Apoptosis In Neurodegenerative Disorders

神经退行性疾病中的细胞凋亡

• 批准号: 8736518
• 负责人: Mark Mattson
• 金额: $ 50.82万
• 依托单位: NATIONAL INSTITUTE ON AGING
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8736518
• 关键词: ADP ribosylation; Aging; Alzheimer' s Disease; Animal Model; Antioxidants; Anxiety; Apoptosis; Attenuated; Base Excision Repairs; Biological; Brain; Brain Injuries; Brain-Derived Neurotrophic Factor; Calcium; Caloric Restriction; Calpain; Cell Culture Techniques; Cell Death; Cell Nucleus; Cell Survival; Cell membrane; Cell model; Cells; Cellular Stress; Cellular Stress Response; Complex; Corpus striatum structure; Cytoplasm; DNA; DNA Damage; DNA Repair; DNA repair protein; Deacetylase; Deacetylation; Development; Disease Progression; Disease model; Dopamine; Enzymes; Event; Excision; Exhibits; Experimental Models; Exposure to; Genes; Glutamate Receptor; Glutamates; Hippocampus (Brain); Homeostasis; Human; Huntington Disease; Imaging technology; Injury; Intravenous Immunoglobulins; Investigation; Ipsilateral; Ischemia; Laboratories; Learning; Ligase; Lipid Peroxidation; Mediating; Mediator of activation protein; Memory; Messenger RNA; Metabolic; Metabolism; Mitochondria; Modeling; Molecular; Molecular Biology; Mono-S; Motor; Movement; Mus; Muscle Cells; Naphthoquinones; Nerve Degeneration; Neurodegenerative Disorders; Neurons; Neurosciences; Neurotrophic Tyrosine Kinase Receptor Type 2; Nuclear; Nucleotides; Nutrient; Oxidation-Reduction; Oxidative Stress; Parkinson Disease; Pathogenesis; Pathway interactions; Peptide Elongation Factor 2; Performance; Phosphorylation; Phytochemical; Play; Post-Translational Protein Processing; Predisposition; Process; Proliferating Cell Nuclear Antigen; Property; Proteins; Proteomics; RTH-1 Nuclease; Rattus; Reperfusion Therapy; Reporting; Response Elements; Ribosomes; Role; Side; Signal Transduction; Stem cells; Stroke; Surgical incisions; TLR4 gene; Telomerase; Telomeric Repeat Binding Protein 2; Testing; Toll-Like Receptor 2; Toll-like receptors; Toxic effect; Translations; Undifferentiated; Up-Regulation; Water; cerebral atrophy; conditioned fear; cumene hydroperoxide; excitotoxicity; experience; functional outcomes; gamma secretase; human APEX1 protein; human FOXO3A protein; memory retention; mitochondrial uncoupling protein; motor function improvement; mouse model; mutant; nerve stem cell; nervous system disorder; neuroblastoma cell; neuron apoptosis; neuroprotection; novel; novel therapeutic intervention; overexpression; oxidative damage; plumbagin; preclinical study; prevent; repaired; telomere

In neurodegenerative disorders such as Alzheimers, Parkinsons and Huntingtons diseases, neurons may die by a form of programmed cell death called apoptosis. A major effort in the Cellular and Molecular Neurosciences section of the Laboratory of Neurosciences is aimed at establishing what triggers apoptosis in neurodegenerative disorders and how neuronal degeneration might be prevented by targeting specific molecular events in the process of apoptosis. Studies of cultured neurons demonstrated that activation of glutamate