RICTOR dependent microglial polarization in aging

衰老是包括阿尔茨海默病(AD)在内的大多数神经退行性疾病的主要危险因素。这个 生物体因衰老而出现的功能障碍在分子水平上是多方面的。 了解关键细胞通路中分子信号的变化如何产生与年龄相关的下降 必须开发更好、更有针对性的治疗方法来改善功能障碍。AD及相关疾病 我们老龄化人口面临的最严重的认知威胁。AD没有治愈的方法，这在一定程度上是由于 对衰老和炎症如何影响发病机制和神经可塑性缺乏了解。这是势在必行的 我们对大脑老化的生物学基础有了更全面的了解。我们假设 神经免疫系统在衰老病理中起着关键作用。小胶质细胞是大脑中常驻的巨噬细胞 和中枢神经系统中的主要免疫效应器。“激活的”小胶质细胞呈现促炎表型， 分泌细胞毒因子。另一个由抗炎分子介导的过程，抑制了促- 炎症表型和小胶质细胞向促进碎片清除的恢复性表型转变 和组织修复。衰老扰乱了小胶质细胞的生物学，使衰老的小胶质细胞对前体细胞变得高度反应 炎症刺激，产生过多的细胞毒性因子，对消炎不敏感 分子。衰老的小胶质细胞也越来越不能呈现抗炎表型。就这样， 功能障碍的小胶质细胞会产生神经毒性，增加中枢神经系统的脆弱性。我们之前已经描述过年龄- 用基于质谱学的蛋白质组学方法研究啮齿动物小胶质细胞的相关变化。我们确认了RICTOR，一个 MTORC2亚基，作为上游调节因子，预计会随着年龄的增长而被抑制。这一预测与 促炎症信号上调，抗炎信号下调，细胞受损 新陈代谢和能量调节。我们通过展示减少RICTOR表达来验证这一发现 与年轻的同龄人相比，衰老的小鼠原代小胶质细胞。我们还进行了有针对性的 RICTOR在BV2细胞中的敲除并观察到类似于衰老的小胶质细胞的表型，证实了原代 RICTOR在衰老表型中的作用。本研究将确定RICTOR的后果 体内基因敲除对幼鼠小胶质细胞表型和认知行为的影响。我们假设在活体内 小胶质细胞RICTOR缺失会导致小胶质细胞表型老化，从而降低大脑的弹性和 对认知和神经可塑性有广泛的影响。我们预测，敲除动物将表现出一种 表型表现为失调的神经炎症，认知和行为能力受损， 并降低了大脑的可塑性。我们的目标是确定体内小胶质细胞功能障碍的分子底物 通过mTORC2信令与提案中详细说明的实验进行比较。

Aging is the primary risk factor for most neurodegenerative disorders including Alzheimer's disease (AD). The dysfunction seen in organisms because of aging has a multifaceted origin at the molecular level. Understanding how altered molecular signaling in key cellular pathways generates age-related decline is essential to develop better, more targeted therapies that ameliorate dysfunction. AD and related diseases are the most crippling cognitive threat to our aging population. There is no cure for AD, and this is partly due to a poor understanding of how aging and inflammation impact pathogenesis and neural plasticity. It is imperative that we gain a more comprehensive understanding of the biological basis for brain aging. We hypothesize that the neuroimmune system plays a critical role in aging pathology. Microglia are the brain's resident macrophage and the major immune effector in the CNS. “Activated” microglia assume a pro-inflammatory phenotype, secreting cytotoxic factors. An additional process mediated by anti-inflammatory molecules, inhibits the pro- inflammatory phenotype and shifts microglial toward a restorative phenotype that promotes debris clearance and tissue repair. Aging perturbs microglial biology such that aging microglia become hyper-responsive to pro- inflammatory stimuli, generate excess cytotoxic factors, and become insensitive to anti-inflammatory molecules. Aged microglia are also increasingly unable to assume an anti-inflammatory phenotype. In this way, dysfunctional microglia become neurotoxic and increase CNS vulnerability. We have previously described age- related changes in rodent microglia using mass-spectrometry-based proteomics. We identified RICTOR, a subunit of mTORC2, as an upstream regulator predicted to be inhibited with age. This prediction coincided with upregulated pro-inflammatory signaling, downregulated anti-inflammatory signaling, and impaired cellular metabolism and energy regulation. We validated this finding by demonstrating reduced RICTOR expression in aged primary mouse microglia when compared with their younger counterparts. We also conducted targeted knockdown of RICTOR in BV2 cells and observed a phenotype resembling aged microglia, validating a primary role for RICTOR in the aging phenotype. The present study will determine the consequences of RICTOR knockdown in vivo on microglial phenotype and cognitive behavior in young mice. We hypothesize that in vivo microglial RICTOR deletion will induce an aging microglial phenotype that will decrease brain resilience and have widespread impact on cognition and neural plasticity. We predict that knockout animals will manifest a phenotype characterized by dysregulated neuroinflammation, impaired cognitive and behavioral performance, and reduced brain plasticity. Our goal is to identify the molecular substrates of in vivo microglial dysfunction via mTORC2 signaling with the experiments detailed in the proposal.