NEUROPROTECTIVE SIGNAL TRANSDUCTION AND ALZHEIMER'S DISEASE

这一建议检验了肿瘤坏死因子(TNF)和 分泌型β-淀粉样前体蛋白对神经元的保护作用 抗淀粉样β-肽(Abeta)毒性的机制 转录因子NFkappaB与抗氧化酶的诱导。 活性氧自由基(ROS)介导Abeta的神经毒性 和钙，初步数据显示，TNFS和SAPS可以保护 海马神经元抗Aβ毒性和诱导NFkappaB DNA 结合活性。该项目的具体目标是：1)测试 肿瘤坏死因子和碱性磷酸酶保护培养的海马神经元的假说 通过诱导NFkappaB的激活来对抗Aβ毒性。我们会 肿瘤坏死因子及其受体诱导NFkappaB DNA结合活性的研究 SAPS，确定其他已知的激活NFkappaB的代理是否 神经保护，并使用反义寡核苷酸(AODN)来对抗IkappaB (NFkappaB抑制亚基)或p50(转录因子亚基)。 2)检验TNF和SAPP激活NFkappaB的假设 防止Abeta诱导的ROS积累，离子动力障碍 ATPase活性和[Ca~(2+)]i.特定ROS的作用 用过氧化氢、羟基自由基、超氧化物和 蛋白质羰基。3)检验以下假设： NFkappaB信号系统对神经细胞的保护作用 抗氧化酶和钙结合蛋白的表达 钙结合蛋白。这将通过抗氧化酶活性来实现。 核糖核酸酶保护实验和Western印迹分析。4)至 确定脑室注射肿瘤坏死因子和碱性磷酸酶是否 成年大鼠保护海马区突触体免受Aβ诱导的损伤。 NFkappaB活性、抗氧化酶活性和钙结合蛋白水平 在输注肿瘤坏死因子和SAPP后，将进行量化和关联 以及肿瘤坏死因子和碱性磷酸酶对突触体对Abeta易感性的影响。 5)检验NFkappaB信号系统被激活的假设 在AD大脑中存在与选择性易损性相关的区域模式。 NFkappaB活性将在脆弱的脑组织中进行量化 区域(中回、海马体、顶下皮质) 和相对不脆弱的大脑区域(小脑、枕极) 来自阿尔茨海默病患者的大脑和年龄匹配的“对照”大脑。这项研究将 识别神经元损伤和神经保护的基本机制 与阿尔茨海默病的发病机制有关。

This proposal tests the hypothesis that tumor necrosis factors (TNFs) and secreted forms of beta-amyloid precursor protein (sAPPs) protect neurons against amyloid beta-peptide (Abeta) toxicity by a mechanism involving the transcription factor NFkappaB and induction of antioxidant enzymes. The neurotoxicity of Abeta is mediated by reactive oxygen species (ROS) and calcium, and preliminary data showed that TNFs and sAPPs can protect hippocampal neurons against Abeta toxicity and induce NFkappaB DNA binding activity. The specific aims of this project are: 1) To test the hypothesis that TNFs and sAPPs protect cultured hippocampal neurons against Abeta toxicity by inducing activation of NFkappaB. We will characterize induction of NFkappaB DNA binding activity by TNFs and sAPPs, determine whether other agents known to activate NFkappaB are neuroprotective, and employ antisense oligonucleotides (AODNs) to IkappaB (inhibitory subunit of NFkappaB) or p50 (transcription factor subunit). 2) To test the hypothesis that activation of NFkappaB by TNFs and sAPPs prevents Abeta-induced accumulation of ROS, impairment of ion-motive ATPase activities, and elevation of [Ca2+]i. Roles of specific ROS will be examined using assays of peroxides, hydroxyl radical, superoxide and protein carbonyl. 3) To test the hypothesis that activation of the NFkappaB signaling system by TNFs and sAPPs protects neurons by inducing the expression of antioxidant enzymes and the calcium-binding protein calbindin. This will be accomplished using antioxidant enzyme activity assays, RNAse protection assays and Western blot analysis. 4) To determine whether intraventricular administration of TNFs and sAPPs to adult rat protects hippocampal synaptosomes from Abeta-induced damage. NFkappaB activity, antioxidant enzyme activities and calbindin levels following infusion of TNF and sAPPs will be quantified and correlated with effects of TNFs and sAPPs on vulnerability of synaptosomes to Abeta. 5) To test the hypothesis that the NFkappaB signaling system is activated in AD brain in a regional pattern related to selective vulnerability. NFkappaB activity will be quantified in tissue from vulnerable brain regions (middle temporal gyrus, hippocampus, inferior parietal cortex) and relatively nonvulnerable brain regions (cerebellum, occipital pole) from AD brains and age-matched "control" brains. This research will identify fundamental mechanisms of neuronal injury and neuroprotection relevant to the pathogenesis of Alzheimer's disease.