Kinetics of induction of DNA adducts, cell proliferation and gene mutations in the liver of Muta™Mice treated with 5,9-dimethyldibenzo[c,g]carbazole

Abstract

5,9-Dimethyldibenzo[c,g]carbazole (DMDBC) is a synthetic derivative of the environmental pollutant 7H-dibenzo[c,g]carbazole. DMDBC is a potent genotoxic carcinogen specific for mouse liver. Using the MutaMouse lacZ transgenic mouse model and a positive selection assay, we measured lacZ mutant frequency (MF) in the liver 28 days after a single s.c. administration of DMDBC at 3, 10, 30, 90 or 180 mg/kg. MF remained low at 3 and 10 mg/kg, but increased markedly from 30 mg/kg onwards. To investigate the reason for this non-linear response, we examined mechanisms potentially involved in mutation induction in the liver. Genotoxic effects such as DNA adduct formation were detected in 32P-post-labelling studies. Liver sections were examined for microscopic changes and cell proliferation. These parameters, and MF, were studied 2, 4, 7, 14, 21 and 28 days after a single s.c. administration of 10 or 90 mg/kg DMDBC. At 10 mg/kg, a dose found to double the MF on day 28, DNA adducts reached a level of 200-600 adducts per 10(8) nucleotides from day 4 to day 28. No changes in histology or cell proliferation were detected at this low dose. At 90 mg/kg, MF increased gradually from day 7 to day 28 (maximum 44-fold). The DNA adduct level ranged from 400 to 4500 adducts per 10(8) nucleotides on day 2, then stabilized at approximately 400 adducts per 10(8) nucleotides on day 4. An early cytotoxic effect was detected microscopically in centrilobular hepatocytes, and was followed by liver cell proliferation. These data suggest that the marked increase in MF in MutaMouse liver after treatment in vivo with DMDBC at 90 mg/kg may be explained by the induction of replicative DNA synthesis due to a cytotoxic effect, allowing the fixation of persistent DNA adducts into mutations.

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

“…The detection of apoptosis at earlier applications was the consequence of using a more sensitive technique (TUNEL) for detecting apoptotic nuclei, and indicated that the critical adduct level from which they induced toxicity could be lower. In our study, large interanimal variations were observed in the apoptotic index of the groups that received a high number of applications of 7H-DBC, and this fact is in agreement with the results of Tombolan et al (1999a) with respect to the variations of the mutation frequency in liver from mice treated with high doses of 7H-DBC. In accordance with our previous results, the apoptosis index revealed that DBC toxicity increased with the number of applications and was higher in periportal areas.…”

Dibenzanthracenes and benzochrysenes elicit both genotoxic and nongenotoxic events in rat liver ‘stem-like’ cells

“…One of the potential mechanisms involved could be the disruption of cell cycle control leading to enhanced cell proliferation, which may potentiate genotoxic effects of PAHs and contribute to tumour promotion (Tombolan et al, 1999;Bishop et al, 2001). We have previously used contact-inhibited WB-F344 cells as a model to study impact of PAHs on cell proliferation, and we have found that the proliferative effects of PAHs in this cell line are AhR-dependent (Chramostová et al, 2004;Andrysík et al, 2007).…”

Genotoxicity of 3-methylcholanthrene in liver of transgenic Big Blue� mice

“…1), because their presence is highly dependent on diet, age, and gender [Lu et al, 1990;Patel et al, 1998]. A recent study using other transgenic mice with a reporter gene (Muta™Mouse) showed a relationship between adduct formation and mutant induction following a 5,9-dimethyldibenzo[c, g]carbazole exposure [Tombolan et al, 1999]. Nevertheless, in this study a linear relationship between the number of detected adducts and the mutation induction factor was evidenced.…”