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：
10631159
负责人：
ARTHI KANTHASAMY
金额：
$ 38.31万
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/10631159
关键词：
Ablation Address Adult Affect Autopsy Biological Assay Biological Availability Brain Brain Diseases CRISPR/Cas technology Calcium Signaling Calcium-Activated Potassium Channel Cells Clinical Trials Development Disease Disease Progression Economic Burden Electrophysiology (science)Exhibits Experimental Models FYN gene Functional disorder Genes Homeostasis Human Image Immune Impairment In Situ Hybridization Inflammation Mediators Inflammatory Knockout Mice 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 Phenotype Play 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 conditional knockout glial activation 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 pharmacologic pre-clinical progressive neurodegeneration protein expression response socioeconomics synucleinopathy therapeutic evaluation transcriptomic profiling

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

抽象的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氏症中的核心作用疾病（PD）发病机理。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潜在有害α-突触核蛋白（αSyn）聚集和进行性神经退行性。但是，我们缺乏对细胞机制调节αSyn诱导的先天免疫的深入了解。所以，确定根据帕金森病病理学调节小胶质细胞功能的信号传导机制可能导致PD的新型免疫调节疗法的发展。我们最近发现成绩单钙激活钾通道KCA3.1的蛋白质表达水平，以其在其中的作用而闻名免疫细胞钙信号传导在验尸脑中的活化小胶质细胞和临床前均升高 PD的模型。我们进一步确定FYN或STAT1的破坏会抑制反应性小胶质细胞激活通过调节聚集的αSyn（αSynagg）刺激的原代小胶质细胞中炎症介质的反应。重要的是，高度选择性和口服的KCA3.1抑制剂senicapoc降低了神经炎症和 PD的临床前小鼠模型中的nigral多巴胺（DA）ERGIC神经毒性，表明KCA3.1扮演A 通过疾病病理学的多方面角色。尽管有这些令人鼓舞的发现，但确切的细胞 KCA3.1在突触核酸的背景下调节小胶质细胞功能的机制仍然很差特征。在此，我们提出了三个综合旨在测试KCA3.1促进的中心假设 αSynagg介导的进行性杀伤性神经退行性过程通过小胶质细胞激活 STAT1信号轴以及KCA3.1的体内抑制可恢复小胶质体稳态，并提供在突触核酸的背景下的Daergic神经保护作用。在AIM-1中，我们将测试上调的假设 KCA3.1诱导促炎的小胶质细胞激活表型和nigral daerronal损失突触核苷的背景。在AIM-2中，我们将测试Fyn-STAT1信号轴的假设小胶质细胞对PD样病理的反应以KCA3.1依赖性方式。在AIM-3中，我们将检验假设抑制KCA3.1激活在减少反应性小胶质细胞激活和进行性PD样的方面有效疾病病理学。提出的研究是创新的，使用了转录组分析，RNA 原位杂交（ISH），成像分析，用于αSynagg播种的RT QUIC测定，CRISPR/CAS9 KCNN4 敲除（KO）小鼠，转基因条件KO小鼠模型和电生理记录，以测试如何小胶质细胞KCA3.1影响PD中的渐进神经退行性过程。这些研究针对关键关于KCA3.1在PD发病机理中的功能作用的机械方面，并可能有助于鉴定可以针对放大或停止PD进展和/或重新利用的新分子确定剂 SENICAPOC用于PD疗法。 1