Mouse liver glutathione S-transferase mediated metabolism of methylene chloride to a mutagen in the CHO/HPRT assay.
Although methylene chloride (MC) is readily detectable as a bacterial mutagen, published studies in mammalian cells have been inconclusive. We have previously shown (Graves et al., 1995) that glutathione S-transferase (GST)-mediated metabolism of MC by mouse liver cytosol (S100 fraction) causes DNA single-strand (ss) breaks in CHO cells. In this study, MC GST metabolites were shown to cause mutations at the HPRT locus of CHO cells. The mutagenicity of MC was enhanced by exposing the cells in suspension rather than as attached cultures. The MC GST metabolite formaldehyde was mutagenic in independent experiments, although the number of mutants induced was lower than with the MC. CHO HPRT mutations were also induced by the reference genotoxin 1,2-dibromoethane (1,2-DBE), which is activated to a mutagen by GST-mediated metabolism. Assay of DNA ss breaks and DNA-protein cross-links at mutagenic concentrations of MC, formaldehyde or 1,2-DBE, showed that all three compounds induced DNA ss breaks, but only formaldehyde induced significant DNA-protein cross-linking. These results suggest that whilst formaldehyde may play a role in MC mutagenesis, its weak mutagenicity and the absence of significant DNA-protein cross-linking after MC exposure, leads to the conclusion that the MC DNA damage and resulting mutations are induced by the glutathione conjugate of MC, S-chloromethylglutathione.
Graves
R J
RJ
Zeneca Central Toxicology Laboratory, Macclesfield, Cheshire, UK.
Green
T
T
D000818:Animals; D001711:Biotransformation; D016466:CHO Cells; D006224:Cricetinae; D004247:DNA / Q000187:drug effects; D004249:DNA Damage; D015946:Ethylene Dibromide / Q000633:toxicity; D005557:Formaldehyde / Q000633:toxicity; D005978:Glutathione / Q000031:analogs & derivatives* / Q000378:metabolism / Q000633:toxicity; D005982:Glutathione Transferase / Q000378:metabolism*; D007041:Hypoxanthine Phosphoribosyltransferase / Q000235:genetics / Q000378:metabolism; D008752:Methylene Chloride / Q000378:metabolism* / Q000633:toxicity*; D051379:Mice; D008862:Microsomes, Liver / Q000378:metabolism; D009152:Mutagenicity Tests; D009153:Mutagens / Q000633:toxicity*; D011506:Proteins / Q000235:genetics / Q000378:metabolism
D009153:Mutagens; D011506:Proteins; C085664:S-chloromethylglutathione; D005557:Formaldehyde; D015946:Ethylene Dibromide; D008752:Methylene Chloride; D004247:DNA; D007041:Hypoxanthine Phosphoribosyltransferase; D005982:Glutathione Transferase; D005978:Glutathione