receptors can induce a transient damage to DNA which is rapidly repaired as the result of calcium-mediated upregulation of the DNA repair protein APE1. In addition, we have identified a mitochondrial uncoupling protein (UCP4) that can protect neurons in models relevant to stroke and Alzheimers disease by a mechanism involving suppression of oxidative stress and stabilization of cellular calcium homeostasis. We have also established roles for brain-derived neurotrophic factor (BDNF) in preventing the apoptosis of neurons produced from stem cells in the hippocampus, a finding that suggests the possibility of increasing the capacity of the brain to replace lost and damaged neurons. In other studies we have found that newly generated neurons are highly sensitive to DNA damage-induced apoptosis because they have low levels of telomerase and the telomere-associated protein TRF2. Preclinical studies have shown that intravenous immunoglobulin and gamma-secretase inhibotors are effective in stroke models. More recently, we have shown that neurons express several toll-like receptors, and have provided evidence that activation of two of these receptors (TLR2 and TLR4) can trigger apoptosis in cell culture and animal models of Alzheimer's disease and stroke. In other studies we have revealed important roles for plasma membrane redox enzymes in protecting neurons against apoptosis in experimental models of aging and neurodegenerative disorders. We previously showed that calorie restriction ameliorated Huntington's disease (HD) pathogenesis and slowed disease progression in mice that model Huntington's disease (Huntington's disease mice). We now report that overexpression of Sirt1, a mediator of the beneficial metabolic effects of calorie restriction, protects neurons against mutant HTT toxicity, whereas reduction of Sirt1 exacerbates mutant HTT toxicity. Overexpression of Sirt1 improves motor function, reduces brain atrophy and attenuates mutant-HTT-mediated metabolic abnormalities in Huntington's disease mice. Further mechanistic studies suggested that Sirt1 prevents the mutant-HTT-induced decline in BDNF concentrations and the signaling of its receptor, TrkB, and restores dopamine concentrations in the striatum. Sirt1 deacetylase activity is required for Sirt1-mediated neuroprotection in Huntington's disease cell models. Notably, we show that mutant HTT interacts with Sirt1 and inhibits Sirt1 deacetylase activity, which results in hyperacetylation of Sirt1 substrates such as forkhead box O3A (Foxo3a), thereby inhibiting its pro-survival function. Overexpression of Sirt1 counteracts the mutant-HTT-induced deacetylase deficit, enhances the deacetylation of Foxo3a and facilitates cell survival. These findings show a neuroprotective role for Sirt1 in mammalian HD models and open new avenues for the development of neuroprotective strategies in HD. Eukaryotic elongation factor 2 (eEF-2) is an important regulator of the protein translation machinery whereby it controls the movement of the ribosome along the mRNA. The activity of eEF-2 is regulated by changes in cellular energy