Microglia as Mediators of Dendritic Spine Loss and Plaque Formation in the AD Brain

项目2：小胶质细胞作为阿尔茨海默病脑内树突状脊髓丢失和斑块形成的媒介 项目摘要/摘要 树突棘丢失与阿尔茨海默病(AD)和其他疾病的认知能力下降密切相关 精神错乱。确定疾病中导致脊柱丢失的机制和刺激因素对疾病的发展至关重要 扭转或防止这些损失的策略，有望导致认知的改善。与并发 脊椎丧失，慢性小胶质细胞激活在AD大脑和其他疾病中被发现。作为我们的一部分 研究炎症在AD发病机制中的作用，我们以集落刺激因子1受体为靶点 (CSF1R)，因为这调节小胶质细胞的增殖。我们发现小胶质细胞在生理上 依赖于CSF1R信号和CSF1R拮抗剂的给药导致快速和 继续从中枢神经系统中消除几乎所有的小胶质细胞。我们已经使用这种方法来确定 在成人脑清除中，小胶质细胞确实在调节树突棘数量方面起着非常重要的作用 小胶质细胞作用8周后，CA1层和V层皮质神经元的棘密度增加约35%。 此外，电生理学显示对神经元的兴奋性突触输入显著增加，显示出直接的 突触活动增加的证据。正如我们已经证明的，小胶质细胞在调节脊柱和 成人大脑中的突触，我们现在提出，这种正常功能在AD时会出错，导致过度修剪 并导致脊椎密度降低和随后的认知能力下降。我们的项目提出了 4个相互关联的目标，将利用人体组织来探索小胶质细胞和树突棘之间的关系 在AD中，斑块的形成和丢失。首先，我们将对小胶质细胞之间进行彻底的关联。 对照组、MCI和AD受试者死后组织中脊椎的密度和形态。我们会 然后用高病理对照受试者的组织来检验我们的假设。我们将利用人类成纤维细胞 来自MCI受试者，具有高数量的AD小胶质风险SNP或低数量的SNP，这些SNP被转换 通过iPS细胞技术分化为多能干细胞，然后分化为小胶质细胞。然后我们将探索 这些MCI患者的小胶质细胞在1)修剪树突棘和2)修剪树突棘的能力上有何不同 吞噬和清除A‘，将这些发现与我们的MCI受试者转化为AD相关。 对我们的方法至关重要的是，我们现在有了消除小鼠体内所有内源性小胶质细胞的技术 通过给予CSF1R抑制剂的CNS，然后通过注入人的IPS重新填充小鼠的大脑- 衍生的小胶质细胞。使用这种方法，我们可以在活体环境中探索这些细胞的影响，从而 确定这些人源性小胶质细胞对树突棘和清除/形成的影响 一块斑块。通过这些实验，我们将能够充分研究人与人之间的关系 小胶质细胞和AD病理/脊柱丢失，这在以前是不可能的。这些结果将 潜在地导致抑制物的发展，这种抑制物可以消除AD大脑中的小胶质细胞，从而防止 脊椎缺失。

Project 2: Microglia as Mediators of Dendritic Spine Loss and Plaque Formation in the AD Brain Project Summary/Abstract Dendritic spine loss is closely associated with cognitive decline in Alzheimer's disease (AD) and other disorders. Identifying the mechanisms and stimuli that lead to spine loss in disease is crucial to developing strategies to reverse or prevent these losses, hopefully leading to improvements in cognition. Concurrent with spine loss, chronic microglial-activation is found in the AD brain and in other disorders. As part of our investigations into inflammation in the pathogenesis of AD, we targeted the colony-stimulating factor 1 receptor (CSF1R), as this regulates the proliferation of microglia. We discovered that microglia are physiologically dependent upon CSF1R signaling and that administration of CSF1R antagonists results in the rapid and continued elimination of virtually all microglia from the CNS. We have used this approach to determine that microglia do play a highly significant role in regulating dendritic spine numbers in the adult brain  elimination of microglia for 8 weeks results in a ~35% increase in spine densities in CA1 and layer V cortical neurons. Additionally, electrophysiology reveals robustly increased excitatory synaptic inputs to neurons, showing direct evidence of increased active synapses. As we have shown that microglia play a role in modulating spine and synapses in the adult brain, we now propose that this normal function goes awry in AD, leading to overpruning of synapses and resulting in reduced spine densities and subsequent cognitive decline. Our project proposes 4 linked aims that will utilize human tissue to explore the relationship between microglia and dendritic spine loss, as well as plaque formation, in AD. Firstly, we will conduct thorough correlations between microglial densities and morphologies with spine loss from post-mortem tissues in control, MCI and AD subjects. We will then test our hypothesis using tissue from high-pathology control subjects. We will utilize human fibroblasts from MCI subjects, with either a high number of AD microglial-risk SNPs or a low number, which are converted to pluripotent stem cells via iPS cell technology, and then differentiated into microglia. We will then explore how microglia derived from these MCI patients differ in their abilities to 1) prune dendritic spines and 2) phagocytose and clear A¿, correlating these findings with the conversion into AD from our MCI subjects. Critical to our approach, we now have the technology to eliminate all endogenous microglia from the mouse CNS via administration of CSF1R inhibitors and then repopulate the mouse brain by infusing in human IPS- derived microglia. Using this method, we can explore the effects of these cells in an in vivo setting and thus determine the effects of these human-derived microglia on both dendritic spines and on clearance/formation of A¿ plaques. Through these experiments we will be able to fully study the relationship between human microglia and AD pathology/spine loss in a fashion that has not been previously possible. These results will potentially lead to the development of inhibitors that can eliminate microglia in the AD brain and hence prevent spine loss.