Generation and persistence of carcinogen-induced repair intermediates in rat liver DNA in vivo

Abstract

Chromatographic separation of native DNA from DNA containing single-stranded regions has been used to determine the relative concentrations of structural intermediates generated during chemically induced DNA repair. Single doses of each of ten compounds were administered to rats. After periods ranging from 90 min to 13 days, hepatic DNA was isolated and analyzed by stepwise elution from benzoylated diethylaminoethyl cellulose with 1.0 M NaCl followed by caffeine solution. The compounds used were benzo(a)pyrene, carbon tetrachloride, diethylnitrosamine, dimethylnitrosamine, ethyl methanesulfonate, galactosamine, N-hydroxy-2-acetylaminofluorene, methyl methanesulfonate, nitrosomorpholine, and beta-propiolactone. Doses of the various agents and/or treatment times were restricted such that hepatic necrosis did not occur. No increase in the amount of caffeine-eluted DNA occurred after administration of carbon tetrachloride or galactosamine. All the remaining chemicals caused a dose-dependent increase in the proportion of hepatic DNA eluted from benzoylated diethylaminoethyl cellulose with caffeine. In most cases, the varying times required to produce maximal increase in the proportion of caffeine-eluted DNA could be related to the rate of metabolism of the carcinogens. A distinction could be made according to whether repair intermediates were detected only within 24 hr of administration (ethyl methanesulfonate, methyl methanesulfonate, and beta-propiolactone) or were present for at least 3 days after treatment (diethylnitrosamine, dimethylnitrosamine, benzo(a)pyrene, N-hydroxy-2-acetylaminofluorene, and nitrosomorpholine). The data, considered with reference to previously ascribed modes of DNA repair for the respective adducts, suggest that base excision repair is immediately operative and rapidly completed in rat liver. However, reactions involved in the completion of nucleotide excision repair may be rate limiting, resulting in persistent structural damage to DNA. Implications of these findings for the use of benzoylated diethylaminoethyl cellulose chromatography as a carcinogen bioassay are considered.