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Probing Heterogeneity of Alzheimer's Disease Using iPSCs

使用 iPSC 探索阿尔茨海默病的异质性

基本信息

批准号：
10657140
负责人：
Tracy L YOUNG-PEARSE
金额：
$ 85.52万
依托单位：
BRIGHAM AND WOMEN'S HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-07-15 至 2028-04-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10657140
关键词：
Address Adult Alzheimer&apos s Disease Alzheimer&apos s disease brain Alzheimer&apos s disease risk Amyloid beta-Protein Astrocytes Biology Brain CASP1 gene CD36 gene CRISPR/Cas technology Cells Coculture Techniques Cognitive Constitution Constitutional Data Detergents Development Disease Disease Progression Environment Enzyme-Linked Immunosorbent Assay Exhibits Experimental Models Formic Acids Functional disorder Genes Genetic Transcription Genetic study Genome engineering Health Heterogeneity Heterozygote Human IL18 gene Immune Immune signaling Impairment Individual Induced pluripotent stem cell derived neurons Inflammasome Inflammation Inflammatory Interleukin-1 beta Interleukin-13 Knockout Mice Late Onset Alzheimer Disease Length Link Literature Lysosomes Measurement Medial Mediating Membrane Microglia Modeling Molecular Nerve Degeneration Neurodegenerative Disorders Neurofibrillary Tangles Neuroimmunomodulation Neurons Pathogenesis Pathway interactions Phagocytosis Phosphoric Monoester Hydrolases Play Prefrontal Cortex Process Proteins Proteomics RNA Receptor Up-Regulation Regulation Reporting Risk Role Sampling Senile Plaques Series Signal Pathway Signal Transduction Stains Synapses System TNF gene Transgenic Mice Up-Regulation Variant Visualization Water Western Blotting Work abeta accumulation brain cell brain health brain tissue cell type cohort conditional knockout cytokine experimental study genetic risk factor genome wide association study human model induced pluripotent stem cell inhibitor inorganic phosphate insight mouse model neuroinflammation pharmacologic phosphoinositide-3,4,5-triphosphate phosphoinositide-3,4-bisphosphate receptor upregulation response scavenger receptor single nucleus RNA-sequencing tau Proteins tau-1 transcriptome sequencing

项目摘要

PROJECT SUMMARY There is an increasing focus on understanding the role of microglia in neurodegenerative disorders including late-onset Alzheimer’s disease (LOAD), a disease defined by the accumulation of amyloid beta rich plaques and neurofibrillary tangles containing tau. Microglia are proposed to play a variety of critical roles during disease progression including synaptic engulfment, cytokine release, and phagocytosis of amyloid beta (Aβ). Further, recent GWAS studies have implicated innate immune processes in LOAD, supporting the importance of understanding the neuroimmunological processes underlying the risk and progression of AD. All recent large- scale GWAS have identified SNPs at the INPP5D locus that are significantly associated with AD. INPP5D expression is largely restricted to microglial cells in the human adult brain and we have confirmed previous findings that RNA levels of INPP5D are elevated in AD brain. However, through quantitative western blotting, we show that protein levels of full length, water-soluble INPP5D are reduced in AD brain. To study the functional consequences of reduced INPP5D, we used both pharmacological inhibition and CRISPR-Cas9 genome engineering to lower INPP5D activity in human iPSC-derived microglia. Through unbiased RNA and proteomic profiling and a series of pharmacological manipulations, we demonstrated that reduction of INPP5D activity induces changes in immune signaling and, more specifically, the activation of the inflammasome. These studies have raised important questions that we aim to address in this proposal regarding the constitution of INPP5D in microglia in the Alzheimer’s brain (aim 1), the molecular mechanism(s) linking INPP5D and inflammasome activation (aim 2), and the functional consequences of loss of INPP5D activity and inflammasome activation in microglia on astrocytes and neurons (aim 3). In aim 1, we will utilize quantitative immunostaining, sequential extraction and western blotting, and ELISA to deeply interrogate INPP5D levels and markers of inflammasome activation across a large cohort of human brain samples and iPSC-derived microglia cultures. Aim 2 interrogates candidate pathways that may link INPP5D activity with inflammasome activation that arose from our preliminary analyses. The first of these involves upregulation of PLA2G7, second through dysregulation of scavenger receptors and the third through disruption of lysosome function. Finally, in aim 3 we utilize co-culture models of iPSC-derived neurons, astrocytes and microglia and a conditional INPP5D knock out mouse model to determine the consequences of reduction of INPP5D levels in microglia on neuron and astrocyte biology in the context of AD relevant environments. Together, these studies will provide important new insights into our understanding of the fundamental role of INPP5D in microglia in health and disease, regulation of inflammasome activation in human microglia, and the consequences of sub-lytic inflammasome activation in microglia on neuron and astrocyte biology.

