title: NADH-dependent microsomal interaction with ferric complexes and production of reactive oxygen intermediates.
abstract: The interaction of NADPH with ferric complexes to catalyze microsomal generation of reactive oxygen intermediates has been well studied. Experiments were carried out to characterize the ability of NADH to interact with various ferric chelates to promote microsomal lipid peroxidation and generation of .OH-like species. In the presence of NADH and iron, microsomes produced .OH as assessed by the oxidation of a variety of .OH scavenging agents. Rates of NADH-dependent .OH production were 50 to 80% those of the NADPH-catalyzed reaction. The oxidation of dimethyl sulfoxide or t-butyl alcohol was inhibited by catalase and competitive .OH scavengers but not by superoxide dismutase or carbon monoxide. NADH-dependent .OH production was effectively catalyzed by ferric-EDTA and ferric-diethylenetriaminepentaacetic acid (DTPA), whereas ferric-ATP and ferric-citrate were poor catalysts. All these ferric chelates were reduced by microsomes in the presence of NADH (and NADPH). H2O2 was produced in the presence of NADH in a reaction stimulated by the addition of ferric-EDTA, consistent with the increase in .OH production. The latter appeared to be limited by the rate of H2O2 generation rather than the rate of reduction of the ferric chelate. NADH-dependent lipid peroxidation was much lower than the NADPH-catalyzed reaction and showed an opposite response to catalysis by ferric complexes compared to .OH generation as production of thiobarbituric acid-reactive material was increased with ferric-ATP and -citrate, but not with ferric-EDTA or- DTPA, and was not affected by catalase, SOD, or .OH scavengers. These results indicate that NADH can support microsomal reduction of ferric chelates, with the subsequent production of .OH-like species and peroxidation of lipids. The pattern of response of the NADH-dependent reactions with respect to catalytic effectiveness of ferric chelates and sensitivity to radical scavengers is similar to that found with NADPH. Many of the metabolic actions of ethanol have been ascribed to production of NADH as a consequence of oxidation by alcohol dehydrogenase. Since the cytosol normally maintains a highly oxidized NAD+/NADH redox ratio, it is interesting to speculate that increased availability of NADH from the oxidation of ethanol may support microsomal reduction of iron complexes, with the subsequent generation of reactive oxygen intermediates.
authors.affiliation: Department of Biochemistry, Mount Sinai School of Medicine, New York, New York 10029.
authors.forename: E
authors.initials: E
authors.lastname: Kukiełka
authors.forename: A I
authors.initials: AI
authors.lastname: Cederbaum
mesh_terms: D000818:Animals; D000440:Butanols / Q000378:metabolism; D004121:Dimethyl Sulfoxide / Q000378:metabolism; D005290:Ferric Compounds / Q000378:metabolism*; D005557:Formaldehyde / Q000378:metabolism; D005609:Free Radicals; D006878:Hydroxides / Q000378:metabolism*; D017665:Hydroxyl Radical; D007700:Kinetics; D008297:Male; D008862:Microsomes, Liver / Q000378:metabolism*; D009243:NAD / Q000378:metabolism*; D010084:Oxidation-Reduction; D051381:Rats; D011919:Rats, Inbred Strains; D020002:tert-Butyl Alcohol
chemical_list: D000440:Butanols; D005290:Ferric Compounds; D005609:Free Radicals; D006878:Hydroxides; D009243:NAD; D005557:Formaldehyde; D017665:Hydroxyl Radical; D020002:tert-Butyl Alcohol; D004121:Dimethyl Sulfoxide