Freshly prepared isolated hepatocytes were shown to metabolize dimethylnitrosamine producing an alkylating agent which reacted with cellular DNA giving rise to 7-methylguanine and O6-methylguanine. Alkylation of DNA was prevented by aminoacetonitrile and was proportional to the dimethylnitrosamine concentration over the range of 1-90 μM. Isolated hepatocytes were able to catalyze the removal of O6-methylguanine from their DNA. After formation of this product by reaction with N-methyl-N-nitrosourea or dimethylnitrosamine to give extents of alkylation in the range of 0.1-15.0 μmol O6-methylguanine per mole of guanine in DNA, the loss produced in 3 h incubation was comparable to that seen in vivo from liver DNA alkylated to the same extent. The isolated hepatocytes therefore, provide a useful system in which factors influencing O6-methylguanine persistence in DNA can be studied.
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