Cytotoxicity of acetaminophen in human cytochrome P4502E1-transfected HepG2 cells.
Acetaminophen (APAP) when administered in excess can cause severe hepatic necrosis in vivo. To study the mechanism of APAP toxicity and the role of cytochrome P450, a previously established human hepatoma HepG2 subline, MVh2E1-9, that constitutively expresses human CYP2E1 was used as a model. At high concentrations (above 5 mM) and when intracellular reduced glutathione (GSH) was depleted, APAP caused severe cytotoxicity in MVh2E1-9, but not in MV-5 cells which lack CYP2E1. The APAP cytotoxicity was dependent on the concentration of APAP and time of exposure, and could be blocked by 4-methylpyrazole, ethanol, diallyl sulfide, N-acetylcysteine and N-t-butyl-alpha-phenylnitrone, but not by propylgallate, an inhibitor of lipid peroxidation. Significantly more 14C-labeled APAP protein adduct was detected in MVh2E1-9 cells than MV-5 cells, especially after depletion of GSH. The formation of the APAP adducts could be inhibited by the same agents which prevent APAP cytotoxicity. At a lower concentration (1-2 mM), APAP inhibited proliferation in both MVh2E1-9 and the control MV-5 cells to similar extents. This antiproliferative action of APAP did not require depletion of GSH as did the cytotoxic action of APAP. These data suggest that APAP has a dual toxic effect on MVh2E1-9 cells: a P450-independent antiproliferative effect and the CYP2E1-dependent cytotoxic effect. These results demonstrate the ability of human CYP2E1 to activate APAP to reactive metabolites which form covalent protein adducts and cause toxicity to a hepatoma cell line.
Dai
Y
Y
Department of Biochemistry, Mount Sinai School of Medicine, City University of New York, USA.
Cederbaum
A I
AI
D000082:Acetaminophen / Q000493:pharmacokinetics / Q000633:toxicity*; D000975:Antioxidants / Q000494:pharmacology; D001711:Biotransformation; D002455:Cell Division / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D019392:Cytochrome P-450 CYP2E1; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism*; D005978:Glutathione / Q000378:metabolism; D006801:Humans; D008861:Microsomes / Q000187:drug effects / Q000378:metabolism; D010084:Oxidation-Reduction; D010089:Oxidoreductases, N-Demethylating / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D014407:Tumor Cells, Cultured
D000975:Antioxidants; D000082:Acetaminophen; D003577:Cytochrome P-450 Enzyme System; D019392:Cytochrome P-450 CYP2E1; D010089:Oxidoreductases, N-Demethylating; D005978:Glutathione