INHIBITION OF FERRITIN-STIMULATED MICROSOMAL PRODUCTION OF REACTIVE OXYGEN INTERMEDIATES BY NITRIC-OXIDE

Experiments were carried out to evaluate the effect of nitric oxide ex posure on the ability of NADPH-dependent microsomal electron transfer to mobilize iron from ferritin, Such interactions could play a role in potential antioxidant actions of nitric oxide (NO), Preincubation of the microsomes from phenobarbital-treated rats with NO donors such as S-nitroso-D,L-N-acetyl penicillamine (SNAP), S-nitroso-L-glutathione, SIN-1, and DETANONOate followed by centrifugation, washing, and resusp ension of the microsomes resulted in a decrease in the ferritin-depend ent oxidation of 2',7'-dichlorofluorescein diacetate (DCFDA) or ferrit in-catalyzed chemiluminescence compared to microsomes pretreated with buffer, The ferritin-stimulated rate of oxidation of DCFDA or of chemi luminescence was completely restored if the microsomal preincubation w ith NO donors was performed in the presence of hemoglobin, In contrast to results with ferritin, ferric-stimulated oxidation of the dye was not affected by any of the tested NO donors. The microsomal oxidation of aminopyrine was inhibited after SNAP treatment, indicating that NO inhibited cytochrome P450 catalyzed activity, Inhibition of cytochrome P450 also resulted in an inhibition of microsomal production of super oxide. Similar results were obtained using microsomes from a cloned ce ll line which express the CYP2E1 isoform, Since superoxide is required for the mobilization of iron from ferritin by microsomes, inhibition of superoxide production as a consequence of NO interaction with cytoc hrome P450 is likely to be responsible for the prevention of ferritin- catalyzed formation of reactive oxygen species by NO donors, The resul ts suggest that NO could exhibit an antioxidant capacity through its a bility of decreasing the activity of iron-heme compounds, such as cyto chrome P450, preventing the release of catalytically active iron from ferritin, and thus decreasing the ability to generate oxygen free radi cals involved in cytotoxicity. (C) 1997 Academic Press.