Metalloproteinase Regulation of Neuronal Death

神经元死亡的金属蛋白酶调节

• 批准号: 6706465
• 负责人: Lee Anna Cunningham
• 金额: $ 27.59万
• 依托单位: UNIVERSITY OF NEW MEXICO
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-01-01 至 2008-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6706465
• 关键词: SDS polyacrylamide gel electrophoresis; apoptosis; biological signal transduction; cell membrane; central nervous system disorders; cerebral ischemia /hypoxia; enzyme activity; enzyme linked immunosorbent assay; gene expression; gene targeting; genetically modified animals; immunocytochemistry; immunoprecipitation; laboratory mouse; membrane proteins; metalloendopeptidases; neurons; neuroprotectants; polymerase chain reaction; stromelysin; terminal nick end labeling; tissue /cell culture; tissue inhibitor of metalloproteinases; ultrasound blood flow measurement; western blottings

DESCRIPTION (provided by applicant): Metalloproteinase activity at the cell surface can strongly influence cell sensitivity to extrinsic death vs. survival signals in a variety of cell types, yet the convergence of metalloproteinases and receptor signaling in the nervous system remains largely unexplored. TIMP-3 (tissue inhibitor of metalloproteinase-3) is a unique natural metalloproteinase inhibitor, in that it is a potent inhibitor of all known metalloproteinase sheddases, and has been shown to play a role in the regulation of receptor-mediated cell death in various non-neuronal cell types. TIMP-3 plays a pro-apoptotic role in many cell types through its ability to inhibit sheddases that target death receptors and their ligands. Our interest in the role of metalloproteinases in CNS damage following stroke led us to investigate the expression of TIMP-3 and the metalloproteinase sheddase, MMP-3, in a rodent model of focal cerebral ischemia. We found that while TIMP-3 and MMP-3 are expressed at low to non-detectable levels in the adult brain, their expression becomes markedly upregulated following ischemia, particularly in cortical neurons undergoing delayed apoptotic death. Using a tissue culture approach, we found that TIMP-3 and MMP-3 are constitutively expressed by embryonic cortical neurons in culture and modulate neuronal sensitivity to receptor-mediated apoptosis induced by the chemotherapeutic drug, doxorubicin (Dox). Metalloproteinase inhibition by TIMP-3 was found to be necessary for Dox-induced apoptosis, whereas addition of exogenous active MMP-3 markedly attenuated apoptosis and blunted death receptor-ligand interactions at the cell surface. These observations strongly implicate a physiologic role for TIMP-3 and MMP-3 in the regulation of receptor-mediated death in the nervous system. While metalloproteinase activity has previously been implicated in vascular damage following cerebral ischemia, the ability of metalloproteinases and their inhibitors to influence neuronal vulnerability to ischemic stress has not been studied. In the proposed studies, experiments are designed to establish a role for metalloproteinase activity in the regulation of receptor-mediated neuronal death following cerebral ischemia, and to explore underlying mechanisms, utilizing both in vitro and in vivo models of ischemic injury, coupled with pharmacological and gene deletion approaches.

描述（由申请人提供）：在细胞表面上的金属蛋白酶活性可以强烈影响细胞对外在死亡的敏感性与多种细胞类型中的生存​​信号，但是金属蛋白酶和受体信号在神经系统中的收敛仍然很大程度上尚未得到探索。 TIMP-3（金属蛋白酶3的组织抑制剂）是一种独特的天然金属蛋白酶抑制剂，因为它是所有已知的金属蛋白酶SHEDDase的有效抑制剂，并且已被证明在各种非神经元细胞类型的受体介导的细胞死亡调节中起作用。 TIMP-3通过抑制靶向死亡受体及其配体的sheddase的能力，在许多细胞类型中起促凋亡的作用。我们对中风后金属蛋白酶在中枢神经系统损伤中的作用的兴趣使我们研究了TIMP-3和金属蛋白酶Sheddase MMP-3的表达，在局灶性脑缺血的啮齿动物模型中。我们发现，尽管TIMP-3和MMP-3在成年大脑中以低至不可检测的水平表达，但它们的表达显着上调，尤其是在延迟凋亡死亡的皮质神经元中。使用组织培养方法，我们发现TIMP-3和MMP-3在培养中由胚胎皮质神经元组成式表达，并调节化学治疗药物，阿霉素（DOX）引起的受体介导的凋亡的神经元敏感性。发现TIMP-3抑制金属蛋白酶酶是DOX诱导的细胞凋亡所必需的，而添加外源活性MMP-3显着减弱了细胞凋亡，并且在细胞表面上呈钝化的死亡受体 - 培养基相互作用。这些观察结果强烈暗示了TIMP-3和MMP-3在神经系统中受体介导的死亡调节中的生理作用。虽然金属蛋白酶活性以前与脑缺血后有关血管损伤，但金属蛋白酶及其抑制剂影响神经元脆弱性的缺血性胁迫的能力尚未得到研究。在拟议的研究中，设计实验旨在确立金属蛋白酶活性在脑缺血后受体介导的神经元死亡调节中的作用，并探索与药理学和基因删除方法相连的体外和体内缺血模型的潜在机制。