Calpain cleavage of α-synuclein and T-cell reactivity in Parkinson’s disease

帕金森病中α-突触核蛋白的钙蛋白酶裂解和 T 细胞反应性

• 批准号: 10042307
• 负责人: NAREN L BANIK
• 金额: $ 41.11万
• 依托单位: MEDICAL UNIVERSITY OF SOUTH CAROLINA
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-05-15 至 2024-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10042307
• 关键词: Antigen Presentation; Asses; Attenuated; Autopsy; B-Lymphocytes; BCL2 gene; Biological Markers; Bone Marrow; Brain; CD4 Positive T Lymphocytes; Calcium; Calpain; Caspase; Cell Death; Cells; Cytoprotection; Data; Development; Disease; Disease Progression; Dopaminergic Cell; Flow Cytometry; Generations; Goals; Human; Immunodeficient Mouse; Immunohistochemistry; In Vitro; Infiltration; Inflammation; Inflammatory; Knock-out; Knockout Mice; Lead; Levodopa; Link; Mediating; Microglia; Movement Disorders; Mus; Nature; Nerve Degeneration; Neurodegenerative Disorders; Neuroglia; Neurons; Parkinson Disease; Pathogenicity; Patients; Peripheral; Pharmacology; Plasma; Play; Process; Production; Protein Isoforms; Rag1 Mouse; Research; Role; Small Interfering RNA; Spinal Cord; Substantia nigra structure; T cell response; T-Cell Activation; T-Lymphocyte; TdT-Mediated dUTP Nick End Labeling Assay; Testing; Tissues; Western Blotting; alpha synuclein; attenuation; calpain inhibitor; chemokine; cytokine; dopaminergic neuron; functional outcomes; improved; improved outcome; inhibitor/antagonist; knock-down; macrophage; migration; mouse model; neuron loss; neuronal survival; novel strategies; novel therapeutics; survival outcome; synuclein

Summary Parkinson's disease (PD) is a debilitating progressive degenerative movement disorder associated with loss of dopaminergic (DA) neurons in the substantia nigra (SN) along with the accumulation of α-synuclein (α-syn) in the brain, activation of microglia, production of inflammatory cytokines/chemokines, infiltration of CD4+ T-cells, and neurodegeneration. The most potent therapy, L-dopa, does not block disease progression, and the mechanism of the progressive nature is unclear. Calpain, a cysteine protease regulated by calcium, plays a pivotal role in SN and SC (spinal cord) degeneration in PD, and its role in α-syn aggregation, activation of microglia, T cells and their migration indicate calpain to be crucial in promoting the inflammatory process and disease progression. While calpain-1 cleavage of α-syn promotes synuclein aggregation in PD-like diseases, the precise involvement of the two major calpain isoforms, calpain-1 and calpain-2, in α-syn presentation to CD4+ T-cells remains unknown. Preliminary studies here identified a subtype of CD4+ T cells in MPTP mice, which was abolished by calpain inhibitor, suggesting that activation of calpain and CD4+ T cells may play critical roles in the inflammatory process and disease progression in PD. Preliminary data also suggest that siRNA-mediated knockdown of calpain-2 diminishes antigen presentation by human B-cells and inhibits activation of CD4+ T cells. Thus, we hypothesize that activation of distinct calpain isoforms may favor expansion of a subtype of α-syn-reactive CD4+ T cells in PD-like disease. We also hypothesize that calpain inhibition may attenuate α-syn aggregation and expansion of inflammatory T cells, reduce inflammation and support neuronal survival and improved outcome in PD patients. Two specific aims are proposed to test the hypothesis: (Aim 1) To investigate whether activation of calpain-1 or calpain-2 is linked with microglial presentation of α-syn to CD4+ T cells resulting in disease progression and neurodegeneration in PD-like disease in mice. (Aim 2) To determine whether inhibition of calpain-2 reduces microglial presentation of α-syn and generation of pathogenic CD4+ T cells, attenuating disease progression in mouse models of PD. The goal of this study is to investigate the role of calpain-1 and calpain-2 in generating α-syn-reactive pathogenic CD4+ T cells, and whether a subpopulation of CD4+ T cells from MPTP mice can induce PD-like disease in immunodeficient mice. Studies are planned to determine whether deletion of calpain-1 attenuates α-syn aggregation and expansion of CD4+ T cells using calpain-1 knockout (KO) mice. In addition, the role of calpain-2 inhibitor will be assessed in the study of alteration of inflammatory CD4+ T cell response and production of cytokines/chemokines in calpain-1 KO mice. Testing the effects of distinct calpain isoforms in the generation of α-syn-reactive pathogenic T cells and induction neuronal death and degeneration may lead to development of a novel approach for treating PD as well as other neurodegenerative disorders.

