Quantitative studies of the toxicity of benzo(a)pyrene to a mouse liver epithelial cell strain in culture

The toxic effects of the carcinogen benzo(a)pyrene (BaP) were studied in a well-characterized epithelial cell strain NMuLi, derived from the livers of weanling Namru mice. These cells were extremely susceptible to the toxicity, 99% dying after a 6-day exposure to BaP, 5mug/ml. The toxic effects began between 11 and 24 hr postapplication of BaP to the cells and increased exponentially with the time of treatment. The toxicity was concentration dependent in cells treated for a specific time period. The survival curves were exponential and extrapolated to a survival fraction of 1.0. The toxic effects of BaP to logarithmically growing NMuLi were inhibited 40% by 7,8 benzoflavone, and the inhibition was concentration dependent. The 7,8-benzoflavone also inhibited aryl hydrocarbon hydroxylase (AHH) from NMuLi cell homogenates and microsomes by 99%. The concentration dependence for AHH inhibition by 7,8-benzoflavone paralleled its inhibition of cellular toxicity. The toxicity of BaP to these cells increased exponentially with the number of population doublings. Hence, the toxicity was 130 times greater in exponentially growing cells than in confluent cells. Levels of AHH, the enzyme that metabolizes BaP to its cytotoxic derivatives, were only 2.4 times higher in exponentially growing than in confluent cells, suggesting that cell division was responsible for the large differential toxicity. In addition, a toxic BaP metabolite was preferentially toxic to log-phase cells. The results indicate that the metabolism of BaP by AHH to produce cytotoxic metabolites, which may cause lesions that are expressed upon cell division, is responsible for the cytotoxicity of BaP to NMuLi.