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The Role of KCa3.1 in Microglial function and in Parkinsons disease pathogenesis

KCa3.1 在小胶质细胞功能和帕金森病发病机制中的作用

基本信息

批准号：
10445079
负责人：
ARTHI KANTHASAMY
金额：
$ 38.41万
依托单位：
UNIVERSITY OF GEORGIA
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：
2021-07-15 至 2026-04-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10445079
关键词：
Ablation Address Adult Affect Autopsy Biological Assay Biological Availability Brain Brain Diseases CRISPR/Cas technology Calcium Calcium Signaling Calcium-Activated Potassium Channel Cells Clinical Trials Development Disease Disease Progression Electrophysiology (science)Exhibits Experimental Models FYN gene Functional disorder Genes Homeostasis Human Image Immune Impairment In Situ Hybridization Inflammation Mediators Knockout Mice Lead Link Mediating Microglia Midbrain structure Mission Molecular Mus National Institute of Neurological Disorders and Stroke Natural Immunity Nerve Degeneration Neurodegenerative Disorders Oral Parkinson Disease Parkinsonian Disorders Pathogenesis Pathology Patients Penetrance Pharmacology Phenotype Play Potassium Potassium Channel Pre-Clinical Model Process Progressive Disease RNA Research Role STAT1 gene Safety Signal Transduction Societies Testing Therapeutic Time Transcript Transgenic Organisms Treatment Efficacy United States National Institutes of Health Up-Regulation alpha synuclein attenuation base conditional knockout immunomodulatory therapies in vivo inflammatory modulation inhibitor innovation insight interest mouse model neurobehavioral neuroinflammation neuron loss neuroprotection neurotoxicity new therapeutic target novel novel therapeutic intervention pre-clinical progressive neurodegeneration protein expression response socioeconomics synucleinopathy therapeutic evaluation transcriptomics

项目摘要

Abstract Recent studies have begun to uncover the central role of microglia-mediated neuroinflammation in Parkinson’s disease (PD) pathogenesis. Increasing evidence suggests that microglia-driven innate immunity could further potentiate deleterious α-synuclein (αSyn) aggregation and progressive neurodegeneration. However, we lack an in-depth understanding of the cellular mechanisms regulating αSyn-induced innate immunity. Therefore, identifying signaling mechanisms that regulate microglial function in response to Parkinsonian pathology may lead to the development of novel immunomodulatory therapies for PD. We recently discovered that the transcript and protein expression levels of the calcium-activated potassium channel KCa3.1, best known for its role in immune cell calcium signaling, are elevated in activated microglia in both postmortem PD brains and in preclinical models of PD. We further identified that disruption of either FYN or STAT1 dampens reactive microglia activation responses via modulation of inflammatory mediators in aggregated αSyn (αSynagg)-stimulated primary microglia. Importantly, the highly selective and orally active KCa3.1 inhibitor Senicapoc reduced neuroinflammation and nigral dopamin(DA)ergic neurotoxicity in a preclinical mouse model of PD, suggesting that KCa3.1 plays a multifaceted role by governing disease pathology. Despite these encouraging findings, the exact cellular mechanisms by which KCa3.1 regulates microglial function in the context of synucleinopathy remain poorly characterized. Herein, we propose three integrated aims to test the central hypothesis that KCa3.1 promotes αSynagg-mediated progressive nigral DAergic neurodegenerative processes via activation of the microglial Fyn- STAT1 signaling axis and that the in vivo inhibition of KCa3.1 restores microglial homeostasis and affords DAergic neuroprotection in the context of synucleinopathy. In Aim-1, we will test the hypothesis that upregulation of KCa3.1 induces the proinflammatory microglial activation phenotype and nigral DAergic neuronal loss in the context of synucleinopathy. In Aim-2, we will test the hypothesis that the Fyn-STAT1 signaling axis drives microglial responses to PD-like pathology in a KCa3.1-dependent manner. In Aim-3, we will test the hypothesis that inhibiting KCa3.1 activation is efficacious in reducing reactive microglial activation and progressive PD-like disease pathology. The proposed studies are innovative, utilizing a combination of transcriptomic profiling, RNA in situ hybridization (ISH), imaging analysis, the RT QuIC assay for αSynagg seeding, CRISPR/Cas9 KCNN4 knockout (KO) mice, transgenic conditional KO mouse models, and electrophysiological recordings to test how microglial KCa3.1 influences progressive neurodegenerative processes in PD. These studies address key mechanistic aspects regarding the functional roles of KCa3.1 in PD pathogenesis and may aid in the identification of new molecular determinants that can be targeted for slowing or halting PD progression and/or repurposing Senicapoc for PD therapy. 1

摘要最近的研究已经开始揭示小胶质细胞介导的神经炎症在帕金森病中的核心作用疾病（PD）发病机制。越来越多的证据表明，小胶质细胞驱动的先天免疫可以进一步增强有害的α-突触核蛋白（αSyn）聚集和进行性神经变性。然而，我们缺乏一个深入了解调节αSyn-induced先天免疫的细胞机制。因此，我们认为，识别响应帕金森病病理学调节小胶质细胞功能的信号传导机制，导致PD的新型免疫调节疗法的发展。我们最近发现和蛋白质表达水平的钙激活钾通道KCa3.1，最有名的作用，免疫细胞钙信号传导，在死后PD脑和临床前PD脑中的活化小胶质细胞中升高。 PD模型我们进一步确定，FYN或STAT 1的破坏抑制了反应性小胶质细胞的激活，通过调节聚集的αSyn（αSynagg）刺激的原代小胶质细胞中的炎症介质的反应。重要的是，高选择性和口服活性的KCa3.1抑制剂Senicapoc减少了神经炎症，黑质多巴胺（DA）能神经毒性在PD的临床前小鼠模型中，表明KCa 3.1起作用，多方面的作用，通过管理疾病病理。尽管有这些令人鼓舞的发现，在突触核蛋白病的背景下，KCa3.1调节小胶质细胞功能的机制仍然很差表征了在此，我们提出了三个综合目标来检验KCa3.1促进 α Synag通过激活小胶质细胞Fyn介导的进行性黑质DA能神经退行性过程 STAT 1信号转导轴和KCa3.1的体内抑制恢复小胶质细胞稳态，突触核蛋白病背景下DA能神经保护作用。在Aim-1中，我们将检验上调 KCa3.1的诱导促炎性小胶质细胞活化表型和黑质DA能神经元损失， synucleinopathy突触核蛋白病在Aim-2中，我们将检验Fyn-STAT 1信号轴驱动小胶质细胞以KCa3.1依赖性方式对PD样病理学的反应。在Aim-3中，我们将检验假设抑制KCa3.1活化在减少反应性小胶质细胞活化和进行性PD样疾病病理学拟议的研究是创新的，利用转录组学分析，RNA 原位杂交（ISH）、成像分析、αSynagg接种的RT QuIC测定、CRISPR/Cas9 KCNN 4 基因敲除（KO）小鼠、转基因条件性KO小鼠模型和电生理记录，以测试小胶质细胞KCa 3.1影响PD中的进行性神经变性过程。这些研究解决了关于KCa3.1在PD发病机制中的功能作用的机制方面，并可能有助于识别新的分子决定因素，可以靶向减缓或停止PD进展和/或再利用 Senicapoc用于PD治疗。 1