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.

总结 帕金森氏病（PD）是一种使人衰弱的进行性退行性运动障碍， 黑质（SN）多巴胺（DA）能神经元沿着α-突触核蛋白（α-syn）的蓄积， 脑，小胶质细胞的活化，炎性细胞因子/趋化因子的产生，CD 4 + T细胞的浸润， 和神经退化最有效的治疗，左旋多巴，不能阻止疾病的进展， 渐进性的机制尚不清楚。钙蛋白酶是一种受钙调节的半胱氨酸蛋白酶， 在PD的SN和SC（脊髓）变性中的关键作用，以及其在α-syn聚集、 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中至关重要， 疾病进展。虽然钙蛋白酶-1切割α-syn促进PD样疾病中的突触核蛋白聚集， 两种主要的钙蛋白酶亚型，钙蛋白酶-1和钙蛋白酶-2，在α-syn呈递中的精确参与， CD 4 + T细胞仍然未知。初步研究在MPTP小鼠中鉴定了一种CD 4 + T细胞亚型， 钙蛋白酶抑制剂可消除这种抑制作用，这表明钙蛋白酶和CD 4 + T细胞的激活可能起作用。 在PD的炎症过程和疾病进展中起关键作用。初步数据还表明， siRNA介导的钙蛋白酶-2敲低减少人B细胞的抗原呈递并抑制 活化CD 4 + T细胞。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-reactive CD 4 + T细胞亚型的扩增我们还假设钙蛋白酶 抑制可以减弱α-syn聚集和炎性T细胞的扩增，减少炎症， 支持PD患者的神经元存活和改善的结果。提出了两个具体目标，以测试 假设：（目的1）研究calpain-1或calpain-2的激活是否与小胶质细胞 α-syn向CD 4 + T细胞呈递导致PD样疾病进展和神经退行性变 老鼠的疾病(Aim 2）确定抑制钙蛋白酶-2是否减少α-syn的小胶质细胞呈递 和致病性CD 4 + T细胞的产生，减弱PD小鼠模型中的疾病进展。目标 本研究的目的是探讨钙蛋白酶-1和钙蛋白酶-2在产生α-syn-reactive致病性CD 4+中的作用。 T细胞，以及来自MPTP小鼠的CD 4 + T细胞亚群是否可以诱导PD样疾病。 免疫缺陷小鼠研究计划确定是否删除钙蛋白酶-1减弱α-syn 使用钙蛋白酶-1敲除（KO）小鼠的CD 4 + T细胞的聚集和扩增。此外， 钙蛋白酶-2抑制剂将在炎性CD 4 + T细胞应答改变的研究中进行评估， 在钙蛋白酶-1 KO小鼠中细胞因子/趋化因子的产生。测试不同的钙蛋白酶同种型在细胞中的作用， α-syn-reactive致病性T细胞的产生和诱导神经元死亡和变性可能导致 开发用于治疗PD以及其他神经退行性疾病的新方法。