Glutathione depletion in CYP2E1-expressing liver cells induces toxicity due to the activation of p38 mitogen-activated protein kinase and reduction of nuclear factor-kappaB DNA binding activity.
Depletion of glutathione (GSH) from CYP2E1-expressing cells by treatment with l-buthionine sulfoximine (BSO) causes decreased cell viability. The possible role of mitogen-activated protein kinases (MAPK) in this toxicity was evaluated. SB203580 [4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)1H-imidazole], an inhibitor of p38 MAPK decreased the BSO-dependent toxicity in HepG2 E47 cells, which express CYP2E1 and in hepatocytes from pyrazole-treated rats. Inhibitors of extracellular signal-regulated kinase, phosphatidylinositol 3-kinase, and c-Jun amino-terminal kinase were not protective. SB203580 did not prevent the loss of GSH nor lower the increase in reactive oxygen production; hence, protection by SB203580 was downstream of the elevated oxidative stress. Treatment with BSO caused activation of p38 MAPK whereas activation of nuclear factor-kappaB (NF-kappaB) was decreased; these effects were prevented by SB203580. We speculated that the decrease in NF-kappaB activation prevented production of hepatoprotective factors. One such factor could be nitric oxide (NO); indeed a NO donor decreased the BSO plus CYP2E1-dependent toxicity, whereas inhibition of inducible NO synthase (iNOS) potentiated toxicity. BSO treatment down-regulated iNOS and lowered NO levels, reactions blocked by SB203580; however, protection by SB203580 was the same in the absence or presence of an iNOS inhibitor, indicating that recovery of iNOS and NO production was not the mechanism by which SB203580 afforded protection against the BSO plus CYP2E1-dependent toxicity. Presumably other protective factors besides nitric oxide may be produced from activated NF-kappaB when p38 MAPK is inhibited by SB203580. These results suggest that the activation of p38 MAPK by BSO treatment in CYP2E1-expressing liver cells cause a loss in NF-kappaB-dependent production of hepatoprotective factors. This loss, coupled to CYP2E1-generated oxidant stress, synergize to promote cell injury.
Department of Pharmacology and Biological Chemistry, Mount Sinai School of Medicine, One Gustave L. Levy Place, New York, NY 10029, USA.
Defeng
D
Wu
Arthur
A
Cederbaum
D000818:Animals; D019328:Buthionine Sulfoximine / Q000037:antagonists & inhibitors / Q000633:toxicity*; D002478:Cells, Cultured; D019392:Cytochrome P-450 CYP2E1 / Q000378:metabolism*; D004247:DNA / Q000187:drug effects / Q000378:metabolism; D004347:Drug Interactions; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005978:Glutathione / Q000378:metabolism*; D022781:Hepatocytes; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors*; D016328:NF-kappa B / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism / Q000494:pharmacology; D019001:Nitric Oxide Synthase / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II; D011725:Pyridines / Q000494:pharmacology; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism; D048051:p38 Mitogen-Activated Protein Kinases
D004791:Enzyme Inhibitors; D007093:Imidazoles; D016328:NF-kappa B; D011725:Pyridines; D017382:Reactive Oxygen Species; D009569:Nitric Oxide; D019328:Buthionine Sulfoximine; D004247:DNA; D019392:Cytochrome P-450 CYP2E1; C496319:NOS2 protein, human; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496318:Nos2 protein, rat; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D005978:Glutathione; C093642:SB 203580