SIRT1 Limits Microglial Toxicity in Alzheimer's Disease

描述（由申请人提供）：长期以来一直认为小胶质细胞活化有助于阿尔茨海默病（AD）的发病机制。除了对神经元和突触功能造成直接毒性作用之外，淀粉样蛋白β（A β）和/或tau的积累刺激小胶质细胞活化和炎性细胞因子的表达，这可以诱导进一步的神经元损伤。阻断小胶质细胞活化中的毒性通路可以有效地防止神经退行性变。然而，调节小胶质细胞环的分子机制仍然难以捉摸。我们先前在原代皮层培养中的研究表明，小胶质细胞中的NF-κB活化在小胶质细胞介导的ApoA毒性中起关键作用。SIRT 1抑制NF-κB，SIRT 1是组蛋白去乙酰化酶sirtuin家族的一员，可保护阿托伐他汀处理的原代培养物免受小胶质细胞毒性。在AD大脑中，SIRT 1水平显著降低。在培养的小胶质细胞中SIRT 1的表达显着减少的ARAPINE治疗。基于这些发现，我们假设SIRT 1减少是导致AD中小胶质细胞毒性的关键事件，并且小胶质细胞SIRT 1通过抑制NF-κB活化来限制Ablast介导的神经元缺陷。为了验证这一假设，我们提出了三个具体目标。在目标1中，我们将在表达人淀粉样前体蛋白（hAPP）的小鼠的小胶质细胞中检测SIRT 1，并系统地研究小胶质细胞SIRT 1失活如何影响炎症反应和AAPP相关的神经元/行为缺陷。在目的2中，为了确定小胶质细胞SIRT 1是否通过抑制NF-κB活化发挥神经保护作用，我们将确定hAPP小鼠小胶质细胞中典型NF-κB信号传导的组成性活化是否以类似于SIRT 1缺失的方式加剧缺陷。在补充实验中，我们将通过在脑中注入有效的NF-κB抑制剂或注射抑制小胶质细胞中NF-κB的病毒载体来确定抑制NF-κ B信号传导是否会改善Ablation相关的神经元缺陷。在目的3中，为了确定小胶质细胞SIRT 1抑制NF-κB的机制，我们将系统地检查SIRT 1是否通过脱乙酰化RelA和/或通过减少髓样细胞中RelA磷酸化来抑制NF-κB。使用染色质免疫沉淀（ChIP）分析，我们将确定SIRT 1诱导的NF-κB活化抑制是否涉及髓样细胞中组蛋白（H3 K56 Ac）的脱乙酰化。这些研究的完成将为神经退行性变中调节小胶质细胞环的分子机制提供新的见解。这些研究也将为我们开发SIRT 1增强策略作为AD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 (Aß) 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-κB activation in microglia plays a critical role in microglial-mediated Aß toxicity. Inhibition of NF-κB by SIRT1, a member of the sirtuin family of histone deacetylases, protected against microglia toxicity in Aß-treated primary cultures. In AD brains, SIRT1 levels were markedly reduced. SIRT1 expression in cultured microglia was significantly diminished by Aß 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 Aß-mediated neuronal deficits by suppressing NF-κB 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 Aß-related neuronal/behavioral deficits. In Aim 2, to determine if microglial SIRT1 exerts neuroprotection by suppressing NF-κB activation, we will determine if constitutive activation of canonical NF-κB 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-κB signaling will ameliorate Aß-associated neuronal deficits by infusing a potent NF-κB inhibitor in the brain or injection of a viral vector that inhibits NF-κB in microglia. In Aim 3, to determine the mechanism by which microglial SIRT1 inhibits NF-κB, we will systematically examine if SIRT1 inhibits NF-κB 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-κB 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.