Astrocyte-specific in vivo molecular signatures of APOE genetic risk in Alzheimer's disease

阿尔茨海默病 APOE 遗传风险的星形胶质细胞特异性体内分子特征

基本信息

批准号：
10679977
负责人：
CHRISTINA CATHERINE RAMELOW
金额：
$ 4.77万
依托单位：
EMORY UNIVERSITY
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：
2023-06-01 至 2026-05-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10679977
关键词：
Address Adopted Alleles Alzheimer&apos s Disease Alzheimer&apos s disease risk Apolipoprotein E Astrocytes Autopsy Biochemical Bioinformatics Biological Process Biotin Biotinylation Brain Brain region Cell Separation Cholesterol Complement Data Diagnosis Disease Disease Progression Exhibits Foundations Future Genetic Genetic Risk Genotype Goals Hippocampus Homeostasis Human Immune response Immunoassay Impaired cognition Individual Inflammation Inflammatory Inflammatory Response Knock-in Knowledge Label Laboratories Late Onset Alzheimer Disease Ligase LoxP-flanked allele MAP Kinase Gene MEKs Mass Spectrum Analysis Mediating Mediator Metabolism Methodology Microglia Modification Molecular Molecular Profiling Mouse Protein Mus Nerve Degeneration Neurodegenerative Disorders Pathogenesis Pathologic Pathway interactions Phenotype Phosphoproteins Play Pre-Clinical Model Production Proteins Proteome Proteomics RNA Risk Risk Factors Role Signal Pathway Signal Transduction Source Synapses TNF gene Testing Time Validation Work aged apolipoprotein E-3 apolipoprotein E-4 brain tissue cell type cytokine differential expression genetic approach glial activation high risk in vivo inhibitor innovation interest lipid metabolism metabolic phenotype mind control neuron loss neuropathology novel novel strategies novel therapeutic intervention novel therapeutics phosphoproteomics pre-clinical research protein aggregation response risk variant selective expression sugar therapeutically effective transcriptome sequencing transcriptomics

项目摘要

PROJECT ABSTRACT Alzheimer’s disease (AD) is a common and burdensome neurodegenerative disease. Apolipoprotein E (APOE) is the most prevalent risk factor for developing AD. APOE genotype may contribute to disease pathogenesis and risk by impacting astrocyte molecular profiles and subsequent biological functions. Data from my lab in human post-mortem AD and control brain proteomes identified a module of co-expressed astrocytic and microglial proteins in APOE ε4 individuals that were involved in sugar metabolism and inflammation and strongly correlated with the MAPK/ERK pathway. Because astrocytes are abundant throughout the brain, produce high levels of apoE protein, and play a critical role in brain homeostasis, they are more likely to contribute to disease vulnerability and pathogenesis of AD in the context of APOE risk. To address how APOE genotype impacts astrocyte function, my laboratory recently developed cell type-specific in vivo biotinylation of proteins (CIBOP) as a novel approach to quantify the total and phospho-proteomes of astrocytes in their native state without the need for cell isolation. In this approach, the biotin ligase, TurboID, is selectively expressed in the cell type of interest using a conditional Cre/lox genetic strategy. Using astrocyte-CIBOP in our preliminary studies, we have successfully obtained native-state proteomes of astrocytes from mouse brain, and quantified astrocyte- derived cytokines and MAPK/ERK signaling phospho-proteins. Leveraging this highly innovative methodology, my goal in the current study is to for the first time define the astrocyte-like immune response to APOE ε4 genotype and determine if the MAPK/ERK pathway is a mechanism of this response. My central hypothesis is that APOE ε4 genotype augments pro-inflammatory profiles of astrocytes via increased cytokine and complement production in an ERK signaling-dependent manner. I will use the astrocyte-CIBOP mice derived on homozygous human APOE 4/4 and APOE 3/3 knock-in genetic backgrounds to determine the differential impacts of APOE genetic risk on molecular signatures of astrocytes in vivo. In Aim 1, I will test the hypothesis that APOE 4/4 astrocytes, compared to APOE 3/3, will exhibit elevated levels of pro-inflammatory cytokines, increased complement protein production, amplified MAPK/ERK signaling, and aberrant changes in lipid metabolism. In Aim 2, I will test the hypothesis that the exaggerated pro-inflammatory response of APOE4 astrocytes, initiated by systemic inflammatory challenge (LPS), is mediated via ERK signaling. Using bioinformatic approaches on proteomic and transcriptomic data and validation IHC approaches in mouse and human brain tissues, I will identify molecular changes in reactivity states of APOE 4/4 and 3/3 astrocytes in vivo and identify ERK- dependent and ERK-independent mechanisms of APOE. My work will lay the foundation to further investigate astrocyte-mediated mechanisms of AD pathogenesis to employ CIBOP in future studies to explore cell type- specific mechanisms of neurodegeneration, beyond AD and astrocytes.

项目摘要阿尔茨海默氏病 (AD) 是一种常见且严重的神经退行性疾病。 (APOE) 是导致 AD 的最普遍的危险因素，APOE 基因型可能导致疾病。通过影响星形胶质细胞分子谱和随后的生物学功能数据来确定发病机制和风险。我的实验室在人类死后 AD 和对照大脑蛋白质组中发现了一个共表达星形胶质细胞的模块 APOE ε4 个体中参与糖代谢和炎症的小胶质蛋白和与 MAPK/ERK 通路密切相关，因为星形胶质细胞在整个大脑中含量丰富，产生高水平的 apoE 蛋白，并在大脑稳态中发挥关键作用，它们更有可能贡献 APOE 风险背景下 AD 的疾病脆弱性和发病机制解决 APOE 基因分型的问题。影响星形胶质细胞功能，我的实验室最近开发了细胞类型特异性蛋白质体内生物素化（CIBOP）作为一种定量星形胶质细胞天然状态下的总蛋白质组和磷酸蛋白质组的新方法无需细胞分离，生物素连接酶 TurboID 在细胞中选择性表达。在我们的初步研究中使用星形胶质细胞-CIBOP，使用条件Cre/lox遗传策略的兴趣类型，我们已经成功地从小鼠大脑中获得了星形胶质细胞的天然蛋白质组，并对星形胶质细胞进行了定量- 利用这种高度创新的方法，衍生细胞因子和 MAPK/ERK 信号磷酸蛋白。我当前研究的目标是首次定义对 APOE ε4 基因型的星形胶质细胞样免疫反应并确定 MAPK/ERK 通路是否是这种反应的机制我的中心假设是 APOE。 ε4 基因型通过增加细胞因子和补体增强星形胶质细胞的促炎特征我将使用源自纯合子的星形胶质细胞-CIBOP 小鼠。人类 APOE 4/4 和 APOE 3/3 敲入遗传背景以确定 APOE 的不同影响在目标 1 中，我将检验 APOE 4/4 的假设。与 APOE 3/3 相比，星形胶质细胞将表现出促炎细胞因子水平升高、补体蛋白的产生、MAPK/ERK 信号传导的放大以及脂质代谢的异常变化。目标 2，我将检验以下假设：APOE4 星形胶质细胞过度的促炎反应，引发了全身炎症反应 (LPS) 是通过 ERK 信号传导介导的。小鼠和人类脑组织中的蛋白质组学和转录组学数据以及验证 IHC 方法，我将识别体内 APOE 4/4 和 3/3 星形胶质细胞反应状态的分子变化并识别 ERK- 我的工作将为进一步研究奠定基础。星形胶质细胞介导的 AD 发病机制，在未来的研究中采用 CIBOP 来探索细胞类型 AD 和星形胶质细胞之外的神经变性的具体机制。