Peroxynitrite, nitrotyrosine and HSP90 in neuronal death

我们的长期目标是了解SOD突变如何增加氧化应激， 导致肌萎缩侧索硬化症（ALS）运动神经元死亡。我们已经证明 通过超氧化物和 一氧化氮诱导培养的胚胎大鼠运动神经元凋亡的营养支持剥夺。 一氧化氮合成抑制剂和铜锌超氧化物歧化酶（SOD）都能 细胞内脂质体保护运动神经元免于凋亡。这些数据表明 一氧化氮和超氧化物之间相互作用在运动神经元凋亡中起作用。突变以 SOD参与ALS运动神经元的选择性变性和ALS-1的表达。 转基因小鼠中的SOD突变体产生运动神经元疾病。一种常见的表型， 迄今为止研究的ALS-SOD突变是降低对锌的亲和力。我们已经证明 缺锌SOD清除超氧阴离子的能力较弱，但对酪氨酸的催化能力较强 硝化作用。此外，缺锌SOD中的铜可作为非特异性单电子 氧化酶，从抗坏血酸和谷胱甘肽等抗氧化剂中夺取可以转移的电子 转化成超氧化物在NO存在下，缺锌SOD可催化 过氧亚硝酸盐的形成。在上一轮的资助中，我们已经证明，缺锌SOD 通过一氧化氮依赖性机制诱导运动神经元凋亡。为了更新，我们的 第一个目的是进一步研究缺锌SOD可以杀死培养的 运动神经元，并确定什么可以保护运动神经元免受这种毒性。我们的第二个目标 是表征营养因子在运动神经元中诱导的超氧化物的来源 戒断我们的第三个目的是测试过亚硝酸盐对酪氨酸硝化在运动神经死亡中的作用。 营养因子剥夺或锌缺乏SOD诱导的神经元。完成 具体的目标将提供一个机制的基础，解释如何运动神经元，特别是 易受SOD突变的影响，并在散发性和家族性SOD之间建立联系。

Our long-term goal is to understand how mutations to SOD can increase oxidative stress and cause the death of motor neurons in amyotrophic lateral sclerosis (ALS). We have shown that endogenous formation of peroxynitrite by the diffusion-limited reaction between superoxide and nitric oxide induces apoptosis in cultured embryonic rat motor neurons deprived of trophic support. Both inhibitors of nitric oxide synthesis as well as Cu.Zn superoxide dismutase (SOD) delivered intracellularly with liposomes protect motor neurons from apoptosis. These data indicate that the interaction between nitric oxide and superoxide has a role in motor neuron apoptosis. Mutations to SOD are implicated in the selective degeneration of motor neurons in ALS and expression of ALS- SOD mutants in transgenic mice produces motor neuron disease. A common phenotype among the ALS-SOD mutations so far investigated is to decrease the affinity for zinc. We have shown that zinc-deficient SOD is both less efficient at scavenging superoxide and a better catalyst of tyrosine nitration. Furthermore, the copper in zinc-deficient SOD can act as a non-specific one-electron oxidase, robbing electrons from antioxidants like ascorbate and glutathione that can be transferred to oxygen to produce superoxide. In the presence of NO, zinc-deficient SOD can catalyze the formation of peroxynitrite. In the previous cycle of funding, we have shown that zinc-deficient SOD induces apoptosis in motor neurons by a nitric oxide-dependent mechanism. For the renewal, our first aim is to further investigate the mechanisms by which zinc-deficient SODs can kill cultured motor neurons and to determine what can protect motor neurons from this toxicity. Our second aim is to characterize the source or sources of superoxide induced in motor neurons by trophic factor withdrawal. Our third aim is to test the role of tyrosine nitration by peroxynitrite in the death of motor neurons induced by either trophic factor deprivation or by zinc-deficient SOD. Completion of the specific aims will provide a mechanistic basis for explaining how motor neurons are particularly vulnerable to SOD mutations and establish a link between sporadic and familial SODs.