Elucidating roles of microglial lipid droplets in neurodegeneration

项目摘要/摘要 阿尔茨海默病(AD)在美国影响着大约600万人，目前没有 明显有效的疾病修正疗法。AD的病理定义是淀粉样蛋白β的堆积 (aβ)斑块和tau神经原纤维在大脑中缠绕在一起，已知 小胶质细胞和其他非神经细胞类型在形成疾病过程中起着重要作用。世界上大部分的 阿尔茨海默病的多基因风险源于小胶质细胞表达的基因变异，特别是那些涉及 内溶酶体和脂质加工途径，以及含有丰富脂滴的小胶质细胞(LD-MG) 在死后的人类阿尔茨海默病的大脑中观察到。由于之前针对Aβ的干预措施尚未 在临床试验中取得成功，调节小胶质细胞脂代谢的概念是一个新的和令人兴奋的途径 然而，在临床前研究中，我们需要更多地了解脂代谢是如何调控的 小胶质细胞功能状态。在淀粉样变性的小鼠模型中，小胶质细胞从稳态转变为 疾病相关(DAM)转录状态，代表一种保护性、吞噬和斑块- 紧凑表型。虽然小胶质细胞的活动在AD的早期淀粉样蛋白阶段可能是有益的， 小胶质细胞的药物或遗传抑制已被证明在小鼠模型中具有保护作用 紧张症。在一种后遗症小鼠模型中，在活跃的变性脑区观察到LD-MG，但 这些细胞在疾病过程中的功能目前尚不清楚。为了开始解决这个问题，我们 将使用FACS和scRNAseq来表征LDHigh和LDlow之间的任何转录差异 从9.5月龄自闭症小鼠体内分离出小胶质细胞。鉴于最近的证据表明LD-MG 在老年小鼠中观察到有促炎、低吞噬的表型，我们推测LD-MG 在我们的肌萎缩侧索硬化症模型中，会有类似的通路改变，因此可能是部分原因 小胶质细胞在神经变性中的作用。为了评估这些发现与人类的相关性，我们将 对死后的人AD脑样本进行染色，以寻找任何有希望的小鼠LD-MG标记。为了澄清 LD-MG在肌萎缩侧索硬化症进展中的功能作用，我们将利用一种新的小鼠模型，该模型允许 二酰甘油酰基转移酶(DGAT)酶的可诱导的、小胶质细胞特异性的敲除，已被 证明了在多种生物环境中LD的生物发生所必需的。大量的文学作品 支持脂滴在隔离潜在有毒脂类中的作用，我们假设 MG通过DGAT KO形成LDS的能力将加速肌张力障碍的进展，我们将使用 结合免疫组织化学、scRNAseq和脂肪组学分析。我们的研究将描述一个 对小胶质细胞亚群的研究不足，增加了我们对髓系功能状态多样性的了解，以及 评价调节小胶质细胞脂代谢治疗阿尔茨海默病的可能性。

PROJECT SUMMARY/ABSTRACT Alzheimer disease (AD) affects approximately 6 million people in the United States and currently has no clearly-effective disease modifying therapies. AD is pathologically defined by the accumulation of amyloid β (Aβ) plaques and tau neurofibrillary tangles in the brain and it is known that alterations in the activities of microglia and other non-neuronal cell types play important roles in shaping the disease course. Much of the polygenic risk for AD is derived from variants in genes expressed by microglia, specifically those involved in endolysosomal and lipid processing pathways, and microglia containing abundant lipid droplets (LD-MG) have been observed in post-mortem human AD brains. As previous interventions targeting Aβ have not yet been successful in clinical trials, the concept of modulating microglial lipid metabolism is a novel and exciting avenue being explored in preclinical studies, however we need to know more about how lipid metabolism governs microglial functional states. In mouse models of amyloidosis, microglia transition from a homeostatic to a disease-associated (DAM) transcriptional state that represents a protective, phagocytic, and plaque- compacting phenotype. While microglial activity may be beneficial in the early amyloid phase of AD, pharmacological or genetic inhibition of microglia has been shown to be protective in mouse models of tauopathy. LD-MG are observed in actively degenerating brain regions in a tauopathy mouse model, but the functions of these cells in the disease process are not currently known. To begin addressing this question, we will use FACS coupled with scRNAseq to characterize any transcriptional differences between LDhigh vs LDlow microglia isolated from 9.5 month old tauopathy mice. Given recent evidence suggesting that the LD-MG observed in aged mice have pro-inflammatory, hypo-phagocytic phenotypes, we hypothesize that the LD-MG in our tauopathy model will have similar pathway alterations and could thus be partially responsible for microglial contributions to neurodegeneration. To assess the relevance of these findings to humans, we will stain postmortem human AD brain samples for any promising mouse LD-MG markers. To elucidate the functional roles of LD-MG in the progression of tauopathy, we will utilize a new mouse model that allows for the inducible, microglial-specific knockout of the diacylglycerol acyltransferase (DGAT) enzymes, which have been demonstrated to be required for LD biogenesis in multiple biological contexts. A substantial amount of literature supports a role for lipid droplets in sequestering potentially toxic lipids and we hypothesize that crippling the ability of MG to form LDs via DGAT KO will accelerate tauopathy progression, which we will assess using a combination of immunohistochemical, scRNAseq, and lipidomic analyses. Our studies will characterize an understudied subset of microglia, increase our knowledge of the diversity of myeloid functional states, and evaluate the potential of modulating microglial lipid metabolism for the treatment of Alzheimer disease.