Evidence for cytochrome P450 2E1-mediated toxicity of N-nitrosodimethylamine in cultured perivenous hepatocytes from ethanol treated rats.

The involvement of cytochrome P450 in the liver toxicity of the potent carcinogen, N-nitrosodimethylamine (NDMA) was investigated in hepatocytes isolated from the periportal or perivenous region by digitonin-collagenase perfusion. Exposure of hepatocytes in culture to NDMA (0.5 or 5 mM) for up to 18 hrs caused little damage, but after 42 hr loss of cell viability became evident, and the extent of cell death was higher in perivenous cells than in periportal cells. Pretreatment of rats with ethanol caused a dramatically enhanced cell damage in perivenous cells (80%) compared to periportal cells (45%). This ethanol pretreatment caused a several-fold induction of cytochrome P450 2E1, as determined both with Western blot and as NDMA demethylase activity, and the effect was observed almost exclusively in perivenous cells. Isoniazid, an inhibitor of cytochrome P450 2E1, completely protected against NDMA toxicity. Glutathione dependent cytoprotective mechanisms and lipid peroxidation did not appear to be critical in NDMA toxicity, as evidence by lack of potentiation of toxicity by buthionine sulfoximine, an inhibitor of glutathione synthesis, and by the absence of increased lipid peroxidation. Instead, the higher expression of cytochrome P450 2E1 in the perivenous cells seems to be the main determinant for the regiospecific toxicity of NDMA, and, consequently, probably also for the associated genotoxicity.