Repair of iron-induced DNA oxidation by the flavonoid myricetin in primaryrat hepatocyte cultures

Oxidative DNA damage and its repair in primary rat hepatocyte cultures was investigated following 4 h of incubation with the toxic iron chelate, ferri c nitrilotriacetate (Fe-NTA), in the presence or absence of the potent prot ective flavonoid myricetin (25-50-100 mu M). Seven DNA base oxidation produ cts were quantified in DNA extracts by gas chromatography-mass spectrometry (GC-MS) in selected ion monitoring mode. Concomitantly, DNA repair capacit y of hepatocytes was estimated by the release of oxidized-base products int o culture media, using the same GC-MS method. A genotoxic effect of Fe-NTA (100 mu M) in hepatocytes was evidenced by a severe increase in DNA oxidati on over basal levels, with accumulation in cellular DNA of five oxidation p roducts derived from both purines and pyrimidines. This prooxidant effect o f iron was also noted by an induction of lipid peroxidation. estimated by f ree malondialdehyde production. Addition of increasing concentrations of my ricetin (25-50-100 mu M) simultaneously with iron prevented both lipid pero xidation and accumulation of oxidation products in DNA. Moreover, as an act ivation of DNA repair pathways, myricetin stimulated the release of DNA oxi dation bases into culture media, especially of purine-derived oxidation pro ducts. This removal of highly mutagenic oxidation products from DNA of hepa tocytes might correspond to an activation of DNA excision-repair enzymes by myricetin. This was verified by RNA blot analysis of DNA polymerase beta g ene expression which was induced by myricetin in a dose-dependent manner. T his represented a novel and original mechanism of cytoprotection by myricet in against iron-induced genotoxicity via stimulation of DNA repair processe s. Since iron-induced DNA damage and inefficient repair in hepatocytes coul d be related to genotoxicity and most probably to hepatocarcinogenesis, mod ulation of these processes in vitro by myricetin might be relevant in furth er prevention of liver cancer derived from iron overload pathologies. (C) 1 999 Elsevier Science Inc.