项目概要人们越来越关注了解小胶质细胞在神经退行性疾病中的作用，包括晚发性阿尔茨海默氏病 (LOAD)，一种由富含 β 淀粉样蛋白的斑块积累和含有 tau 蛋白的神经原纤维缠结。小胶质细胞被认为在疾病过程中发挥多种关键作用进展包括突触吞噬、细胞因子释放和β淀粉样蛋白（Aβ）的吞噬作用。更远，最近的 GWAS 研究表明 LOAD 中存在先天免疫过程，支持了 LOAD 的重要性了解 AD 风险和进展背后的神经免疫过程。所有最近的大型- 大规模 GWAS 已确定 INPP5D 基因座上的 SNP 与 AD 显着相关。 INPP5D 表达主要限于成人大脑中的小胶质细胞，我们已经证实之前研究结果表明，AD 大脑中 INPP5D 的 RNA 水平升高。然而，通过定量蛋白质印迹，我们表明 AD 脑中全长水溶性 INPP5D 的蛋白质水平降低。来研究泛函 INPP5D 减少的后果，我们同时使用药理学抑制和 CRISPR-Cas9 基因组工程降低人 iPSC 衍生的小胶质细胞中的 INPP5D 活性。通过公正的RNA和蛋白质组学通过分析和一系列药理学操作，我们证明 INPP5D 活性的降低诱导免疫信号的变化，更具体地说，诱导炎症小体的激活。这些研究提出了我们旨在在有关 INPP5D 组成的提案中解决的重要问题阿尔茨海默病大脑中的小胶质细胞（目标 1），连接 INPP5D 和炎症小体的分子机制激活（目标 2），以及 INPP5D 活性丧失和炎症小体激活的功能后果星形胶质细胞和神经元上的小胶质细胞（目标 3）。在目标 1 中，我们将利用定量免疫染色、序贯分析提取和蛋白质印迹以及 ELISA 来深入探究 INPP5D 水平和炎症小体标记物大量人脑样本和 iPSC 衍生的小胶质细胞培养物的激活。目标2审问可能将 INPP5D 活性与我们初步产生的炎症小体激活联系起来的候选途径分析。第一个涉及 PLA2G7 的上调，第二个涉及清除剂的失调第三个是通过破坏溶酶体功能。最后，在目标 3 中，我们利用共培养模型 iPSC 衍生的神经元、星形胶质细胞和小胶质细胞以及条件性 INPP5D 敲除小鼠模型以确定小胶质细胞中 INPP5D 水平降低对神经元和星形胶质细胞生物学的影响 AD相关环境。总之，这些研究将为我们的理解提供重要的新见解 INPP5D 在小胶质细胞中在健康和疾病中的基本作用，炎症小体激活的调节人类小胶质细胞，以及小胶质细胞中亚裂解炎症小体激活对神经元和神经元的影响星形胶质细胞生物学。

项目成果

期刊论文数量（10）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

RNA-binding protein ELAVL4/HuD ameliorates Alzheimer's disease-related molecular changes in human iPSC-derived neurons.

DOI：
10.1016/j.pneurobio.2022.102316
发表时间：
2022-10
期刊：
PROGRESS IN NEUROBIOLOGY
影响因子：
6.7
作者：
van der Linden, Robert J.;Gerritsen, Jacqueline S.;Liao, Meichen;Widomska, Joanna;Pearse II, Richard V.;White, Forest M.;Franke, Barbara;Young-Pearse, Tracy L.;Poelmans, Geert
通讯作者：
Poelmans, Geert

Loss of endosomal exchanger NHE6 leads to pathological changes in tau in human neurons.

DOI：
10.1016/j.stemcr.2022.08.001
发表时间：
2022-09-13
期刊：
STEM CELL REPORTS
影响因子：
5.9
作者：
Fernandez, Marty A.;Bah, Fatmata;Ma, Li;Lee, YouJin;Schmidt, Michael;Welch, Elizabeth;Morrow, Eric M.;Young-Pearse, Tracy L.
通讯作者：
Young-Pearse, Tracy L.

Lost in translational biology: Understanding sex differences to inform studies of diseases of the nervous system.

迷失在转化生物学中：了解性别差异为神经系统疾病的研究提供信息。

DOI：
10.1016/j.brainres.2019.146352
发表时间：
2019
期刊：
Brain research
影响因子：
2.9
作者：
Pearse2nd,RichardV;Young-Pearse,TracyL
通讯作者：
Young-Pearse,TracyL

Mosaic loss of Chromosome Y in aged human microglia.

DOI：
10.1101/gr.276409.121
发表时间：
2022-10
期刊：
Genome research
影响因子：
7
作者：
Vermeulen MC;Pearse R;Young-Pearse T;Mostafavi S
通讯作者：
Mostafavi S

Small Molecule Regulators of microRNAs Identified by High-Throughput Screen Coupled with High-Throughput Sequencing.

通过高通量筛选结合高通量测序鉴定 microRNA 的小分子调节因子。

DOI：
10.21203/rs.3.rs-2617979/v1
发表时间：
2023
期刊：
Research square
影响因子：
0
作者：
Krichevsky,Anna;Nguyen,Lien;Wei,Zhiyun;Silva,M;Barberán-Soler,Sergio;Rabinovsky,Rosalia;Muratore,Christina;Stricker,Jonathan;Hortman,Colin;Young-Pearse,Tracy;Haggarty,Stephen
通讯作者：
Haggarty,Stephen