Effects of 1,1-Dichloroethene and of Some of Its Metabolites on the Functional Viability of Mouse Hepatocytes

1,1-Dichloroethene (DCE) is hepatotoxic in rodents, and theexpression of its toxicity involves probably its metabolism.In this study the role of DCE metabolites in the generationof the hepatotoxic lesion was investigated. Hepatocytes frommale BALB/c mice in suspension were used as the experimentalmodel. Cells were incubated with DCE for up to 5 hr and cellularviability was assessed by measurement of the release of lactatedehydrogenase into the medium and by alterations in the reductionof the dye 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazoliumbromide. After incubation for 3 hr DCE at 0.5 mM caused maximaltoxicity, whereas at 0.1 mM DCE was only marginally toxic. Cytotoxicitywas exacerbated by pretreatment of mice with buthionine sulfoximine(1.6 g/kg), an inhibitor of glutathione biosynthesis, given4 hr prior to hepatocyte isolation. Inclusion of N-acetylcysteine(10 mM) into the incubate protected cells against DCE-inducedcytotoxicity. Coincubation with octylamine (0.5 mM), an inhibitorof cytochrome P450, abolished the cytotoxic potential of 0.5mM DCE during incubation for 3 hr. DCE toxicity was increasedin hepatocytes from mice which had received ethanol or acetonein their drinking water, both of which induce levels of thehepatic cytochrome P450 isozyme P450 2E1. Incubation of cellswith the P450 2E1 inhibitors N,N-dimethylformamide (10 mM) ordiethyldithiocarbamate (100 µM) protected liver cellsagainst the detrimental effect of DCE. Pretreatment of animalswith phenobarbital, which induces the P450 2B subfamily, or3-methylcholan-threne, which induces P450 1A1, did not affectthe degree of hepatocytotoxicity elicited by DCE. The DCE metaboliteschloroacetic acid and dichloroacetaldehyde at 0.75 mM were toxictoward the cells; however, their toxic potency was inferiorto that of DCE. Dichloroacetic acid, another product of metabolicDCE oxidation, and S-(chloroacetyl)glutathione and glutathionylacetylglutathione,both of which are generated by conjugation of DCE metaboliteswith glutathione, at concentrations of up to 5 mM did not interferewith hepatocyte viability. The results suggest that (i) DCEundergoes metabolic toxification in mouse hepatocytes, (ii)P450 2E1 is responsible for the metabolic activation of DCE,and (iii) conjugation with glutathione is a detoxification step.