Elucidating the role of SORL1 as an APOE receptor in human astrocytes

阐明 SORL1 作为人星形胶质细胞中 APOE 受体的作用

基本信息

批准号：
9983410
负责人：
Hyo Lee
金额：
$ 3.88万
依托单位：
HARVARD MEDICAL SCHOOL
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-06-01 至 2022-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9983410
关键词：
Abeta clearance Address Affect Alzheimer&apos s Disease Alzheimer&apos s disease patient Alzheimer&apos s disease risk Amyloid beta-Protein Apolipoproteins Astrocytes Binding Biochemical Biological Biological Assay Biology Biotinylation Blood - brain barrier anatomy Brain CRISPR/Cas technology Cell surface Cells Cerebrospinal Fluid Characteristics Cholesterol Homeostasis Co-Immunoprecipitations Collecting Cell Complex Confocal Microscopy Data Dementia Drainage procedure Endocytosis Endosomes Endothelial Cells Enzyme-Linked Immunosorbent Assay Family Gene Family Generations Genes Genetic Genetic Transcription Genetic study Human Individual Inflammation Intercellular Fluid Knock-out LDL-Receptor Related Protein 1 Label Late Onset Alzheimer Disease Lead Ligand Binding Ligands Link Lipoprotein Receptor Lipoproteins Low Density Lipoprotein Receptor Lysosomes Measures Mediating Meta-Analysis Missense Mutation Mus Mutation Nervous system structure Neurodegenerative Disorders Neurofibrillary Tangles Neurons Pathology Pathway Analysis Pathway interactions Patients Play Polymorphism Analysis Presenile Alzheimer Dementia Property Protein Family Protein Isoforms Proteins RNA Splicing Risk Role Senile Plaques Technology Testing Transcript Variant Work abeta accumulation amyloid precursor protein processing apolipoprotein E-4 base brain cell causal variant cell type experimental study extracellular familial Alzheimer disease genetic risk factor genome wide association study human subject induced pluripotent stem cell loss of function neuron loss novel preference protein expression receptor risk variant stem cell technology tau Proteins trafficking transcriptome sequencing uptake

项目摘要

Project Summary/Abstract Alzheimer’s disease (AD) is a progressive neurodegenerative disease affecting more than 30 million people worldwide. It is the most common cause of dementia, and its pathology includes extensive neuronal loss, accumulation of extracellular amyloid-beta (Aβ) plaques and intracellular neurofibrillary tangles. A number of large scale meta-analyses of genome wide association studies (GWAS) has analyzed polymorphisms in many human subjects, and identified more than 20 genes associated with AD risk, which can be classified based on their known functions: cholesterol metabolism, inflammation, and endocytosis. Our lab seeks to utilize induced pluripotent stem cells (iPSCs) from patients with familial and sporadic AD to understand the cellular mechanisms underlying AD. My proposed project will investigate the potential convergent mechanisms of APOE and SORL1 function in the brain. Both APOE and SORL1 are AD risk genes, and APOE (ε4) is the strongest genetic risk factor for AD. Not only is SORL1 a GWAS hit for late onset AD (LOAD), but recent studies identified SORL1 loss-of-function variants in familial AD patients, suggesting that SORL1 can be a potential causal gene for AD. Here, I will study the function of SORL1 as a receptor for APOE, which can address the potential convergent mechanism between these genes. I hypothesize that SORL1 will function as an APOE receptor in human astrocytes and regulate Aβ clearance. In support of this, publicly available RNA-seq data show that SORL1 transcript is highly expressed in astrocytes over other brain cell types. In addition, APOE has a known role in Aβ clearance, and SORL1 is in the APOE receptor gene family. These findings, however, do not directly examine the role of SORL1 in regulating Aβ uptake in astrocytes. Here, I will utilize iPSC-derived astrocytes with different SORL1 and APOE variants to study the ability of SORL1 to function as an APOE receptor and uptake Aβ. In Aim 1, I will generate iPSC-derived astrocytes and determine SORL1 expression, subcellular localization, and ligand binding characteristics. Then, In Aim 2, I will use iPSC-derived astrocytes generated from individuals with 1) different APOE isoforms 2) SORL1 knock out (KO), risk SNPs and missense mutations and 3) other AD risk SNPs. I then will meticulously characterize Aβ generation, uptake and clearance in these astrocytes. Our preliminary data suggest that SORL1 is highly expressed in human astrocytes, and SORL1 KO in these astrocytes results in elevated extracellular Aβ levels. Also, I have shown that iPSC-derived astrocytes reduce extracellular Aβ levels when treated with exogenous Aβ, opening up the possibility/potential for this assay to examine the Aβ clearance ability of cells from individuals with various genetic backgrounds. In summary, my proposal will utilize iPSC-derived astrocytes to examine the novel role of SORL1 as an APOE receptor in astrocytes, and investigate a potential convergent mechanism that APOE and SORL1 play in Aβ clearance.

