SIRT1 Limits Microglial Toxicity in Alzheimer's Disease

描述（由申请人提供）：小胶质细胞激活长期以来一直被认为与阿尔茨海默病（AD）的发病机制有关。除了对神经元和突触功能造成直接毒性作用外，β淀粉样蛋白（A¿）和/或tau蛋白的积累刺激小胶质细胞的激活和炎症细胞因子的表达，这可以诱导进一步的神经元损伤。阻断小胶质细胞激活的毒性通路可有效预防神经退行性变。然而，调节小胶质环的分子机制仍然难以捉摸。我们之前对原代皮层培养的研究表明NF-？小胶质细胞中的B活化在小胶质细胞介导的a¿毒性中起关键作用。抑制NF-？B通过SIRT1（组蛋白去乙酰化酶sirtuin家族的一员）在a¿处理的原代培养物中保护小胶质细胞免受毒性。在AD大脑中，SIRT1水平显著降低。A¿处理显著降低了培养小胶质细胞中SIRT1的表达。基于这些发现，我们假设SIRT1减少是导致AD小胶质毒性的关键事件，并且小胶质SIRT1通过抑制NF- 1来限制a -介导的神经元缺陷。B激活。为了验证这一假设，我们提出了三个具体目标。在Aim 1中，我们将在表达人类淀粉样蛋白前体蛋白（hAPP）的小鼠小胶质细胞中灭活SIRT1，并系统地检查小胶质细胞SIRT1灭活如何影响炎症反应和A¿相关的神经元/行为缺陷。在Aim 2中，为了确定小胶质细胞SIRT1是否通过抑制NF-？B活化，我们将确定是否本构活化标准NF-？hAPP小鼠小胶质细胞中的B信号以类似SIRT1缺失的方式加剧了缺陷。在补充实验中，我们将确定抑制NF-？B信号可通过注入强效NF- 1改善A -相关的神经元缺陷。脑内B抑制剂还是注射抑制NF-的病毒载体？B在小胶质细胞。在Aim 3中，为了确定小胶质细胞SIRT1抑制NF-？B，我们将系统地检查SIRT1是否抑制NF-？B通过去乙酰化RelA和/或减少髓细胞中RelA的磷酸化。使用染色质免疫沉淀（ChIP）分析，我们将确定SIRT1-是否诱导NF-？B活化涉及髓细胞组蛋白（H3K56Ac）的去乙酰化。本研究的完成将为神经退行性变中调节小胶质环的分子机制提供新的见解。这些研究也将为我们开发sirt1增强策略作为阿尔茨海默病的新治疗方法的长期目标奠定基础。

DESCRIPTION (provided by applicant): Microglial activation has long been proposed to contribute to the pathogenesis of Alzheimer's disease (AD). Besides causing direct toxic effects on neuronal and synaptic functions, accumulation of amyloid beta (A¿) and/or tau stimulates microglial activation and expression of inflammatory cytokines, which can induce further neuronal damage. Blocking the toxic pathway in microglial activation could effectively protect against neurodegeneration. However, the molecular mechanisms modulating the microglial loop remain elusive. Our previous studies in primary cortical cultures suggest that NF-?B activation in microglia plays a critical role in microglial-mediated A¿ toxicity. Inhibition of NF-?B by SIRT1, a member of the sirtuin family of histone deacetylases, protected against microglia toxicity in A¿-treated primary cultures. In AD brains, SIRT1 levels were markedly reduced. SIRT1 expression in cultured microglia was significantly diminished by A¿ treatment. Based on these findings, we hypothesize that SIRT1 reduction is a key event leading to microglial toxicity in AD and that microglial SIRT1 limits A¿-mediated neuronal deficits by suppressing NF-?B activation. To test this hypothesis, we propose three Specific Aims. In Aim 1, we will inactivate SIRT1 in microglia of mice expressing human amyloid precursor protein (hAPP) and systematically examine how microglial SIRT1 inactivation affects inflammatory responses and A¿-related neuronal/behavioral deficits. In Aim 2, to determine if microglial SIRT1 exerts neuroprotection by suppressing NF-?B activation, we will determine if constitutive activation of canonical NF-?B signaling in microglia of hAPP mice exacerbates the deficits in a manner similar to SIRT1 deletion. In complementary experiments, we will determine if inhibiting NF-?B signaling will ameliorate A¿-associated neuronal deficits by infusing a potent NF-?B inhibitor in the brain or injection of a viral vector that inhibits NF-?B in microglia. In Aim 3, to determine the mechanism by which microglial SIRT1 inhibits NF-?B, we will systematically examine if SIRT1 inhibits NF-?B by deacetylating RelA and/or by reducing RelA phosphorylation in myeloid cells. Using chromatin immunoprecipitation (ChIP) analyses, we will then determine if SIRT1- induced suppression of NF-?B activation involves deacetylation of histones (H3K56Ac) in myeloid cells. Completion of the proposed studies will provide new insight into the molecular mechanisms modulating the microglial loop in neurodegeneration. These studies will also lay the foundation for our long-term goal of developing SIRT1-enhancing strategies as a new therapeutic approach for AD. PUBLIC HEALTH RELEVANCE: This projects aims at investigating mechanisms regulating the proinflammatory responses and microglial toxicity in Alzheimer's disease. This study may provide new therapeutic avenue for treating this devastating disease.