Investigating the cause of APOE4-associated microglial activation and its resulting neurotoxicity of tauopathy-afflicted neurons

项目摘要 载脂蛋白E的ɛ4等位基因是晚发性阿尔茨海默病的主要危险因素 (Ad)，尽管直接原因仍然是争论的来源。最近的数据表明，载脂蛋白E基因是 在AD和其他神经退行性疾病中发现的一种新的小胶质细胞激活状态上调 疾病相关小胶质细胞(DAM)。此外，APOE4的表达已经被证明诱导了一种神经毒性 在各种刺激存在的情况下激活小胶质细胞，包括患有自闭症的神经元，尽管 这种激活的机制尚不清楚。为了发现APOE4的作用和机制- 相关的小胶质细胞激活及其导致的神经毒性，我已经设计了一种 一套创新的实验，既利用尖端技术，又利用新资源。在目标1中，单一- 细胞RNA测序将对从一种新的小鼠系中纯化的小胶质细胞进行，在该小鼠系中，人APOE3 APOE4小鼠与EC-Tau小鼠杂交，EC-Tau小鼠主要表达促聚集突变tau蛋白 在内嗅皮层(EC)。在目标2中，将研究载脂蛋白4的S对小胶质细胞激活的影响 人类小胶质细胞的设置，使用新创建的等基因APOE人类干细胞系和强大的 生物信息学分析。在目标3中，将利用一种强大的体外共培养试验来揭示 APOE4相关的小胶质细胞激活和由此产生的神经毒性的机制 确定抑制这些事件的新的治疗策略。作为载脂蛋白4和小胶质细胞的激活 显著影响AD，发现这两个重要参与者之间的机制联系和治疗 调控APOE4相关的小胶质细胞激活的策略将是在 并将极大地推进我们预防或减缓AD进展的目标，特别是在 载脂蛋白E4携带者。

Project Summary Possession of the ɛ4 allele of apolipoprotein E (APOE) is a major risk factor for late onset Alzheimer's disease (AD), although the direct cause remains a source of debate. Recent data has shown that the APOE gene is upregulated in a novel microglial activation state found in AD and other neurodegenerative diseases termed disease associated microglia (DAM). Furthermore, APOE4 expression has been shown to induce a neurotoxic activation of microglia in the presence of various stimuli, including tauopathy-afflicted neurons, although the mechanism of this activation has yet to be elucidated. In order to discover the role and mechanism of APOE4- associated microglial activation and its resulting neurotoxicity of tauopathy-afflicted neurons, I have devised an innovative set of experiments that utilize both cutting-edge technology and novel resources. In Aim 1, single- cell RNA-sequencing will be performed on microglia purified from a novel mouse line in which human APOE3 and APOE4 mice are crossed with EC-Tau mice, which express a pro-aggregating mutant tau protein primarily in the entorhinal cortex (EC). In Aim 2, APOE4's effects on microglial activation will be investigated in the setting of human microglia, using both a newly created isogenic APOE human stem cell line and powerful bioinformatics analyses. And in Aim 3, a robust in vitro co-culture assay will be utilized in order to both uncover the mechanism responsible for APOE4-associated microglial activation and the resulting neurotoxicity and to identify novel therapeutic strategies for inhibiting these events. As both APOE4 and microglial activation significantly impact AD, discovery of the mechanistic link between these two important players and therapeutic strategies for modulating APOE4-associated microglial activation would represent a major breakthrough in the AD field and would greatly advance our goal of preventing or slowing the progression of AD, especially among APOE4 carriers.