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Elucidating endolysosomal trafficking dysregulation induced by APOE4 in human astrocytes

阐明人星形胶质细胞中 APOE4 诱导的内溶酶体运输失调

基本信息

批准号：
10835321
负责人：
Julia TCW
金额：
$ 4.57万
依托单位：
BOSTON UNIVERSITY MEDICAL CAMPUS
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-04-01 至 2028-03-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10835321
关键词：
Affect Alzheimer&apos s Disease Alzheimer&apos s disease brain Alzheimer&apos s disease patient Alzheimer&apos s disease risk Alzheimer&apos s disease therapeutic Apolipoprotein E Astrocytes Autopsy Brain CRISPR interference Cholesterol Clinical Data Defect Dementia Demyelinations Development Disease Drug Targeting Exhibits Future Genes Genetic Screening Genetic Transcription Genotype Goals Human Impairment In Vitro Intracellular Accumulation of Lipids Knowledge Late Onset Alzheimer Disease Lipids Longitudinal Studies Lysosomes Mediating Microglia Modality Molecular Nerve Degeneration Neuroglia Pathogenesis Pathology Pathway interactions Phagocytosis Phenotype Research Project Grants Risk Symptoms Testing Therapeutic Variant Xenograft procedure apolipoprotein E-4 autosome brain cell brain endothelial cell causal variant cell type cholesterol biosynthesis disorder risk effective therapy endophenotype genetic risk factor genetic variant genome wide association study in vivo induced pluripotent stem cell lipid transport neuropathology novel risk variant stem cell model targeted treatment trafficking transcriptome

项目摘要

Project Summary Alzheimer’s disease (AD) is the most common form of dementia without effective treatments, underscoring the need for a better understanding of AD pathogenesis. Longitudinal studies in autosomal dominant and sporadic AD have demonstrated that pathology begins 10-20 years before clinical symptoms, but developmental effects of AD-associated genetic variants likely provide a substrate for future neuropathological changes. Many AD causal GWAS variants are associated with genes involved in endolysosomal pathways in glia. However, how these causal genes are affecting cellular mechanisms has to be further investigated to tackle the disease. One of the important questions is whether these endolysosomal pathway genes converge on ideally one clear phenotype that can be targeted for therapeutics. Among others, Apolipoprotein E (APOE) is the most significant risk for late-onset AD—homozygosity for the risk allele APOE ɛ4 (APOE 44) increases AD risk by more than 15- fold. To comprehensively assess the effect of human APOE4 on human brain cell types, we characterized the APOE4 genotype-phenotype relationship in four brain cell types: microglia, astrocytes, brain microvascular endothelial cells and mixed cortical cultures derived from human induced pluripotent stem cells (iPSCs). Global transcriptome analyses and in vitro mechanism study reveal that human APOE 44 astrocytes sequester cholesterol in lysosomes, leading to upregulated cholesterol biosynthesis despite elevated intracellular cholesterol. Our data suggests that the APOE4-mediated lipid accumulation impairs multiple intracellular trafficking pathways that converge on the lysosome. Therefore, we hypothesize that intracellular lipid accumulation in APOE 44 astrocytes jams trafficking to the lysosome (Aim 1), which are controlled by upstream regulators that can be identified by CRISPRi genetic screening (Aim 2). The APOE4-led endolysosomal defects in vitro astrocytes can be exacerbated in vivo by excessive lipid challenge induced by neurodegeneration (Aim 3). To test these hypotheses, in Aim 1, we will determine the mechanistic defects of lipid-mediated endolysosomal trafficking in vitro human APOE 44 astrocytes in transcriptional and functional changes. The identified phenotypes will be validated in AD post-mortem brain. In Aim 2, using CRISPRi screen on APOE 44 astrocytes, we will identify targets to reverse defected phagocytosis and intracellular lipid accumulation and determine the mechanisms by CROP-seq. In Aim 3, we will investigate mechanistic endolysosomal defects in vivo xenotransplanted astrocytes at baseline and during demyelination-associated lipid debris challenge and further test if CRISPRi-targeted astrocytes exhibit rescued phenotypes in vivo. The goal of this project is to assess the molecular mechanisms of APOE4-driven endolysosomal and autophagic defects in lipid trafficking and identify regulatory targets that reverse the phenotype. Further, this proposed research project sets out to uncover potential therapeutic drug targets to tackle APOE4-driven endolysosomal trafficking endophenotypes found in APOE4 carrier AD patients.

项目总结