CYTAKINE-MEDIATED PRODUCTION OF NITRIC-OXIDE IN ISOLATED RAT HEPATOCYTES IS DEPENDENT ON CYTOCHROME P-450III ACTIVITY

To investigate the role of the cytochrome P-450 system in NO synthesis , cytochrome P-450IIIA, IIE and IA activities were specifically inhibi ted by cimetidine (IIIA), clotrimazole (IIIA), benzoflavone (IA) and d isulfiram (IIE) in a model of cultured rat hepatocytes. Cytokine-induc ed NO synthesis was significantly decreased in the presence of cimetid ine and clotrimazole. Kinetic analysis revealed a non-competitive mode of inhibition (K-i = 21 mM, cimetidine; K-i = 13 mu M, clotrimazole), Reverse transcriptase-PCR and immunoblot analysis revealed no signifi cant change in steady state levels of iNOS mRNA and protein expression with P-450IIIA inhibition. Purified iNOS enzyme activity was not alte red. These data suggest that cytokine-mediated hepatocyte synthesis of NO is dependent upon P-450IIIA activity, which functions in a post-tr anslational capacity.