APOE e4 negative regulation of microglia-astrocytes crosstalk in Alzheimer's disease

阿尔茨海默病(AD)是最常见的神经退行性疾病。小胶质细胞在人类免疫缺陷中起着重要作用 支持正常的中枢神经系统功能，但在疾病中可能导致神经变性。我们确认了 内稳态(M0)和神经退行性(MGnD)小胶质细胞，也称为疾病相关小胶质细胞 (DAM)，受TGFb的相互抑制和APOE信号的诱导在不同的 包括AD在内的神经退行性变小鼠模型。然而，这两种主要表型的作用以及它们如何 影响疾病进展仍然是一个主要问题。重要的是，靶向小胶质细胞APOE信号的恢复 小胶质细胞的动态平衡特征与急性和慢性脑缺血模型的神经保护相关 神经退行性变。APOE信号以细胞自主的方式抑制PU1和转化生长因子β信号。人类 载脂蛋白E有三个变异体：ε2、ε3和ε4。载脂蛋白ε4是晚发性阿尔茨海默病最强的遗传风险因素 疾病(负荷)。我们的初步数据显示：1)人源化APOE的小胶质细胞中PU1的表达增加 ε-4小鼠；2)小胶质细胞表达载脂蛋白E-ε-4抑制小胶质细胞反应，诱导MGnD表型； 3)小胶质细胞载脂蛋白ε4的缺失恢复了与星形胶质细胞相关的APP/PS1小鼠的MGnD表型 向抗体斑块募集和神经炎斑块减少。这些数据表明，APOEε4信号 是APP/PS1小鼠小胶质细胞的重要调控途径。基于这些发现，我们假设在 载脂蛋白ε4携带者，载脂蛋白ε4‘锁定’小胶质细胞处于动态平衡状态导致吞噬功能受损 功能和小胶质细胞-星形胶质细胞串扰失调，这可能会产生根本后果 用于AD开发。我们将从以下几个方面阐述我们的假设： 目的1：确定载脂蛋白Eε4在小胶质细胞转录调控中的作用及其与 AD和非痴呆患者携带不同载脂蛋白E等位基因的星形胶质细胞。我们将确定是否 人载脂蛋白Eε4影响细胞反应、相互作用途径和功能的转录信号 调节，侧重于小胶质细胞和星形胶质细胞。 目的2：验证载脂蛋白Eε4小胶质细胞和星形胶质细胞在单细胞分辨率下的空间差异。我们 将利用免疫组织化学方法验证小胶质细胞和星形胶质细胞基因在脑内表达的差异 特异标记和多重错误稳健的荧光原位杂交(MerFish)。 综上所述，本研究旨在验证载脂蛋白ε4是否损害转录、空间和功能 阿尔茨海默病脑内MGnD-小胶质细胞的调节及其与星形胶质细胞的相互作用。

Alzheimer’s disease (AD) is the most prevalent neurodegenerative disorder. Microglia play an essential role in supporting normal CNS functions, but in disease may contribute to neurodegeneration. We identified homeostatic (M0) and neurodegenerative (MGnD) microglia, also referred to as disease associated microglia (DAM), that are regulated by the reciprocal suppression of TGFb and induction of APOE signaling in different neurodegenerative mouse models including AD. However, the role of these two major phenotypes and how they affect disease progression remains a major question. Importantly, targeting microglial APOE signaling restored the homeostatic signature of microglia associated with neuroprotection in acute and chronic models of neurodegeneration. APOE signaling suppressed PU.1 and TGFβ signaling in a cell-autonomous manner. Human APOE has three variants: ε2, ε3, and ε4. APOE ε4 is the strongest genetic risk factor for late-onset Alzheimer’s disease (LOAD). Our preliminary data show: 1) Increased expression of PU.1 in microglia from humanized APOE ε4 mice; 2) Expression of APOE ε4 in microglia impaired microglial response to induce a MGnD phenotype and 3) Deletion of microglial APOE ε4 restored MGnD phenotype in APP/PS1 mice associated with astrocytes recruitment towards Ab-plaques and reduction in neuritic plaques. These data indicate that APOE ε4 signaling is a crucial regulatory pathway in microglia in APP/PS1 mice. Based on these findings, we hypothesize that in APOE ε4 carriers, APOE ε4 ‘locks’ microglia in their homeostatic state leading to impaired phagocytic functions and dysregulated microglia-astrocyte crosstalk, which could have fundamental consequences for AD development. We will address our hypothesis in the following aims: Aim 1: Determine the role of APOE ε4 on transcriptional regulation of microglia and their crosstalk with astrocytes in AD and non-demented subjects carrying different APOE alleles. We will determine whether human APOE ε4 affects transcriptional signatures of cellular response, interactive pathways, and functional regulation, focusing on microglia and astrocytes. Aim 2: Validate the spatial differences in APOE ε4 microglia and astrocytes at single-cell resolution. We will validate the differences in microglia and astrocytic gene expression in brain, using immunohistochemistry of specific markers and multiplexed error-robust fluorescence in situ hybridization (MERFISH). IN SUMMARY, this study aims to validate whether APOE ε4 impairs transcriptional, spatial, and functional regulation of MGnD-microglia and their crosstalk with astrocytes in AD brain.