Astrocyte Lipid Droplets as a Novel Mechanism for Impaired Glucose Metabolism in the E4 and AD Brain

项目总结 阿尔茨海默病(AD)是一种社会负担，每年给国家造成2770亿美元的损失，是 第六大死因。随着对阿尔茨海默病的疾病修饰疗法的探索继续，更深层次的 了解阿尔茨海默病大脑中的细胞代谢对指导研究至关重要。载脂蛋白E(ApoE) 基因是晚发性AD(LOAD)的最强遗传预测因子。在人类中，有三种主要的同工酶 APOE：E2、E3和E4。E3是人类表达的主要亚型(约占总人口的60%)。E4在 与E3相比，负载风险增加2倍(杂合子)到15倍(纯合子)。载脂蛋白E(ApoE) 主要由星形胶质细胞分泌，作为大脑中的主要脂质载体，并已获得作为调节剂的牵引力 大脑新陈代谢。例如，E4携带者表现出葡萄糖摄取减少，通过18fludeoxy- 葡萄糖正电子发射断层扫描(FDG-PET)。在AD患者中，FDG-PET同样较低，但E4 携带者在出现症状性认知功能下降前几十年表现出低FDG-PET。我们最近展示了 与表达E4的星形胶质细胞相比，表达E4的星形胶质细胞以脂滴(LDs)的形式积累更多的脂肪 E3，这是一个有趣的发现，根据1907年阿尔茨海默病的最初报告，它牵涉到神经胶质脂质堆积 作为疾病的标志。因为LDS被证明与代谢性疾病有关--包括调节 葡萄糖摄取-我们假设，星形胶质细胞LDS的过度积累推动了葡萄糖摄取的受损 见于E4和AD的大脑。因此，这项提议测试了一种机制，通过这种机制，LD相关蛋白 隔离是葡萄糖转运蛋白运输的关键媒介，导致星形胶质细胞葡萄糖减少 领悟。我们将在体内人类E3和E4纯合子小鼠身上测试这一机制(目标1A)，并(目标1B- C)体外培养表达人E3和E4的原代星形胶质细胞。我们还将通过以下方式翻译我们的调查结果(目标2) APOE基因分型人死后脑组织中LD形成的特征，与脂质丰度相关 来自英国阿尔茨海默病中心队列的临床数据。这一奖学金的主要焦点是揭示 驱动E4星形胶质细胞葡萄糖摄取受损的细胞机制，以及使受训者掌握 质谱学、共聚焦显微镜、免疫组织化学、放射性示踪、转化人 标本处理、数据分析等。这些技能和出色的指导将有助于 实习生成为一名独立的内科医生兼科学家的道路。

PROJECT SUMMARY Alzheimer’s disease (AD) is a societal burden, costing the nation 277 billion dollars per year and is the sixth leading cause of death. As the search for disease modifying therapies for AD continues, a deeper understanding of cellular metabolism in the AD brain is critical to guide research. The apolipoprotein E (APOE) gene is the strongest genetic predictor of late onset AD (LOAD). In humans, there are three major isoforms of apoE: E2, E3, and E4. E3 is the major isoform expressed in humans (~60% of population). E4 confers between a 2 (heterozygous) to 15-fold (homozygous) increase in risk of LOAD compared to E3. Apolipoprotein E (apoE) is primarily secreted by astrocytes as a primary lipid carrier in the brain, and has gained traction as a regulator of cerebral metabolism. For example, E4 carriers exhibit decreased glucose uptake as measured by 18fludeoxy- glucose positron emission tomography (FDG-PET). FDG-PET is similarly lower in AD individuals, however E4 carriers show low FDG-PET decades before the onset of symptomatic cognitive decline. We recently showed that E4 expressing astrocytes accumulate significantly more lipid in the form of lipid droplets (LDs) compared to E3, an intriguing finding in light of the initial 1907 report from Alzheimer that implicated glial lipid accumulation as a hallmark of disease. Since LDs have been shown to be involved in metabolic disease – including regulation of glucose uptake – we hypothesize that over accumulation of astrocyte LDs drive the impaired glucose uptake seen in E4 and AD brains. Therefore, this proposal tests a mechanism by which an LD-associated protein sequesters a critical mediator of glucose transporter trafficking, leading to a decrease in astrocyte glucose uptake. We will test this mechanism (Aim 1A) in vivo in mice homozygous for human E3 and E4, and (Aim 1B- C) in vitro in primary astrocytes that express human E3 and E4. We will also translate our findings (Aim 2) by characterizing LD formation in APOE genotyped human post-mortem brain tissue, correlating lipid abundance with clinical data from the UK Alzheimer’s Disease Center cohort. The key focus of this fellowship is to uncover a cellular mechanism driving impaired glucose uptake in E4 astrocytes, as well as equip the trainee with skills in mass spectrometry, confocal microscopy, immunohistochemistry, radioactive tracing, translational human specimen handling, data analysis, and more. These skills and excellent mentorship will be instrumental in the trainee’s path to become an independent physician-scientist.