Induction of DNA Synthesis in Mouse Liver Following Increases of DNA Adduct Levels Elicited by Very Low Cumulative Doses of the Genotoxic Hepatocarcinogen 7H-dibenzo[c,g]carbazole

Abstract

The purpose of this work was to investigate the administration of very low but repeated doses of a genotoxic carcinogen and an eventual correlation with cellular DNA synthesis. The compound 7H-dibenzo[c,g]carbazole is a genotoxic carcinogen in the mouse liver and was administered topically at the dose of 13.35 microg per animal every 2 days to give a total of 13 applications. Animals were sacrificed 48 hours after every 2 applications until the 10th treatment, then 48 hours after every treatment. Postulated genotoxic effects such as DNA adduct formation were detected by the 32P-post labeling assay. Liver sections were examined for microscopic changes and DNA synthesis. Results showed an increase of the total DNA adduct level in the liver throughout the study with a slowing down in the level after the sixth application of the compound. This change could correspond to the onset of DNA synthesis and to the moderate hepatocellular apoptosis which was observed. The DNA synthesis, which was considered to be secondary to the cytotoxicity induced by the high level of DNA adducts altering normal cellular activity, could also be the opportunity to fix the DNA adducts into heritable mutations. These results raise the question regarding the risk assessment in humans exposed regularly to very low doses of chemicals in the environment: for non-proliferating tissue, the regular accumulation of DNA adducts could remain silent until a "threshold level" is reached from which stimulation of the DNA synthesis may fix the DNA adducts into heritable mutations, eventually leading to tumors.

Histopathological and Molecular Changes During Apoptosis Produced by 7H-Dibenzo[c,g]-Carbazole in Mouse Liver

“…In this work we have analyzed the early damage produced by this compound in mouse liver by means of the quantification of apoptosis and the analysis of apoptosis related genes. We have previously reported the effect of this compound in mouse liver after repeated administration of low doses of DBC (Dorchies et al, 2001). Although a marked increase of the total adduct level in the liver was observed from the second application of the compound until the thirteenth, in that work we only observed hepatocellular apoptosis after the 8th cumulative application of DBC.…”

“…We have previously reported the lesions in mouse liver produced by the cutaneous administration of 7H-DBC (Dorchies et al, 2001). No significant compoundrelated histological changes in the liver of mice were observed until the eight application of 7H-DBC, where a moderate apoptosis and single cell necrosis (presence of unicellular necrosis associated with few inflammatory cells) were observed.…”

“…As for the results obtained with Bax and Fas, the areas of stained hepatocytes were much more extensive in liver from mice treated with 12 applications of 7H-DBC (Figure 3). DISCUSSION 7H-DBC is described as a genotoxic compound (Périn et al, 1988) that forms DNA adducts (Périn-Roussel et al, 1995) and produces tumours in mouse liver (Valéro et al, 1985;Dorchies et al, 2001). In this work we have analyzed the early damage produced by this compound in mouse liver by means of the quantification of apoptosis and the analysis of apoptosis related genes.…”

“…In addition, Tombolan and colleagues (Tombolan et al, 1999a(Tombolan et al, , 1999b) demonstrated this mutagenic activity in vivo using the lacZ transgenic mouse assay. In previous work (Dorchies et al, 2001) we have described the kinetics of adduct formation and cell proliferation in liver from female mice treated with low cumulative doses of 7H-DBC. The protocol used in that work reproduced closely the risks for humans exposed to very low but regular doses of this molecule or similar compounds.…”

“…This aromatic hydrocarbon is widely found in various sources such as tobacco smoke and coal, and induces both local and distant tumors in mice, having greater effects in female than in male animals (Périn et al, 1984;Valéro et al, 1985;Dorchies et al, 2001). The genotoxic activity of this compound has been established using different in vitro approaches (Périn et al, 1988) and 15 separate DNA adducts induced by 7H-DBC have been detected in mouse liver (Périn-Roussel et al, 1995).…”