Increase of mtNOS contributes to oxidative and nitrative stress in aging.

CONVERSO DP, LEVISMAN D, CARRERAS MC, PODEROSO JJ.

Washington, EEUU

Congreso; 14th Annual Meeting of the Society for Free Radical Biology and Medicine.; 2007

Society for Free Radical Biology and Medicine

Over the past few decades, the role of oxidative stress in the process of aging was extensively discussed, being established that an increase in free radical production plus low antioxidant defenses are responsible for this phenomenon. In addition, it is known that nitric oxide (NO) elicits the mitochondrial production of superoxide anion (O2-) and hydrogen peroxide (H2O2). In this context, we wondered whether mitochondrial NO increases during aging, how this increase would be regulated, and a putative action on mitochondrial function.  Liver (L) and brain (B) Wistar rats of 3 (C) to 24 (O) months were utilized. The activity of mitochondrial nitric oxide synthase (mtNOS) increased up to 100% at  24 months (LC:  24±3  vs  LO: 44±6; BC: 17±2  vs BO: 36±3 pmoles NO/min.mg prot, p<0.05), which was accompanied by an increment in the enzyme expression.  This observation went in parallel to the low expression level of heme oxigenase-1 (HO-1) at 24 months, enzyme that negatively regulates mtNOS. In this context, mitochondrial functionality was evaluated by measuring spectrophotometrically the activity of respiratory chain complexes by either reduction or oxidation of citochrome c. We observed 40% decay of electron transfer rate at complex I-III (LC: 341±48 vs LO:  203±19 nmoles NADH/min.mg prot, p<0.05). Mitochondrial superoxide dismutase (MnSOD)  activity was  diminished  at  24  months  as  well  (LC:  1394±95  vs  LO: 921±11;  BC: 3350±150  vs BO:  2398±60 pmoles SOD/mg  prot, p<0.05). These two effects contributed here to increase O2- that also reacts with NO to produce peroxynitrite (ONOO-) that is capable of nitrating proteins.  In accord, western blot revealed a rise over basal nitration of mitochondrial proteins in aged animals, being particularly identified complex I and Mn-SOD nitration by coimmunoprecipitation.  These results lead us to conclude that in the aging process there is an increase in the production of matrix NO and O2-  that cannot be reverted by low antioxidant defenses and that, augmented  formation  of highly reactive ONOO- modifies mitochondrial proteins and causes the loss of complex I activity and the progression of oxidative stress during lifespan.