PAF (Platelet activating factor) is a pro-inflammatory phospholipid implicated in the pathogenesis of necrotizing enterocolitis. We have previously shown that PAF induces TNF production in the small intestine. The rapid burst of cytokine production seen during inflammation can be triggered by the activation of transcription factors of the REL/NF-kB family. To examine if PAF activates NF-kB in the ileal epithelial cells, we injected PAF at 1.25 or 1.5 μg/kg iv, into young Sprague-Dawley rats. These doses cause only transient hypotension and no gross bowel necrosis. (Preliminary results showed that 1 μg/kg of PAF had no effect on NF-kB activation and 2 μg/kg produces bowel necrosis). Ileal epithelial cells were isolated 30 minutes after PAF injection. Nuclei were isolated, proteins extracted and nuclear translocation of NF-kB was analyzed by electrophoretic mobility shift assay using a [32P]-labeled oligonucleotide probe specific for NF-kB. We found that ileal epithelial cells of shams controls have a constitutive low level of NF-kB activity within nuclei. PAF at both 1.25 and 1.5 μg/kg significantly increased NF-kB translocation into the nucleus. These results suggest that NF-kB may mediate the upregulation of the cytokine production by epithelial cells in response to PAF. (funded by NIH grant HD31840).