NITRIC-OXIDE PREVENTS OXIDATIVE DAMAGE PRODUCED BY TERT-BUTYL HYDROPEROXIDE IN ERYTHROLEUKEMIA-CELLS VIA NITROSYLATION OF HEME AND NONHEME IRON - ELECTRON-PARAMAGNETIC-RESONANCE EVIDENCE

We studied protective effects of NO against tert-butyl-hydroperoxide ( t-BuOOH)-induced oxidations in a subline of human erythroleukemia K562 cells with different intracellular hemoglobin (Hb) concentrations. t- BuOOH-induced formation of oxoferryl-Hb-derived free radical species i n cells was demonstrated by low temperature EPR spectroscopy, Intensit y of the signals was proportional to Hb concentrations and was correla ted with cell viability, Peroxidation of phosphatidylcholine, phosphat idylethanolamine, phosphatidylserine, phosphatidylinositol, and cardio lipin metabolically labeled with oxidation-sensitive cis-parinaric aci d was induced by t-BuOOH. An NO donor, pyl)-N-(n-propyl)amino]-diazen- 1-ium-1,2-diolate], produced nonheme iron dinitrosyl complexes and hex a- and pentacoordinated Hb-nitrosyl complexes in the cells. Nitrosylat ion of non heme iron centers and Hb-heme protected against t-BuOOH-ind uced: (a) formation of oxoferryl-Hb-derived free radical species, (b) peroxidation of cis-parinaric acid-labeled phospholipids, and (c) cyto toxicity. Since NO did not inhibit peroxidation induced by an ate-init iator of peroxyl radicals, 2,2'-azobis(2,4-dimethylvaleronitrile), pro tective effects of NO were due to formation of iron-nitrosyl complexes whose redox interactions with t-BuOOH prevented generation of oxoferr yl-Hb-derived free radical species.