Role of nitric oxide in down-regulation of CYP2B1 protein, but not RNA, inprimary cultures of rat hepatocytes

There are conflicting reports about the role of nitric oxide in the down-re gulation of cytochrome P450 that occurs when animals or cultured hepatocyte s are exposed to inflammatory stimuli. Here, we investigated the participat ion of NO in the down-regulation of CYP2B1 by bacterial endotoxin (LPS) in rat hepatocytes cultured on Matrigel. LPS caused the down-regulation of CYP 2B1 mRNA to 20% of control values within 12 h of treatment, and this was no t reversed by concentrations of NO synthase inhibitors that completely bloc ked NO production. LPS was several orders of magnitude more potent in the d own-regulation of CYP2B1 mRNA than in induction of NO production. In contra st, concentrations of LPS in the 1 to 100 ng/ml range induced NO production and produced a rapid down-regulation of CYP2B1 protein to 30% and <5% of c ontrol at 6 and 24 h, respectively, that could be completely prevented both by inhibitors of NO synthase and by LY83583, which prevents NO synthase-2 induction. The blockade of CYP2B1 down-regulation by NO synthase inhibitors was reversed by arginine, and the NO donors S-nitrosoglutathione and S-nit roso-N-acetylpenicillamine mimicked CYP2B1 protein suppression. Taken toget her, these experiments demonstrate two independent mechanisms of CYP2B1 dow n-regulation by LPS: a rapid, NO-dependent suppression of the protein occur ring at high concentrations of LPS and a slower, NO-independent pretranslat ional suppression occurring at low concentrations of LPS.