosis, mediator-induced apoptosis was assayed in IEC-6 rat small intestinal epithelial cells using fluorescence TUNEL staining, or by quantifying histone-bound DNA fragments. PAF caused only slight increase in the rate of apoptosis in IEC-6 cells. However, treatment of IEC-6 cells with a chemical •NO donor caused up to six fold increase of the rate of apoptosis in a time- and concentration -dependent manner. Apoptosis was induced following a 3 hrs delay with a T1/2 of 50 minutes and half maximal induction of apoptosis was observed at 0.6 μM/hr •NO production by the •NO donor. These data show that PAF is a potent inducer of apoptosis in the intestinal epithelium in vivo, but acts as a weak inducer of epithelial apoptosis in a tissue culture model. Given our data indicating that •NO is a potent inducer of apoptosis in IEC-6 cells, and PAF has been shown to mediate •NO production by a variety of cell types in the gut, we hypothesize that PAF-induced •NO production might mediate epithelial apoptosis in experimental models of NEC.