Immunocytochemical visualization of DNA adducts in mouse tissues and human white blood cells following treatment with benzo[a]pyrene or its diol epoxide. A quantitative approach

The formation and stability of benzoa]pyrene DNA adducts were studied in tissues of BALB/c mice exposed to benzoa]pyrene (Ba]P). The DNA adducts were visualized with an immunocytochemical peroxidase staining technique using an antiserum specific for the major Ba]P-derived adduct in DNA (+/-)trans-7,8-dihydroxy-anti-9,10-epoxy-7,8,9,10- tetrahydrobenzoa]pyrene (BPDE-N2-dG)]. The nuclear staining density was measured by microdensitometry. When mice were treated with an increasing dose of Ba]P the nuclear staining increased in the tissues studied (lung, heart and kidney). A linear relationship was found between the immunocytochemical nuclear staining signal and the actual DNA adduct level in the lung as measured by 32P-postlabeling. Maximum adduct formation was found 5 days after a single i.p. injection of Ba]P. Adduct levels decreased gradually after 7 days, but even after 61 days a slight specific staining was still present, suggesting that not all adducts had disappeared at that time. As judged from the disappearance of 3H]thymidine from prelabeled DNA the loss of adducts from the lung was not a result of DNA repair but one of cell turnover. In human white blood cells Ba]P-derived adducts could be detected after in vitro incubation with the reactive metabolite of Ba]P (BPDE). Dose-response studies demonstrated a positive relationship between BPDE-DNA adduct formation, the immunocytochemical staining signal and the BPDE concentration in the culture medium.