status and nutrient availability and by posttranslational modifications such as phosphorylation and mono-ADP-ribosylation. However, the mechanisms regulating protein translation under conditions of cellular stress in neurons are unknown. Here we show that when rat hippocampal neurons experience oxidative stress (lipid peroxidation induced by exposure to cumene hydroperoxide; CH), eEF-2 is hyperphosphorylated and ribosylated, resulting in reduced translational activity. The degradation of eEF-2 requires calpain proteolytic activity and is accompanied by accumulation of eEF-2 in the nuclear compartment. The subcellular localization of both native and phosphorylated forms of eEF-2 is influenced by CRM1 and 14.3.3, respectively. In hippocampal neurons p53 interacts with nonphosphorylated (active) eEF-2, but not with its phosphorylated form. The p53-eEF-2 complexes are present in cytoplasm and nucleus, and their abundance increases when neurons experience oxidative stress. The nuclear localization of active eEF-2 depends upon its interaction with p53, as cells lacking p53 contain less active eEF-2 in the nuclear compartment. Overexpression of eEF-2 in hippocampal neurons results in increased nuclear levels of eEF-2 and decreased cell death after exposure to CH. Our results reveal novel molecular mechanisms controlling the differential subcellular localization and activity state of eEF-2 that may influence the survival status of neurons during periods of elevated oxidative stress Nuclear factor E2-related factor 2 (Nrf2)/antioxidant response element (ARE) pathway is an important cellular stress response pathway involved in neuroprotection. We previously screened several natural phytochemicals and identified plumbagin as a novel activator of the Nrf2/ARE pathway that can protect neurons against ischemic injury. Here we extended our studies to natural and synthetic derivatives of plumbagin. We found that 5,8-dimethoxy-1,4-naphthoquinone (naphthazarin) is a potent activator of the Nrf2/ARE pathway, up-regulates the expression of Nrf2-driven genes in primary neuronal and glial cultures, and protects neurons against glutamate-induced excitotoxicity. Neurons are terminally differentiated cells with a high rate of metabolism and multiple biological properties distinct from their undifferentiated precursors. Previous studies showed that nucleotide excision DNA repair is downregulated in postmitotic muscle cells and neurons. Here, we characterize DNA damage susceptibility and base excision DNA repair (BER) capacity in undifferentiated and differentiated human neural cells. The results show that undifferentiated human SH-SY5Y neuroblastoma cells are less sensitive to oxidative damage than their differentiated counterparts, in part because they have robust BER capacity, which is heavily attenuated in postmitotic neurons. The reduction in BER activity in differentiated cells correlates with diminished protein levels of key long patch BER components, flap endonuclease-1, proliferating cell nuclear antigen, and ligase I. Thus, because of their higher BER capacity, proliferative