SURVIVAL AND DEATH OF NEURONS

神经元的生存和死亡

• 批准号: 6802530
• 负责人: Timothy J Timothy J Cunningham Cunningham
• 金额: $ 2.5万
• 依托单位: DREXEL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1980
• 资助国家: 美国
• 起止时间: 1980-07-01 至 2004-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6802530
• 关键词: bradykinin; calcium flux; calreticulin; cell death; cell line; central neural pathway /tract; cytoprotection; experimental brain lesion; immunologic assay /test; laboratory rat; neural degeneration; neurotransmitter transport; bradykinin; calcium flux; calreticulin; cell death; cell line; central neural pathway /tract; cytoprotection; experimental brain lesion; immunologic assay /test; laboratory rat; neural degeneration; neurotransmitter transport

This is an application for continued support for studies of cell death in the nervous system. The proposal is concerned mainly with the mechanism of action and the therapeutic potential of a cell survival peptide that was identified and synthesized during the previous period of support. This peptide, called Y-P30 because of its amino acid composition, has several interesting and potentially important properties: 1) it promotes the survival of cell lines in vitro and of cortical neurons in vivo after application to cerebral cortex lesions; 2) it inhibits the differentiation of a promonocyte cell line (HL-60) in vitro and CNS monocyte derivatives (macrophages/microglia) in vivo in the cortical lesion model; 3) it binds specifically to membrane-associated calreticulin, a rediscovered calcium binding protein that may be important for Ca signaling associated with integrins and other ligands, including those related to neurotransmission; 4) the peptide may destabilize the calreticulin at membranes via its phosphatase activity causing calreticulin release from cells; 5) intravenous-delivery of Y-P30 inhibits macrophages/microglia cells, promotes neuron survival, and causes calreticulin release from cortical cells in rats with cortical lesions, and in unoperated rats. A model has therefore evolved in which Y-P30 acts to inhibit aspects of the cascade of inflammation-neuron death that characterizes several disorders of primary neuronal degeneration. The proposed experiments will further explore mechanisms related to Y-P30 biological activities, especially as these relate to calreticulin function in the nervous system. First, Y-P30's effects on integrin-mediated functions will be tested in an in vitro chemotaxis chamber with brain and peritoneal macrophages. Second the effects of Y- P30 stimulated release of calreticulin on complement-mediated mechanisms (calreticulin as a Clq receptor homologue) will be determined in human neuroblastoma cells 5Y5Y. Third, Y-P 30 will be tested for its effects on Ca2+ handling in resting 5Y5Y cells and after stimulation with bradykinin. A major part of this research will involve further comparisons of direct application of Y-P30 to intravenous delivery in order to determine dosages and potential for long term and retrograde/transneuronal survival effects in the visual and somatosensory systems. Antibodies to the peptide and to brain calreticulin will be made for use in these studies as well as to investigate the possibility of endogenous peptide like activity in the CNS. It is expected that these experiments will provide further insights into mechanisms and potential therapies for neuron degeneration after CNS injury and in nervous system diseases.

这是对神经系统中细胞死亡研究的持续支持的应用。该提案主要涉及作用机理和细胞存活肽的治疗潜力，该肽在上一个支持期间被鉴定出和合成。该肽由于其氨基酸组成而称为Y-P30，具有几种有趣且潜在的重要特性：1）在应用于脑皮质病变后，它促进了体外和体内皮质神经元的存活； 2）在皮质病变模型中，它抑制体外和CNS单核细胞衍生物（巨噬细胞/小胶质细胞）的体外和CNS单核细胞衍生物（巨噬细胞/小胶质细胞）的分化； 3）它与膜相关的钙网蛋白专门结合，膜相关的钙网蛋白是一种重新发现的钙结合蛋白，可能对与整联蛋白和其他配体相关的CA信号传导很重要，包括与神经传递相关的配体； 4）肽可能通过其磷酸酶活性使膜上的钙网蛋白不稳定，从而导致细胞从细胞中释放出钙网蛋白； 5）Y-P30的静脉递送抑制巨噬细胞/小胶质细胞，促进神经元的存活，并导致具有皮质病变的大鼠皮质细胞和无手术大鼠在皮质细胞中释放的钙网蛋白。因此，一个模型进化出来，Y-P30起作用抑制炎症 - 神经元死亡的级联反应的各个方面，该死亡表征了原发性神经元变性的几种疾病。提出的实验将进一步探索与Y-P30生物活性有关的机制，尤其是这些机制与神经系统中的钙网蛋白功能有关。首先，Y-P30对整联蛋白介导的功能的影响将在与脑和腹膜巨噬细胞的体外趋化室中进行测试。其次，Y-P30刺激钙网蛋白释放对补体介导的机制（钙网蛋白作为CLQ受体同源物）的作用将在人类神经母细胞瘤细胞5Y5Y中确定。第三，将测试Y-P 30对静止的5Y5Y细胞和用心动激肽刺激后对Ca2+处理的影响。这项研究的主要部分将涉及将Y-P30直接应用于静脉输送的直接应用，以确定视觉和体感系统中的长期和逆行/跨性别生存效应的剂量和潜力。将在这些研究中使用对肽和脑钙网蛋白的抗体，并研究中枢神经系统中内源性肽的可能性。预计这些实验将进一步了解中枢神经系统损伤和神经系统疾病后神经元变性的机制和潜在疗法。