项目概要/摘要阿尔茨海默病 (AD) 是一种进行性神经退行性疾病，影响超过 3000 万人它是世界范围内痴呆症最常见的原因，其病理包括广泛的神经功能丧失，细胞外β-淀粉样蛋白（Aβ）斑块和细胞内神经原纤维缠结的积累。全基因组关联研究（GWAS）的大规模荟萃分析分析了许多基因组中的多态性人类受试者，并鉴定了 20 多个与 AD 风险相关的基因，这些基因可以根据它们的已知功能：胆固醇代谢、炎症和内吞作用。来自家族性和散发性 AD 患者的多能干细胞 (iPSC)，以了解细胞机制我提出的项目将研究 APOE 和 SORL1 的潜在收敛机制。 APOE 和 SORL1 都是 AD 风险基因，其中 APOE (ε4) 是最强的遗传风险基因。 SORL1 不仅是迟发性 AD (LOAD) 的 GWAS 命中目标，而且最近的研究还发现了 SORL1。家族性 AD 患者的功能丧失变异，表明 SORL1 可能是 AD 的潜在致病基因。在这里，我将研究SORL1作为APOE受体的功能，它可以解决潜在的收敛问题我推测 SORL1 将作为人类的 APOE 受体发挥作用。公开的 RNA-seq 数据显示 SORL1 可以支持星形胶质细胞并调节 Aβ 清除。转录物在星形胶质细胞中的表达高于其他脑细胞类型。此外，APOE 在 Aβ 中具有已知的作用。然而，这些发现并没有直接检查。 SORL1 在调节星形胶质细胞 Aβ 摄取中的作用在这里，我将利用具有不同功能的 iPSC 衍生的星形胶质细胞。 SORL1 和 APOE 变体用于研究 SORL1 作为 APOE 受体发挥作用并摄取 Aβ 的能力。 1、我将生成iPSC衍生的星形胶质细胞并确定SORL1表达、亚细胞定位和配体然后，在目标 2 中，我将使用由具有 1) 的个体生成的 iPSC 衍生星形胶质细胞。不同的 APOE 亚型 2) SORL1 敲除 (KO)、风险 SNP 和错义突变以及 3) 其他 AD 风险然后，我将仔细描述这些星形胶质细胞中 Aβ 的生成、摄取和清除。初步数据表明，SORL1 在人星形胶质细胞中高表达，并且 SORL1 在这些星形胶质细胞中被敲除。此外，我还发现 iPSC 衍生的星形胶质细胞会降低细胞外 Aβ 水平。当用外源 Aβ 处理时，细胞外 Aβ 水平，开启了该测定的可能性/潜力检查具有不同遗传背景的个体细胞的 Aβ 清除能力。该提案将利用 iPSC 衍生的星形胶质细胞来检查 SORL1 作为 APOE 受体的新作用星形胶质细胞中，并研究 APOE 和 SORL1 在 Aβ 中发挥的潜在收敛机制清除。