The enterocyte is an active participant in the inflammatory and metabolic response to sepsis, endotoxemia and other critical illnesses and is the site for cytokine and acute phase protein production in these conditions. The role of the CCAAT/enhancer binding protein (C/EBP) family of transcription factors in the response to inflammatory stimuli in the enterocyte is not well understood. In the present study, we treated Caco-2 cells with IL-1beta and determined C/EBP DNA binding activity by electrophoretic mobility shift assay. The involvement of the alpha, beta, and delta isoforms was determined by supershift analysis and Western blot analysis of proteins from the nuclear fraction. The role of the mitogen-activated protein kinase (MAPK) signaling pathway was assessed by treating cells with the MAPK inhibitor PD-98059. Treatment of the Caco-2 cells with IL-1beta resulted in increased CCAAT/enhancer binding protein DNA binding activity. Supershift analysis and Western blotting indicated that this response to IL-1beta mainly reflected the delta isoform, and to a lesser degree the beta isoform. Treatment of the cells with PD-98059 inhibited the IL-1beta-induced increase in beta and delta activity. The results suggest that members of the CCAAT/enhancer binding protein family of transcription factors are activated in enterocytes during inflammatory conditions characterized by high levels of IL-1beta.