Using Peripheral Microglial Exosomes to predict brain inflammation in the human Parkinson’s brain

使用外周小胶质细胞外泌体预测人类帕金森病大脑的炎症

• 批准号: 10665477
• 负责人: Diego F Mastroeni
• 金额: $ 28.19万
• 依托单位: ARIZONA STATE UNIVERSITY-TEMPE CAMPUS
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-18 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10665477
• 关键词: Address; Affect; Anti-Inflammatory Agents; Antibodies; Antigens; Binding Proteins; Biological Markers; Blood Circulation; Brain; Brain Diseases; Brain region; Cell Nucleus; Cell membrane; Cells; Central Nervous System; Cerebrospinal Fluid; Chemicals; Clinical; Communication; Complement; Data; Diagnosis; Disease; Disease Progression; Disease associated microglia; Encapsulated; Encephalitis; Etiology; Event; Family; Genes; Human; Immune; Individual; Infection; Inflammation; Inflammatory; Institution; Leukocytes; Life; Membrane; Membrane Proteins; Messenger RNA; Meta-Analysis; Methods; MicroRNAs; Microglia; Molecular; Molecular Sieve Chromatography; Mononuclear; Multivesicular Body; Nerve Degeneration; Neurobiology; Neurodegenerative Disorders; Parents; Parkinson Disease; Pathogenesis; Pathologic; Pathway interactions; Patients; Peripheral; Peripheral Blood Mononuclear Cell; Pharmaceutical Preparations; Physiological; Physiological Processes; Play; Publishing; RNA; Role; Sampling; Secure; Serum; Signaling Molecule; Stimulus; Substantia nigra structure; TNF gene; Testing; Therapeutic; Time; Tissues; Work; blood-brain barrier crossing; cell type; clinical diagnosis; comorbidity; cytokine; density; design; differential expression; disorder risk; exosome; extracellular; extracellular vesicles; fighting; glial activation; human data; immune stimulant; liquid chromatography mass spectrometry; member; monocyte; neuroinflammation; novel; particle; protective effect; response; tool; transcriptome sequencing; treatment strategy; vesicular release

In this proposal, we will determine if we can use microglial extracellular vesicles (EV) that have been shed into the cerebral spinal fluid (CSF) and serum to predict the microglial activation state in the Parkinson’s brain (PD). Currently, a major obstacle in the field is that there are no established and validated methods to detect brain inflammation in response to neurodegeneration during life. Microglia, the resident immune cell of the CNS constantly patrol the brain, looking for signs of infection or inflammation caused by a host of immune stimulants. The role of microglia is to clear potential threats to the CNS, but their chemical signatures based on their presence are continuously released (cytokines). These cytokines activate neighboring microglia initiating a cascade of events that are believed to drive disease pathogenesis. Although cytokines are generally considered to function as soluble molecules, recent efforts have shown that cytokines are encapsulated in EVs. These EVs contain a host of inflammatory-associated mRNAs that encode cytokine-associated genes among other signaling molecules that are known to reflect the physiological state of the parent cell. Unlike microglia, EVs can cross the blood-brain barrier under leaky and inflamed conditions, both of which are known physiological processes in PD. To determine if we can identify microglial EVs in the periphery we will characterize EVs isolated from disease-associated microglia (DAM) using RNAseq and Liquid chromatography-mass spectrometry (LC/MS). This novel data will be used to generate targets for discovery work in the periphery. This proposal aims to address two unmet needs 1) the possibility to detect brain inflammation in the living and 2) the ability to distinguish EVs released from microglia from those released from peripheral blood mononuclear cells. We hypothesize that microglial EVs extracted from the serum/CSF will reflect EV profiles from DAM. To test our hypothesis, we propose to 1) Analyze membrane-bound proteins and EV cargo in disease-associated microglia (DAM), 2) analyze membrane-bound proteins and EV cargo in peripheral EVs isolated from PD peripheral blood mononuclear cells, 3) identify and validate unique microglial-specific EV membrane antigens for antibody discovery work and 4) use these validated antibodies to pull down microglial-specific EVs from CSF and serum. All the tissues proposed in the application were collected from premortem PD patients who have since passed and have pathological confirmation of disease without comorbidities. We believe that each of these individual aims is high impact, and novel on their own, but collectively could provide us the tools necessary to predict brain inflammation.

在这项提案中，我们将确定是否可以使用小胶质细胞外囊泡(EV)，这些小胶质细胞已经进入 脑脊液(CSF)和血清以预测帕金森病脑(PD)的小胶质细胞激活状态。 目前，该领域的一个主要障碍是没有建立和验证的方法来检测大脑 在生活中对神经退化作出反应的炎症。中枢神经系统的常驻免疫细胞--小胶质细胞 不断地巡视大脑，寻找由大量免疫兴奋剂引起的感染或炎症迹象。 小胶质细胞的作用是清除对中枢神经系统的潜在威胁，但它们的化学特征基于它们的 存在不断地释放(细胞因子)。这些细胞因子激活邻近的小胶质细胞，启动 被认为是推动疾病发病机制的一连串事件。尽管细胞因子通常被认为是 为了发挥可溶性分子的作用，最近的研究表明，细胞因子被包裹在电动汽车中。这些电动汽车 包含大量炎症相关的mRNAs，编码细胞因子相关基因和其他信号 已知能反映亲本细胞生理状态的分子。与小胶质细胞不同，电动汽车可以穿越 血脑屏障在渗漏和炎症条件下，这两个都是已知的生理过程 警察。为了确定我们是否能在外周识别出小胶质细胞EV，我们将对从 用RNAseq和液质联用(LC/MS)检测疾病相关小胶质细胞(DAM)。 这种新颖的数据将被用来为外围的发现工作生成目标。这项提案旨在解决 两个未得到满足的需求：1)检测活人脑部炎症的可能性；2)辨别EV的能力 从外周血单核细胞释放的小胶质细胞释放的。我们假设 从血清/脑脊液中提取的小胶质细胞EV将反映DAM的EV特征。为了检验我们的假设，我们 建议1)分析疾病相关小胶质细胞(DAM)中的膜结合蛋白和EV货物，2) 帕金森病患者外周血中EV的膜结合蛋白和EV载量分析 单个核细胞，3)鉴定和验证独特的小胶质细胞特异性EV膜抗原抗体 发现工作和4)使用这些经过验证的抗体从脑脊液和血清中提取小胶质细胞特异性EV。 申请中提出的所有组织都是从死前帕金森病患者身上收集的，这些患者后来去世了 并有疾病病理证实，无合并症。我们相信这些人中的每一个 AIMS本身影响很大，而且很新颖，但它们共同可以为我们提供预测大脑所需的工具 发炎。