Rat liver microsomal NADPH-supported oxidase activity and lipid peroxidation dependent on ethanol-inducible cytochrome P-450 (P-450IIE1).
The liver microsomal ethanol-inducible cytochrome P-450 (P-450IIE1) form is known to exhibit a high rate of oxidase activity in the absence of substrate and it was therefore of interest to evaluate whether this form of P-450 could contribute to microsomal and liposomal NADPH-dependent oxidase activity and lipid peroxidation. The rate of microsomal NADPH-consumption, O2--formation, H2O2-production and generation of thiobarbituric acid (TBA) reactive substances correlated to the amount of P-450IIE1 in 28 microsomal samples from variously treated rats. Anti-P-450IIE1 IgG inhibited, compared to control IgG, microsomal H2O2-formation by 45% in microsomes from acetone-treated rats and by 22% in control microsomes. NADPH-dependent generation of TBA-reactive products was completely inhibited by these antibodies, whereas preimmune IgG was essentially without effect. Liposomes containing reductase and P-450IIE1 were peroxidized in a superoxide dismutase (SOD) sensitive reaction at a 5-10-fold higher rate than membranes containing 3 other forms of cytochrome P-450. Lipid peroxidation in reconstituted vesicles dependent on the presence of P-450IIB1 was by contrast not inhibited by SOD. Microsomal peroxidase activities, using 15-(S)-hydroperoxy-5-cis-8,11,13-trans-eicosatetraenoic acid as a substrate were high in microsomes from phenobarbital- or ethanol-treated rats but low in membranes from isoniazid-treated rats, having the highest relative level of P-450IIE1. It is suggested that the oxidase activity of P-450IIE1 contributes to microsomal NADPH-dependent lipid peroxidation. The combined action of the oxidase activity by P-450IIE1 and the peroxidase activities by P-450IIB1 and other forms of P-450 may be important for the high rate of lipid peroxidation observed in e.g. microsomes from ethanol- or acetone-treated rats. The possible importance of cytochrome P-450IIE1-dependent lipid peroxidation in vivo after ethanol abuse is discussed.
Ekström
G
G
Department of Physiological Chemistry, Karolinska Institute, Stockholm, Sweden.
Ingelman-Sundberg
M
M
D000818:Animals; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis / Q000378:metabolism*; D004492:Edetic Acid / Q000494:pharmacology; D004790:Enzyme Induction; D000431:Ethanol / Q000494:pharmacology*; D005609:Free Radicals; D007074:Immunoglobulin G / Q000494:pharmacology; D015227:Lipid Peroxidation; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology*; D009249:NADP / Q000378:metabolism*; D009251:NADPH-Ferrihemoprotein Reductase; D010088:Oxidoreductases / Q000378:metabolism*; D010100:Oxygen / Q000378:metabolism; D010544:Peroxidases / Q000378:metabolism; D051381:Rats; D011919:Rats, Inbred Strains
D005609:Free Radicals; D007074:Immunoglobulin G; D000431:Ethanol; D009249:NADP; D003577:Cytochrome P-450 Enzyme System; D004492:Edetic Acid; D010088:Oxidoreductases; D010544:Peroxidases; D009251:NADPH-Ferrihemoprotein Reductase; D010100:Oxygen