Role of calpastatin in modulating function following TBI

创伤性脑损伤(TBI)导致钙调蛋白的长时间激活，导致细胞骨架的形成。 损伤、神经元死亡和行为障碍。然而，令人惊讶的是，几乎没有体内的细胞底物 在创伤损伤的大脑中已经发现了钙调蛋白，因此，人们对此知之甚少。 肌钙蛋白在创伤后发病中的作用途径。内源性抑制因子--钙化抑素 在钙调蛋白中，是唯一已知的专门抑制调理蛋白的蛋白质。因此，Calastatin代表着一种 理想的分子工具，可以用来分离受伤大脑中钙调蛋白的活动。翻译的努力 激动人心的临床前数据显示，脑损伤啮齿动物在治疗后功能得到改善 外源性钙蛋白酶抑制剂因其溶解性、特异性和生物利用度的挑战而放缓 小分子抑制剂。增强内源性钙调蛋白活性可能代表着一种新的和有效的 治疗方法。因此，项目1的总体目标是评估钙调蛋白在 调节创伤后钙蛋白酶介导的蛋白分解，并评估神经保护和行为 提高钙调蛋白活性在脑外伤治疗中的作用。使用基因改造的小鼠 过表达人钙蛋白酶抑素或缺乏钙蛋白酶抑素，项目1将：1)评估钙蛋白酶抑素的作用 在调节局灶性或弥漫性脑损伤后的行为结果时，2)量化改变的影响 脑局灶性或弥漫性脑损伤后神经细胞存活和轴突损伤后钙调蛋白的表达 Calastatin在限制创伤诱导的神经元细胞骨架蛋白降解中的体内作用，以及4) 评价创伤后钙蛋白水解酶激活在修饰钙相关膜蛋白中的作用 内流和调控线粒体相关的细胞死亡事件。我们的中心假设是钙调蛋白 过度表达将阻止Calain介导的对细胞生存至关重要的神经元底物的切割， 从而减轻创伤后神经元死亡和功能障碍。拟议的实验将提供 Calastatin在创伤后病理中功能作用的第一个证据及阐明区别 钙蛋白酶/钙调蛋白系统在局灶性和弥漫性脑损伤中的作用。此外，该项目还将提供 基于对钙调蛋白系统的操纵，为新的治疗方法奠定基础，旨在 减轻因脑外伤和其他CMS损伤和疾病状态而造成的脑损伤和功能障碍。

Traumatic brain injury (TBI) results in the prolonged activation of calpains, which contributes to cytoskeletal damage, neuronal death and behavioral dysfunction. However, surprisingly few in vivo cellular substrates of calpains have been identified in the traumatically injured brain and, consequently, little is understood about the pathways through which calpains mediate posttraumatic morbidity. Calpastatin, the endogenous inhibitor of calpains, is the only known protein that exclusively inhibits calpains. As such, calpastatin represents an ideal molecular tool with which to isolate the actions of calpains within the injured brain. Efforts to translate exciting preclinical data demonstrating functional improvement in brain-injured rodents treated with exogenous calpain inhibitors have been slowed by challenges with solubility, specificity and bioavailability of small molecule inhibitors. Enhancing endogenous calpastatin activity may represent a novel and potent therapeutic approach. The overall goals of Project 1, then, are to evaluate the role of calpastatin in regulating posttraumatic calpain-mediated proteolysis and to assess the neuroprotective and behavioral efficacy of increasing calpastatin activity in the setting of TBI. Using genetically altered mice that either overexpress human calpastatin or are calpastatin deficient, Project 1 will: 1) evaluate the role of calpastatin in modulating behavioral outcome following focal or diffuse brain injury, 2) quantify the effects of altered calpastatin expression on neuronal survival and axonal injury after focal or diffuse brain injury, 3) determine the in vivo role of calpastatin in limiting trauma-induced proteolysis of neuronal cytoskeletal proteins, and 4) evaluate the role of posttraumatic calpain activation in modifying membrane proteins involved in calcium influx and in modulating mitochondria-related cell death events. Our central hypothesis is that calpastatin overexpression will prevent calpain-mediated cleavage of neuronal substrates critical for cell survival, thereby attenuating posttraumatic neuronal death and dysfunction. The proposed experiments will provide the first evidence for a functional role for calpastatin in posttraumatic pathology and elucidate differential roles for the calpain/calpastatin system in focal and diffuse TBI. In addition, this Project will provide the groundwork for novel therapeutic approaches, based on manipulation of the calpastatin system, aimed at attenuating brain damage and dysfunction due to TBI as well as other CMS injury and disease states.