The impact of AD-associated genetic variants in 3D human mixed neural-glial models of AD

标题：AD 相关遗传变异对 AD 3D 人类混合神经胶质模型的影响 项目概要 越来越多的阿尔茨海默病 (AD) 相关基因与先天免疫和 神经炎症途径。基于网络的 AD 相关基因综合分析表明 小胶质细胞基因网络与 AD 神经病理学密切相关 (1,5)。我们已经证明了 AD 相关的保护性 CD33 变体 rs3865444 可导致 CD33 表达减少和 CD33 水平降低 Aβ42)(1)。相反，小胶质细胞 TREM2 变异会增加 AD 风险，并降低小胶质细胞对抗体的清除。在 除了 CD33 和 TREM2 中与 AD 相关的功能变异之外，我们的 AD 全基因组测序 (WGS) 和 来自 AD 家族和病例对照的全外显子组测序 (WES) 数据集揭示了功能 AD 相关小胶质细胞基因的变异与先天免疫和神经炎症有关，包括 CD33、 TREM2、MS4A 簇、ABCA7、ABI3、PLGC2、CR1 等。测试小胶质细胞遗传的影响 针对人类遗传背景上 AD 发病机制的变异，我们开发了一种新型 3D 人类神经元 使用独特的 3D 微流体系统的星形胶质细胞-小胶质细胞三培养 AD 模型。我们证明了人类 小胶质细胞通过小胶质细胞特异性被招募到 3D AD（产生抗体）神经元星形胶质细胞培养物中 迁移通道以趋化因子依赖性方式导致神经炎症和神经变性。 在这里，我们建议使用我们广泛收集的 AD WGS 和 WES 数据集，以及我们的 3D 人体 三培养 AD 模型，评估 AD 相关先天免疫功能变异的致病作用 与神经炎症有关的基因。在目标 1 中，我们将鉴定功能基因组变异和富集基因 与先天免疫和神经炎症相关的网络。然后我们将检查致病作用 3D 人类混合神经星形胶质细胞中小胶质细胞 AD 风险或保护基因及其功能变异的研究 AD 小胶质细胞模型（目标 2），探索与小胶质细胞 AD 风险基因相关的 AD 致病途径 及其功能变体，使用集成的多组学方法，并验证选择性阻断这些 途径（目标 3）。该提案的总体目标是全面评估致病影响 先天免疫 AD 风险基因的功能变异对 AD 发病机制的影响，并探索潜在的分子机制 网络，这将为 AD 患者提供新的治疗靶点。

Title: The impact of AD-associated genetic variants in 3D human mixed neural-glial models of AD Project summary A growing number of Alzheimer’s disease (AD)-associated genes are associated with innate immunity and neuroinflammatory pathways. Network-based integrative analyses of AD-related genes have shown that microglial gene networks are strongly associated with AD neuropathology (1,5). We have shown that the protective AD-associated CD33 variant, rs3865444, leads to reduced CD33 expression and lower levels of Aβ42)(1). Conversely, microglial TREM2 variants, which increase AD risk, reduce microglial clearance of Ab. In addition to AD-linked functional variants in CD33 and TREM2, our AD whole genome sequencing (WGS) and whole exome sequencing (WES) datasets from AD families and case-controls, have revealed functional variants in AD-associated microglial genes linked to innate immunity and neuroinflammation, including CD33, TREM2, MS4A cluster, ABCA7, ABI3, PLGC2, CR1, and others. To test the impact of microglial genetic variants on AD pathogenesis on human genetic background, we developed a novel 3D human neuron- astrocyte-microglia tri-culture AD model using a unique 3D microfluidic system. We demonstrated that human microglial cells are recruited towards 3D AD (Ab-producing) neuron-astrocyte cultures via microglia-specific migration channels, in a chemokine-dependent manner, leading to neuroinflammation and neurodegeneration. Here, we propose to use our extensive collection of AD WGS and WES datasets, together with our 3D human tri-culture AD model, to evaluate the pathogenic effects of functional variants in AD-associated innate immune genes linked to neuroinflammation. In Aim 1, we will identify functional genomic variants and enriched gene networks that are linked to innate immunity and neuroinflammation. We will then examine the pathogenic roles of microglial AD risk or protective genes and their functional variants in 3D human mixed neural-astrocyte- microglial models of AD (Aim 2), explore AD pathogenic pathways that are linked to microglial AD risk genes and their functional variants using integrated multi-omics approaches, and validate selectively blocking these pathway (Aim 3). The overarching goals of this proposal are to comprehensively assess the pathogenic effects of functional variants in innate immune AD-risk genes on AD pathogenesis and explore underlying molecular networks, which will provide novel therapeutic targets for AD patients.