NOVEL EXOSOME BIOMARKERS OF IRON PATHOLOGY IN AD

AD 中铁病理学的新型外泌体生物标志物

基本信息

批准号：
10223789
负责人：
Utkan Demirci
金额：
$ 43.3万
依托单位：
STANFORD UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-05-01 至 2023-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10223789
关键词：
Address Alzheimer&apos s Disease Alzheimer&apos s disease brain Alzheimer&apos s disease pathology American Amyloid Antibodies Astrocytes Atlases Autopsy Biological Assay Biological Markers Biology Biomedical Engineering Blood Brain Carrier Proteins Cell Death Cell Surface Proteins Cell physiology Cell surface Cells Cellular biology Ceruloplasmin Contralateral Data Development Diagnosis Disease Disease Progression Electron Microscopy Electron energy loss spectroscopy Exhibits Ferritin Freezing Future Gene Expression Glutathione Goals Hippocampus (Brain)Homeostasis Human Image Immune Inductively Coupled Plasma Mass Spectrometry Inflammation Intervention Investigation Iron Knowledge Lead Link Lipids Literature Magnetic Resonance Imaging Mass Spectrum Analysis Measurable Measures Metabolic Pathway Metals Methods MicroRNAs Microglia Molecular Nerve Degeneration Neurodegenerative Disorders Neurons Nucleic Acids Oligodendroglia Oxidative Stress Pathologic Processes Pathology Pensions Pharmacologic Substance Play Prevalence Proteins Publishing RNA Raman Spectrum Analysis Reproducibility Roentgen Rays Role Specificity Specimen Synchrotrons TFRC gene Technology Tissues Transferrin Translating Translations Vesicle Western Blotting Work X ray microscopy base brain cell cell type cost disorder control divalent metal effective therapy efficacious treatment exosome extracellular vesicles hepcidin human tissue image reconstruction in vivo innovation insight intercellular communication iron metabolism iron oxidation machine learning algorithm metal transporting protein 1 multidisciplinary multimodality nanoscale neuropathology next generation sequencing novel novel therapeutics oxidation population based specific biomarkers tau Proteins tau aggregation transcriptome sequencing

项目摘要

PROJECT SUMMARY/ABSTRACT Alzheimer’s disease (AD) afflicts millions of Americans, yet no effective treatments exist. Iron has been shown to be involved in key AD pathologic processes, including amyloid and tau aggregation, inflammation, oxidative stress, and cell death mechanisms. Despite this growing evidence, it is challenging to ascertain alterations in iron metabolism in vivo, limiting potential translation to biomarkers and novel therapies. Exosomes are nanometer-sized vesicles shed by cells to transport proteins, nucleic acids, metals, lipids or metabolites. While exosomes reflect cellular processes and can reveal disease-related pathologies in human tissues and biofluids, iron abnormalities in AD exosomes have not yet been investigated. We will address this knowledge gap through state-of-the-art exosome isolation technology combined with advanced iron imaging, protein quantification and next generation sequencing methods. Our goal is to investigate iron dysregulation in exosomes from post- mortem AD brains, in order to unveil AD-specific biomarkers and facilitate the development of novel therapies. The project aims are: (1) To determine whether the quantity, oxidation state, and cellular origin of exosomal iron is altered in AD. Using MRI and synchrotron X-ray microscopy, we will quantify tissue iron content and oxidation state in human AD and control hippocampal specimens. We will then use our novel exosome isolation platform, ExoTIC, to isolate exosomes from regions of high hippocampal iron content in the same specimens. Using antibodies that target cell-surface proteins, we will enrich the isolated exosomes based on their cellular origin (e.g. neurons, microglia, etc.). We will quantify exosomal iron content from each cell type using mass spectrometry (ICP-MS), and measure exosomal iron oxidation state using electron microscopy. Taken together, we will determine whether iron content and oxidation state are altered in Alzheimer’s exosomes compared to controls, in particular in exosomes originating in microglia, the brain’s immune cells. (2) Detect dysregulation of iron-related proteins and RNAs in AD exosomes. Using Western blotting on the enriched exosomes, we will determine whether levels of proteins that play a role in iron metabolism are altered in AD compared to controls. Because exosomes are generally rich in microRNAs that are known to regulate gene expression, we will use RNA-Seq to determine whether exosomal microRNAs regulating these same iron-related proteins are also altered in AD. Machine learning algorithms will enable the creation of an atlas of microRNAs linking iron, iron-related proteins, and neuropathology, which should provide a deeper understanding of AD biology. Characterization of exosome content in the AD brain should result in cell-specific signatures of iron dysregulation associated with neurodegeneration. This approach may elucidate novel aspects of AD biology, lead to novel assays to detect early AD, and facilitate a much-needed future therapy.

项目概要/摘要阿尔茨海默病（AD）困扰着数百万美国人，但目前尚无有效的治疗方法。参与关键的 AD 病理过程，包括淀粉样蛋白和 tau 蛋白聚集、炎症、氧化尽管有越来越多的证据，但确定这些变化仍然具有挑战性。铁在体内的代谢，限制了外泌体向生物标志物和新疗法的潜在转化。细胞脱落的纳米大小的囊泡，用于运输蛋白质、酸、金属、脂质或代谢物。外泌体反映细胞过程，可以揭示人体组织和生物体液中与疾病相关的病理学， AD 外泌体中的铁异常尚未得到研究，我们将通过以下方式解决这一知识差距。最先进的外泌体分离技术与先进的铁成像、蛋白质定量和我们的目标是研究后外泌体中的铁失调。尸检 AD 大脑，以揭示 AD 特异性生物标志物并促进新疗法的开发。该项目的目标是：（1）确定外泌体的数量、氧化态和细胞来源是否使用 MRI 和同步加速器 X 射线显微镜，我们将量化组织铁含量和然后我们将使用我们的新型外泌体来检测人类 AD 和对照海马样本的氧化态。分离平台 ExoTIC，用于从同一海马铁含量高的区域分离外泌体使用针对细胞表面蛋白的抗体，我们将基于以下内容富集分离的外泌体：它们的细胞起源（例如神经元、小胶质细胞等）我们将量化每种细胞类型的外泌体铁含量。使用质谱（ICP-MS），并使用电子显微镜测量外泌体铁氧化态。综合起来，我们将确定铁含量和氧化态是否与阿尔茨海默病有关外泌体与对照相比，特别是源自小胶质细胞（大脑的免疫细胞）的外泌体。 (2) 使用蛋白质印迹法检测 AD 外泌体中铁相关蛋白和 RNA 的失调。富集的外泌体，我们将确定在铁代谢中发挥作用的蛋白质水平是否与对照组相比，AD 中的外泌体通常富含已知具有调节作用的 microRNA。基因表达，我们将使用 RNA-Seq 来确定外泌体 microRNA 是否调节这些相同的基因 AD 中与铁相关的蛋白质也发生了改变，机器学习算法将能够创建铁图谱。连接铁、铁相关蛋白和神经病理学的 microRNA，这应该提供更深入的研究了解 AD 生物学。 AD 大脑中外泌体含量的表征应该会产生铁的细胞特异性特征这种方法可能会阐明 AD 生物学的新方面。导致检测早期 AD 的新方法，并促进未来急需的治疗。