概括 帕金森病 (PD) 是一种使人衰弱的进行性退行性运动障碍，与丧失能力有关 黑质 (SN) 中的多巴胺能 (DA) 神经元以及 α-突触核蛋白 (α-syn) 的积累 大脑、小胶质细胞的激活、炎症细胞因子/趋化因子的产生、CD4+ T 细胞的浸润、 和神经退行性变。最有效的治疗方法是左旋多巴，但它不能阻止疾病进展，而且 渐进性质的机制尚不清楚。钙蛋白酶是一种受钙调节的半胱氨酸蛋白酶，在 在 PD 的 SN 和 SC（脊髓）变性中起关键作用，及其在 α-syn 聚集、激活 小胶质细胞、T 细胞及其迁移表明钙蛋白酶在促进炎症过程和 疾病进展。虽然 calpain-1 裂解 α-syn 会促进 PD 样疾病中的突触核蛋白聚集， 两种主要钙蛋白酶异构体 calpain-1 和 calpain-2 精确参与 α-syn 呈递 CD4+ T 细胞仍然未知。初步研究在 MPTP 小鼠中鉴定出了 CD4+ T 细胞的一种亚型， 被钙蛋白酶抑制剂消除，表明钙蛋白酶和 CD4+ T 细胞的激活可能发挥作用 在 PD 的炎症过程和疾病进展中发挥关键作用。初步数据还表明 siRNA 介导的 calpain-2 敲低可减少人类 B 细胞的抗原呈递并抑制 CD4+ T 细胞的激活。因此，我们假设不同的钙蛋白酶亚型的激活可能有利于 PD 样疾病中 α-syn 反应性 CD4+ T 细胞亚型的扩增。我们还假设钙蛋白酶 抑制可能会减弱 α-syn 聚集和炎症 T 细胞的扩张，减少炎症和 支持神经元存活并改善 PD 患者的预后。提出了两个具体目标来测试 假设：（目标 1）研究 calpain-1 或 calpain-2 的激活是否与小胶质细胞相关 α-syn 呈递给 CD4+ T 细胞，导致 PD 样疾病进展和神经变性 小鼠疾病。 （目标 2）确定抑制 calpain-2 是否会减少小胶质细胞 α-syn 的呈递 和致病性 CD4+ T 细胞的产生，减轻 PD 小鼠模型的疾病进展。目标 本研究的目的是研究 calpain-1 和 calpain-2 在产生 α-syn 反应性致病性 CD4+ 中的作用 T 细胞，以及来自 MPTP 小鼠的 CD4+ T 细胞亚群是否可以诱导 PD 样疾病 免疫缺陷小鼠。计划进行研究以确定删除 calpain-1 是否会减弱 α-syn 使用 calpain-1 敲除 (KO) 小鼠进行 CD4+ T 细胞的聚集和扩增。此外，作用 calpain-2 抑制剂将在炎症 CD4+ T 细胞反应改变的研究中进行评估 calpain-1 KO 小鼠细胞因子/趋化因子的产生。测试不同钙蛋白酶亚型的影响 α-syn反应性致病性T细胞的产生和诱导神经元死亡和变性可能导致 开发治疗帕金森病和其他神经退行性疾病的新方法。