Deciphering isogenic APOE isoform dependent neurodegenerative response in human glia

破译人类神经胶质细胞中同基因 APOE 亚型依赖性神经退行性反应

基本信息

批准号：
10580481
负责人：
Julia TCW
金额：
$ 12.3万
依托单位：
BOSTON UNIVERSITY MEDICAL CAMPUS
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-05-01 至 2024-02-29
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10580481
关键词：
AD transgenic mice Address Affect Age Aging Alleles Alzheimer&apos s Disease Alzheimer&apos s disease model Alzheimer&apos s disease pathology Alzheimer&apos s disease risk Amyloid Amyloid beta-Protein Apolipoprotein E Astrocytes Autophagocytosis Bioinformatics Biological Assay Brain Brain region CRISPR/Cas technology Cell Differentiation process Cell physiology Cells Clinical Clinical Trials Data Development Disease Disease associated microglia E protein Environment Exhibits Future Gene Expression Gene Expression Profiling Genetic Genetic Risk Genetic Transcription Genotype Goals Human In Vitro Infant Inflammation Inflammatory Response Inherited Knock-out Knowledge Late Onset Alzheimer Disease Lead Life Lipids Longitudinal Studies Membrane Lipids Metabolic Microglia Modality Modeling Molecular Myelin Nerve Degeneration Neurodegenerative Disorders Neurofibrillary Tangles Neuroglia Neurons Organoids Pathogenesis Pathology Pathway Analysis Patients Phagocytes Phagocytosis Phenotype Population Production Protein Isoforms Regulation Research Research Project Grants Senile Plaques Series Signal Transduction Symptoms System Systems Biology Testing Tissue-Specific Gene Expression Tissues Variant autosomal dominant Alzheimer&apos s disease base brain cell cell injury cell type cytokine disorder risk embryo tissue gene network genetic risk factor genetic signature genome editing gray matter human model imaging study induced pluripotent stem cell mouse model neuroimaging neuroinflammation novel therapeutics particle response risk variant single-cell RNA sequencing stem cell model tau Proteins tool transcriptome transcriptome sequencing transcriptomics

项目摘要

PROJECT SUMMARY Apolipoprotein E (APOE) is the most significant risk gene for late-onset Alzheimer's disease (AD). APOE ɛ4/ɛ4 homozygosity increases AD risk by >14-fold. Although an association between the APOE ε4 allele and increased AD risk is well-established, the mechanisms underlying this genetic risk remain elusive. In brain, microglia and astrocytes induce inflammation and degrade (engulf, digest, store or recycle) lipid-rich cellular debris that accumulates during aging and neurodegeneration. We hypothesize that APOE ɛ4/ɛ4 genotype has cell autonomous effects on astrocytes and microglia which affects other cell types. Specifically, we hypothesize that APOE ε4/ε4 astrocytes and microglia exhibit altered response to lipid-rich debris (myelin fragments), detectable as changes in global transcription and cellular function. To test this hypothesis we have generated a unique series of isogenic iPSCs differing solely in APOE isoform using a CRISPR/Cas9 genome-editing tool. We have established a platform to recapitulate cellular systems of human brain in culture by differentiation of iPSC-derived astrocytes and microglia that express APOE. We seek to bring together this established patient- derived isogenic human iPSC model, powerful systems biology, bioinformatic approaches and in vitro metabolic assays including neuroinflammation, phagocytosis and autophagy to understand the mechanism underlying APOE risk for AD. This proposed research project sets out to unravel the APOE isoform-dependent effects in glia and their responses to lipid challenge. In aim 1, we will generate homogenous populations of astrocytes and microglia and mixed cultures of cortical neurons/glia from isogenic APOE isoforms and APOE knockout derived from patient iPSCs. We will perform differential gene expression analysis to determine the downstream effects of APOE genotype in each cell type. In aim 2, we will study APOE isoform-dependent glial responses to challenge with lipid-rich particles. Following an unbiased transcriptomic approach, we will analyze which of the disease associated microglia signatures or AD-associated networks are APOE isoform-dependent upon challenge. In aim 3, we will investigate APOE isoform-dependent effects on the inflammatory response and phagocytic/autolysosomal clearance of lipid particles. The goal of this project is to identify the transcriptomic networks and cellular functions governed by APOE genotype in the presence or absence of a disease relevant environment (myelin debris) to pinpoint the earliest and potentially most treatable mechanisms involved in AD pathogenesis.

项目概要载脂蛋白 E (APOE) 是晚发性阿尔茨海默病 (AD) 的最重要风险基因。尽管 APOE ε4 等位基因与纯合性之间存在关联，但 AD 风险会增加 14 倍以上。 AD 风险增加已是公认的事实，但这种遗传风险背后的机制在大脑中仍然难以捉摸。小胶质细胞和星形胶质细胞诱导炎症并降解（吞噬、消化、储存或回收）富含脂质的细胞我们发现 APOE ɛ4/ɛ4 基因型具有衰老和神经退行性变过程中积累的碎片。对星形胶质细胞和小胶质细胞的细胞自主作用会影响其他细胞类型。 APOE ε4/ε4 星形胶质细胞和小胶质细胞对富含脂质的碎片（髓磷脂碎片）表现出不同的反应，可以通过整体转录和细胞功能的变化来检测。为了检验这一假设，我们生成了一个假设。使用 CRISPR/Cas9 基因组编辑工具，获得一系列独特的同基因 iPSC，仅 APOE 亚型不同。我们建立了一个平台，通过区分培养物中的人脑细胞系统我们力求将这一已建立的患者聚集在一起。衍生的同基因人类 iPSC 模型、强大的系统生物学、生物信息方法和体外代谢检测，包括神经炎症、吞噬作用和自噬，以了解其机制 AD 的潜在 APOE 风险该拟议研究项目旨在解开 APOE 异构体依赖性。神经胶质细胞的影响及其对挑战脂质的反应在目标 1 中，我们将产生同质的群体。星形胶质细胞和小胶质细胞以及来自同基因 APOE 亚型和 APOE 的皮质神经元/神经胶质细胞的混合培养物我们将进行差异基因表达分析，以确定来自患者 iPSC 的敲除。 APOE 基因型对每种细胞类型的下游影响在目标 2 中，我们将研究 APOE 亚型依赖性神经胶质细胞。我们将采用无偏见的转录组学方法来分析对富含脂质颗粒的挑战的反应。哪些疾病相关的小胶质细胞特征或 AD 相关网络是 APOE 同工型依赖性的在目标 3 中，我们将研究 APOE 同工型依赖性对炎症反应的影响。和脂质颗粒的吞噬/自溶酶体清除该项目的目标是确定脂质颗粒的吞噬/自溶酶体清除。在存在或不存在 a 的情况下，由 APOE 基因型控制的转录组网络和细胞功能疾病相关环境（髓磷脂碎片），以查明最早且最可能可治疗的疾病 AD 发病机制。