Nitric oxide donors prevent while the nitric oxide synthase inhibitor L-NAME increases arachidonic acid plus CYP2E1-dependent toxicity.
Polyunsaturated fatty acids such as arachidonic acid (AA) play an important role in alcohol-induced liver injury. AA promotes toxicity in rat hepatocytes with high levels of cytochrome P4502E1 and in HepG2 E47 cells which express CYP2E1. Nitric oxide (NO) participates in the regulation of various cell activities as well as in cytotoxic events. NO may act as a protectant against cytotoxic stress or may enhance cytotoxicity when produced at elevated concentrations. The goal of the current study was to evaluate the effect of endogenously or exogenously produced NO on AA toxicity in liver cells with high expression of CYP2E1 and assess possible mechanisms for its actions. Pyrazole-induced rat hepatocytes or HepG2 cells expressing CYP2E1 were treated with AA in the presence or absence of an inhibitor of nitric oxide synthase L-N(G)-Nitroarginine Methylester (L-NAME) or the NO donors S-nitroso-N-acetylpenicillamine (SNAP), and (Z)-1-[-(2-aminoethyl)-N-(2-aminoethyl)]diazen-1-ium-1,2-diolate (DETA-NONO). AA decreased cell viability from 100% to 48+/-6% after treatment for 48 h. In the presence of L-NAME, viability was further lowered to 23+/-5%, while, SNAP or DETA-NONO increased viability to 66+/-8 or 71+/-6%. The L-NAME potentiated toxicity was primarily necrotic in nature. L-NAME did not affect CYP2E1 activity or CYP2E1 content. SNAP significantly lowered CYP2E1 activity but not protein. AA treatment increased lipid peroxidation and lowered GSH levels. L-NAME potentiated while SNAP prevented these changes. Thus, L-NAME increased, while NO donors decreased AA-induced oxidative stress. Antioxidants prevented the L-NAME potentiation of AA toxicity. Damage to mitochondria by AA was shown by a decline in the mitochondrial membrane potential (MMP). L-NAME potentiated this decline in MMP in association with its increase in AA-induced oxidative stress and toxicity. NO donors decreased this decline in MMP in association with their decrease in AA-induced oxidative stress and toxicity. These results indicate that NO can be hepatoprotective against CYP2E1-dependent toxicity, preventing AA-induced oxidative stress.
Wu
Defeng
D
Department of Pharmacology and Biological Chemistry, Box 1603, One Gustave L. Levy Place, Mount Sinai School of Medicine, New York, NY 10029, USA.
Cederbaum
Arthur
A
D000818:Animals; D000975:Antioxidants / Q000494:pharmacology; D016718:Arachidonic Acid / Q000633:toxicity*; D006528:Carcinoma, Hepatocellular / Q000188:drug therapy / Q000201:enzymology / Q000473:pathology; D002470:Cell Survival / Q000187:drug effects; D019392:Cytochrome P-450 CYP2E1 / Q000378:metabolism*; D004338:Drug Combinations; D004347:Drug Interactions; D004791:Enzyme Inhibitors / Q000633:toxicity*; D005978:Glutathione / Q000378:metabolism; D022781:Hepatocytes / Q000187:drug effects / Q000201:enzymology / Q000473:pathology; D006801:Humans; D007425:Intracellular Membranes / Q000187:drug effects / Q000502:physiology; D015227:Lipid Peroxidation / Q000187:drug effects; D008113:Liver Neoplasms / Q000188:drug therapy / Q000201:enzymology / Q000473:pathology; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D008928:Mitochondria / Q000187:drug effects; D019331:NG-Nitroarginine Methyl Ester / Q000633:toxicity*; D009336:Necrosis / Q000139:chemically induced / Q000473:pathology; D009569:Nitric Oxide / Q000378:metabolism*; D020030:Nitric Oxide Donors / Q000494:pharmacology*; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors; D009603:Nitroso Compounds / Q000494:pharmacology; D018384:Oxidative Stress / Q000187:drug effects; D051381:Rats; D017207:Rats, Sprague-Dawley; D026423:S-Nitroso-N-Acetylpenicillamine / Q000494:pharmacology; D014162:Transfection
D000975:Antioxidants; D004338:Drug Combinations; D004791:Enzyme Inhibitors; D020030:Nitric Oxide Donors; D009603:Nitroso Compounds; C094210:2,2'-(hydroxynitrosohydrazono)bis-ethanamine; D016718:Arachidonic Acid; D009569:Nitric Oxide; D026423:S-Nitroso-N-Acetylpenicillamine; D019392:Cytochrome P-450 CYP2E1; D019001:Nitric Oxide Synthase; D005978:Glutathione; D019331:NG-Nitroarginine Methyl Ester