neural progenitor cells are more efficient at repairing DNA damage compared with their neuronally differentiated progeny. In a related study we found that mice lacking NEIL1 exhibit impaired memory retention in a water maze test, but no abnormalities in tests of motor performance, anxiety, or fear conditioning. NEIL1 deficiency results in increased brain damage and a defective functional outcome in a focal ischemia/reperfusion model of stroke. The incision capacity on a 5-hydroxyuracil-containing bubble substrate was lower in the ipsilateral side of ischemic brains and in the mitochondrial lysates of unstressed old NEIL1-deficient mice. These results indicate that NEIL1 plays an important role in learning and memory and in protection of neurons against ischemic injury.

在神经退行性疾病中，例如阿尔茨海默氏症，帕金森氏症和亨廷顿疾病，神经元可能会因一种称为凋亡的程序性细胞死亡而死亡。 神经科学实验室的细胞和分子神经科学部分的主要努力旨在确定哪些触发神经退行性疾病中的凋亡以及如何通过在凋亡过程中靶向特定的分子事件来预防神经元变性。 培养神经元的研究表明，谷氨酸受体的激活可以诱导对DNA的短暂损害，而DNA迅速修复了DNA修复蛋白APE1的上调。 此外，我们已经确定了一种线粒体解偶联蛋白（UCP4），该蛋白可以通过涉及抑制氧化应激和稳定细胞钙稳态的机制来保护与中风和阿尔茨海默氏病有关的模型中的神经元。 我们还确定了脑衍生的神经营养因子（BDNF）在防止海马中由干细胞产生的神经元的凋亡中的作用，这一发现表明有可能增加大脑替代损失和受损神经元的能力。 在其他研究中，我们发现新近产生的神经元对DNA损伤诱导的凋亡高度敏感，因为它们的端粒酶水平较低和与端粒相关的蛋白TRF2。临床前研究表明，静脉免疫球蛋白和γ-分泌酶抑制剂在中风模型中有效。 最近，我们表明神经元表达了几种类似Toll的受体，并提供了证据表明，这些受体中的两个（TLR2和TLR4）的激活可以触发阿尔茨海默氏病和中风的细胞培养和动物模型中的细胞凋亡。 在其他研究中，我们揭示了质膜氧化还原酶在衰老和神经退行性疾病的实验模型中保护神经元免受凋亡的重要作用。 我们先前表明，卡路里限制改善了亨廷顿氏病（HD）的发病机理，并减慢了模拟亨廷顿氏病（亨廷顿病小鼠）的小鼠的疾病进展。现在，我们报告说，SIRT1的过表达是卡路里限制的有益代谢作用的介体，可保护神经元免受突变htt毒性的侵害，而SIRT1的降低加剧了突变体HTT毒性。 SIRT1的过表达可改善运动功能，减少脑萎缩，并减轻亨廷顿病小鼠中突变体HTT介导的代谢异常。进一步的机械研究表明，SIRT1可防止突变体HTT诱导的BDNF浓度下降及其受体，TRKB的信号传导，并恢复纹状体中的多巴胺浓度。 SIRT1脱乙酰基酶活性是SIRT1介导的亨廷顿病细胞模型中的神经保护需要的。值得注意的是，我们表明突变型HTT与SIRT1相互作用并抑制SIRT1脱乙酰基酶活性，从而导致SIRT1底物（例如Forkhead Box O3A（FoxO3A））的过度乙酰化，从而抑制其促卵巢功能。 SIRT1的过表达抵消了突变-HTT诱导的脱乙酰基酶缺损，增强了FOXO3A的脱乙酰基化并促进细胞存活。 这些发现显示了SIRT1在哺乳动物HD模型中的神经保护作用，并开放了新的HD神经保护策略的新途径。 真核伸长因子2（EEF-2）是蛋白质翻译机制的重要调节剂，它可以控制核糖体沿mRNA的运动。 EEF-2的活性受细胞能量状态和养分可用性的变化以及翻译后修饰（例如磷酸化和单ADP-核糖基化）的调节。然而，在神经元细胞应激条件下调节蛋白质翻译的机制尚不清楚。在这里，我们表明，当大鼠海马神经元经历氧化应激（暴露于Cumene hydrocoxide; CH）时，EEF-2被过度磷酸化并核糖基化，从而减少了翻译活性。 EEF-2的降解需要钙蛋白酶蛋白水解活性，并伴随着EEF-2在核室中的积累。 EEF-2的天然和磷酸化形式的亚细胞定位分别受CRM1和14.3.3的影响。在海马神经元中，p53与非磷酸化（活性）EEF-2相互作用，但与其磷酸化形式不相互作用。 p53-EEF-2复合物存在于细胞质和核中，当神经元经历氧化应激时，其丰度会增加。主动EEF-2的核定位取决于其与p53的相互作用，因为缺乏p53的细胞在核室中含有较少的活性EEF-2。海马神经元中EEF-2的过表达导致EEF-2的核水平升高，暴露于CH后的细胞死亡降低。我们的结果揭示了控制氧化应激升高期间神经元可能影响神经元生存状态的差异亚细胞定位和活性状态的新型分子机制 核因子E2相关因子2（NRF2）/抗氧化剂响应元件（AS）途径是参与神经保护的重要细胞应力反应途径。我们先前筛选了几种天然植物化学物质，并确定了铅铅制是NRF2/是可以保护神经元免受缺血性损伤的途径的新型激活剂。在这里，我们将研究扩展到了铅垂的天然和合成衍生物。我们发现5,8-二甲氧基-1,4-萘醌（萘氮蛋白）是NRF2/是途径的有效激活剂，在原发性神经元和神经胶质培养物中上调了NRF2驱动的基因的表达，可在神经元中保护神经元免受粘液氨酸酯诱导的兴奋性兴奋性。 神经元是最终分化的细胞，具有高代谢率和与未分化的前体不同的多种生物学特性。先前的研究表明，核苷酸切除DNA修复在有丝分裂后肌肉细胞和神经元中下调。在这里，我们表征了未分化和分化的人类神经细胞中DNA损伤敏感性和碱基切除DNA修复（BER）的能力。结果表明，与分化的对应物相比，未分化的人类SY5Y神经母细胞瘤细胞对氧化损伤的敏感性不大，部分原因是它们具有强大的BER能力，这在丝状神经后神经元中大量减弱。分化细胞中BER活性的降低与蛋白质水平降低的关键长斑块成分，皮瓣内核酸酶-1，增殖细胞核抗原和连接酶I的降低相关。因此，由于其较高的BER能力，增殖性神经祖细胞的增殖性神经祖细胞在修复DNA损伤方面具有更高的效率，而与其神经元分化相比。在一项相关研究中，我们发现缺乏Neil1的小鼠在水迷宫测试中表现出记忆力保留受损，但在运动性能，焦虑或恐惧调节的测试中没有异常。 NEIL1缺乏会导致脑部缺血/再灌注模型的脑损伤增加和功能结果有缺陷。在缺血性大脑的同侧和无数旧尼尔1缺陷型小鼠的线粒体裂解物中，含5-羟基尿素的起泡泡沫底物的切口能力较低。这些结果表明，Neil1在学习和记忆以及保护神经元免受缺血性损伤方面起着重要作用。