Microcystin-LR (MCYST-LR) and nodularin (NOD) are known as tumor promoters in experimental animals and so present potential health threats for humans. Although their hepatotoxic mechanisms have been very well documented, many other effects of these toxins are relatively undescribed, indeed controversial, notably those related to their genotoxicity. In the present investigation, we examined how these toxins could induce DNA damage using a combination of in vitro and in vivo approaches. We first used the (32)P-postlabeling assay to test hydrophobic adduct formation on DNA from primary cultured rat hepatocytes treated with noncytotoxic concentrations of MCYST-LR and NOD (2 and 10 ng/mL). Analysis of the autoradiograms of DNA digests isolated from the hepatocytes did not show any hydrophobic DNA adduct formation. However, these toxins significantly decreased the amount of hydrophobic endogenous adducts, termed I compounds. We next investigated oxidative DNA damage by using the (32)P-postlabeling assay to analyze 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxo-dG) content as a biomarker of possible DNA lesions. Both MCYST-LR and NOD significantly enhanced 8-oxo-dG in time- and dose-dependent manner in vitro in primary cultured hepatocytes and in vivo in rat liver cells. Thus, it appears that the depletion of endogenous DNA adducts (I compounds) and/or the increase of 8-oxo-dG levels by MCYST-LR and NOD could be involved in the formation of hepatic tumors during long-term exposure to these cyanobacterial hepatotoxins.

No abstract

No abstract

The comparison of the behaviour of three dibenzoanthracene (DBA) isomers, dibenzo[a,c]anthracene (DB[a,c]A), dibenzo[a,h]anthracene (DB[a,h]A) and dibenzo[a,j]anthracene (DB[a,j]A), polycyclic aromatic hydrocarbons (PAHs), whose carcinogenicity varies from very potent to apparently inactive, has been carried out. Influence of norharman (NH; 9H-pyrido[3,4-b]indol) was investigated for mutagenicity (reversion of histidine prototrophy) on Salmonella typhimurium TA 100, using 3-methylcholanthrene (3-MC)-induced rat liver microsomes or S9 (post-mitochondrial fractions). A correlation with its influence, on the in vitro metabolism of radiolabelled molecules by the same enzymatic systems, was carried out. NH enhances the mutagenicities of DB[a,c]A and DB[a,h]A which are very well known mutagenic and carcinogenic PAHs. Contrary to its two isomers, the mutagenic potency of DB[a,j]A, which is considered as a weak mutagen and not a carcinogen, is strongly inhibited by NH. The balance sheets of the in vitro metabolism by microsomal enzymes, where the conjugation is excluded, were reported with or without NH. In the presence of the latter, the amounts of remaining DBAs slightly decreased while the metabolites covalently bound to microsomal proteins strongly decreased and the amount of hydrophobic metabolites highly increased. At the same time, the HPLC elution profiles of the metabolism pathways of the three DBAs are found to be modified in a similar way by NH: some of the metabolites are highly enhanced, and for all three DBAs, a tetraol, not detectable in the absence of NH, emerges. The results are discussed with regard to possible effects of NH.

The recent observation of a high level of adducts in mitochondrial DNA (mtDNA) of cells exposed to chemical carcinogens aroused new interest in the hypothesis that carcinogen-induced damage in mitochondria plays a role in one or more stages of carcinogenesis. In order to investigate whether differences in the metabolic activation of carcinogens have qualitative and quantitative effects on ml- and nuclear DNA (nuDNA) adduct formation, mice were exposed to the potent hepatocarcinogenic and sarcomagenic polycyclic hydrocarbon 7H-dibenzo[c,g]carbazole (DBC) and to three of its derivatives that show large differences in enzymatic activation: N-acetyl-DBC (N-AcDBC), which is carcinogenic for several tissues; 5,9-dimethyl-DBC (DiMeDBC), which is exclusively hepatocarcinogenic; and N-methyl-DBC (N-MeDBC), which is exclusively sarcomagenic. Adduct formation and toxic effects were measured over 48 hr. With a moderate 5 mumol/kg dose of DBC, the adduct level in liver 24 hr after treatment was always higher in nuDNA than in mtDNA; after 48 hr a substantial increase in the level of adducts in mtDNA was observed, with a parallel decrease in the level in nuDNA. With DiMeDBC, a 4.9-fold increase in mtDNA was seen at 48 hr, whereas, at the same dose, the non-hepatocarcinogenic N-MeDBC induced a very small number of adducts. In order to obtain a nearly identical level of adducts in nu- and mtDNA at 24 hr, the dose of DBC must be three times higher (15 mumol/kg); this and higher dose levels had a strong cytotoxic effect in liver cells. Qualitative differences in adduct distribution were observed on chromatograms of mtDNA and nuDNA, showing that the access to mtDNA is a complex process. Our results confirm that mouse liver mtDNA is a major target for DBC and its hepatocarcinogenic derivatives. The possible interference of genotoxic alterations in mtDNA with carcinogenic mechanisms is discussed.