Mechanisms of Dantrolene Neuroprotection in Alzheimer's Disease

丹曲林对阿尔茨海默病的神经保护机制

• 批准号: 10227272
• 负责人: HUAFENG WEI
• 金额: $ 33.28万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-03-01 至 2023-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10227272
• 关键词: 2019-nCoV; ACE2; Address; Affect; Alzheimer' s Disease; Alzheimer' s disease patient; Apoptosis; Binding; COVID-19; COVID-19 patient; COVID-19 treatment; Calcineurin; Cathepsin L; Cathepsins; Cell Death; Cell Nucleus; Cell membrane; Cells; Cessation of life; Clinical; Clinical Research; Cytosol; Dantrolene; Data; Dose; Endocytosis; Endoplasmic Reticulum; Endosomes; Epithelial; Exposure to; Fibroblasts; Functional disorder; Future; Genetic Transcription; Goals; Grant; Homeostasis; Human; Impairment; Infection; Kidney; Knock-in; Lysosomes; Mediating; Membrane; Mitochondria; Modeling; Mutation; Neurons; Nuclear Pore Complex; Oxygen; Oxygen Consumption; Parents; Pathologic; Pathway interactions; Patients; Pharmaceutical Preparations; Pharmacotherapy; Play; Process; Production; Proteins; RNA replication; Reactive Oxygen Species; Reporting; Risk Factors; Role; Ryanodine Receptor Calcium Release Channel; Ryanodine Receptors; SARS-CoV-2 infection; SARS-CoV-2 inhibitor; SARS-CoV-2 spike protein; Skin; Stress; Testing; Toxic effect; Transfection; Virus; Virus Diseases; Virus Replication; aged; cell injury; cytokine; cytokine release syndrome; drug repurposing; efficacy evaluation; endoplasmic reticulum stress; experimental study; induced pluripotent stem cell; inhibitor/antagonist; neuroblastoma cell; neurogenesis; neuroprotection; nuclear factors of activated T-cells; pandemic disease; presenilin-1; prevent; receptor; sensor; side effect; stem cells; synaptogenesis; translational study; viral RNA

Abstract Coronavirus disease 2019 (COVID-19), a pandemic affecting millions of patients around the world, is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) with no effective drug treatments. Both infection and transfection of virus in host cells require abnormally elevated Ca2+ concentrations in cytosol and endosome via overactivation of ryanodine receptors (RyRs) Ca2+ channel located on the membrane of the endoplasmic reticulum (ER). Dantrolene, an antagonist of RyRs, has been demonstrated to inhibit SARS- CoV-2-mediated host cell toxicity and damage. In the presence of RyRs overactivation and excessive and abnormal elevation of cytosolic and endosome Ca2+ concentrations, the spike proteins (S1, S2) of the virus play important roles in binding, fusion, and virus replication in the host cell, eventually leading to cell damage or death. Our long-term goal is to examine the efficacy and mechanisms of dantrolene to treat AD. The overall objective of this study is to investigate the effects and underlying mechanisms of dantrolene to protect against host cell damage or death induced by SARS-CoV-2 spike proteins-mediated overactivation of RyRs and associated Ca2+ dysregulation. Our central hypothesis supported by preliminary data is that dantrolene inhibits SARS-CoV-2 spike proteins-mediated cell damage by inhibiting the overactivation of RyRs and by restoring intracellular Ca2+ homeostasis in AD cells. We will test this hypothesis with the following specific aims. Specific Aim 1 (SA1). To determine the effects of dantrolene on SARS-CoV-2 spike (S) proteins-mediated Ca2+ dysregulation and cell damage by apoptosis in AD cells using induced pluripotent stem cells (iPSC), from skin fibroblasts of patients with either sporadic (SAD) or familial (FAD), and human neuroblastoma cells (SH-SY5Y), with knocked-in AD presenilin 1 mutation (M146L). Specific Aim 2 (SA2). To determine the effects and mechanisms of dantrolene on SARS-CoV-2 spike (S) proteins-mediated pathological pathways leading to host cell damage. We will examine the effects of S proteins on mitochondria oxygen consumption, ATP production, reactive oxygen species (ROS) production, ER stress, and cytokine release. We will correlate the results between SA1 and SA2. We expect that dantrolene will inhibit S proteins-mediated overactivation of RyRs and associated disruption of intracellular Ca2+ homeostasis and cell death by apoptosis.

摘要 2019冠状病毒病（COVID-19），一场影响数百万患者的大流行病 严重急性呼吸道综合征冠状病毒2型（SARS-CoV-2）引起的全球范围内， 有效的药物治疗。病毒在宿主细胞中的感染和转染都需要 细胞质和核内体中Ca 2+浓度异常升高，通过过度激活 Ryanodine receptor（RyRs）是位于内质网膜上的钙通道 网织膜（ER）。丹曲林是RyRs的拮抗剂，已被证明可以抑制SARS- CoV-2介导的宿主细胞毒性和损伤。在RyRs过度激活的情况下， 胞浆和内体Ca 2+浓度过度和异常升高， 病毒的蛋白质（S1，S2）在结合、融合和病毒复制中起重要作用。 宿主细胞，最终导致细胞损伤或死亡。我们的长远目标是研究 丹曲林治疗AD疗效及机制本研究的总体目标是 研究丹曲林对宿主细胞的保护作用及其机制 SARS-CoV-2刺突蛋白介导的RyR过度激活诱导的损伤或死亡， 相关的Ca 2+失调。我们的中心假设得到了初步数据的支持， 丹曲林通过抑制SARS-CoV-2刺突蛋白介导的细胞损伤， RyR过度激活和恢复AD细胞内Ca 2+稳态。我们 我将以下列具体目标来检验这一假设。具体目标1（SA 1）。以确定 丹曲林对SARS-CoV-2刺突蛋白介导的Ca 2+失调的影响 以及使用诱导多能干细胞（iPSC）在AD细胞中通过凋亡引起的细胞损伤， 散发性（SAD）或家族性（FAD）患者的皮肤成纤维细胞，以及人 神经母细胞瘤细胞（SH-SY 5 Y），具有敲入的AD早老素1突变（M146 L）。具体 目标2（SA 2）。探讨丹曲林对SARS冠状病毒2型的作用及其机制 刺突（S）蛋白介导的导致宿主细胞损伤的病理途径。我们将 研究S蛋白对线粒体耗氧量，ATP产生， 活性氧（ROS）产生、ER应激和细胞因子释放。我们将把 SA 1和SA 2之间的结果。我们预期丹曲洛林将抑制S蛋白介导的 RyR的过度激活和相关的细胞内Ca 2+稳态和细胞内Ca 2+稳态的破坏 凋亡死亡。