Genotoxic potential of Microcystin‐LR and nodularin in vitro in primary cultured rat hepatocytes and in vivo in rat liver

Abstract

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.

Guidance values for microcystins in water and cyanobacterial supplement products (blue-green algal supplements): a reasonable or misguided approach?

“…The results of these assays strongly suggest that moderate to high concentrations of MC-LR are not directly genotoxic (Bouaicha and Maatouk, 2004a;Fessard et al, 2004;Zegura et al, 2003), however, do promote the generation of reactive oxygen species and subsequently lead to oxidative DNA damage and transient DNA strand breaks (Zegura et al, 2003;Maatouk, 2004a, 2004b). Consequently a tumour initiating capacity of MC-LR has been suggested.…”

The Kidney Vero-E6 Cell Line: A Suitable Model to Study the Toxicity of Microcystins

“…This data was obtained by the Comet assay, which measures DNA strand breaks that constitute primary DNA lesions with relevance for the formation of gene and chromosome mutations [48]. Several mutagenicity studies excluded the hypothesis of MCLR being a mutagen [49,50], which was further supported by the fact that MCLR do not form adducts with DNA that would represent a pre-mutagenic lesion [51]. On the other hand, some authors reported that MCLR-induced DNA damage was a consequence of early apoptosis due to oxidative stress and not a real genotoxic effect [52].…”

Microcystin-LR is not Mutagenic in vivo in the .LAMBDA./lacZ Transgenic Mouse (Muta Mouse)

“…So it may be inferred that some other mutagenic toxins present in the extracts or diŠerent routes of administration might be responsible for the positive results observed in those studies. However, MCLR treatment caused enhanced formation of 8-oxo-7,8-dihydro-2?-deoxyguanosine in a time-and dosedependent manner in vitro in primary cultured hepatocytes and in vivo in rat liver cells that could involve in the formation of hepatic tumors during long-term exposure to this cyanobacterial hepatotoxin (33). In contrast, in our study, under present experimental conditions, MCLR failed to induce mutation in both target genes (lacZ and cII) in liver and lungs of TG mouse.…”