It has been proposed that brain disorders such as AIDS dementia complex, Alzheimer's dementia, Parkinson's disease, stroke, head trauma, and possibly schizophrenia may be caused by progressive neurodegeneration. Oxidative brain damage occurs when the endogenous antioxidative defense systems are severely weakened by aging, ischemia, viral infection, and neurotoxins. The mission of our research unit is to retard progressive brain damage and to improve life quality by developing neuroprotective agents and strategies against neurodegeneration caused by free radicals. We employed both cell cultures and animal models for investigating neuroprotective and/or neurorescue strategies including antioxidant therapy, hypothermia therapy, and gene induction/therapy. Preconditioning stress induced by a transient ischemia has been shown to increase brain tolerance to oxidative stress and the underlying neuroprotective mechanisms are not well understood. We first investigated the pathophysiology of hydroxyl radicals (OH) and nitric oxide (NO) free radicals in apoptosis caused by serum deprivation in SH-SY5Y cells derived from human brain. We have also developed a preconditioning cell model for inducing stress-responsive genes such as redox factor-1 (Ref-1), Fos, and neuronal nitric oxide synthase (NOS1). We propose studying the beneficial effects of NO and cGMP in gene induction, cell viability, and adaptive tolerance to lethal oxidative stress. In a series of experiments, we found that endogenous NO, S-nitrosoglutathione (GSNO) and N-acetylcysteine blocked oxidative stress-induced apoptosis in human brain cells. Similar to nuclear redox factor-1 (Ref-1), mRNA of human neuronal nitric oxide synthase (NOS1) was maximally up regulated within 2 hours after oxidative stress and down regulated by NO/GSNO and OH scavenger (i.e., serum and salicylate). A brief preconditioning stress induced by serum deprivation for 2 hours caused a delayed increase in the expression of human NOS1 mRNA, protein and the associated formation of NO and cGMP, which in turn decreased OH generation and oxidative stress-induced cell death. In addition to inhibiting caspase-3 through a dithiothreitol-sensitive S-nitrosylation process, preconditioning stress concomitantly up-regulated the expression of the anti-apoptotic bcl-2 protein and down-regulated the p66shc adaptor protein. This beneficial neuroprotective process of preconditioning stress is mediated by newly synthesized NO since it can be suppressed by the inhibition of NOS1 and guanylyl cyclase. Therefore, the constitutive isoform of human NOS1 is dynamically redox-regulated to meet both functional and compensatory demands of NO for gene regulation, antioxidant defense, and tolerance to oxidative stress.
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