Role of Cell Cycle Pathways in Traumatic Brain Injury

创伤性脑损伤可导致神经细胞死亡，并伴有星形胶质细胞的增殖和炎症 与小胶质细胞的激活有关。细胞周期蛋白上调发生在CMS创伤后，并出现 导致有丝分裂后细胞(如神经元)的细胞死亡。它还可能导致创伤后 胶质细胞增多症和小胶质细胞活化。我们实验室最近的研究表明， 啮齿动物脑损伤或脊髓损伤后多种细胞周期蛋白的表达及其在神经元中的共表达 显示caspase-3激活及细胞凋亡的形态特征。此外，在几个经典的模型中， Caspase-3依赖于原代培养神经细胞的凋亡，损伤与许多 这些相同的细胞周期蛋白。此外，初步研究表明，抑制关键细胞周期调节 在体外和体内，通路都可以减少损伤诱导的细胞死亡。因此，使用细胞周期抑制剂进行治疗 大鼠脑外伤后损伤体积明显减少，周围胶质瘢痕明显减少，也明显改善 脑损伤后的运动和认知功能。 拟议的研究旨在解决以下假设：(1)TBI上调关键细胞周期 神经元、星形胶质细胞和小胶质细胞中mRNA和蛋白质水平的成分；(2)这种上调 促进神经元的凋亡和星形胶质细胞的增殖；(3)细胞周期蛋白的上调 与小胶质细胞活化和随后相关炎症因子的释放有关；以及(4)细胞治疗 周期抑制物具有神经保护作用，其机制包括抑制体内caspase途径。 神经元减少，神经胶质细胞激活减少，小胶质细胞介导的炎症因子释放减少。 具体目的是证明：(1)脑损伤导致一些关键细胞周期的表达增加 相关基因/蛋白，包括神经元和神经胶质细胞中的细胞周期蛋白D1、CDK4、CDK5和Rb； 表达与caspase依赖的神经元凋亡、星形胶质细胞增殖、活化 小胶质细胞和促进小胶质细胞相关炎症因子的释放；c.细胞周期蛋白D1基因敲除小鼠表现较少 强烈的损伤诱导的病理生物学包括神经细胞凋亡，脑损伤，星形胶质瘢痕形成，释放 小胶质细胞相关炎症因子与创伤后神经功能障碍；(2)结构不同的细胞 周期抑制剂以剂量依赖的方式缩小病变体积，改善认知和运动功能 在两种不同病理生物学的大鼠和小鼠脑损伤模型中；B.细胞周期抑制剂降低细胞周期 脑损伤后的激活，从而减少随后的神经细胞死亡、反应性胶质增生和小胶质细胞的激活； 延迟全身给药细胞周期抑制剂是一种更具临床意义的范例，具有神经保护作用。

Traumatic brain injury (TBI) causes neuronal cell death combined with astroglial proliferation and inflammation associated with activation of microglia. Upregulation of cell cycle proteins occurs after CMS trauma, and appears to contribute to apoptotic cell death of post-mitotic cells such as neurons. It also likely contributes to posttraumatic gliosis and microglial activation. Recent studies in our laboratory have shown significantly increased expression of many cell cycle proteins after TBI or spinal cord injury in rodents, with the proteins co-expressed in neurons showing caspase-3 activation and morphological features of apoptosis. Moreover, in several classical models of caspase-3 dependent apoptosis in primary neuronal cell cultures, injury is associated with up-regulation of many of these same cell cycle proteins. In addition, pilot studies have indicated that inhibition of key cell cycle regulatory pathways reduces injury-induced cell death both in vitro and in vivo. Thus, treatment with a cell cycle inhibitor after TBI in rats markedly reduces lesion volumes and the surrounding glial scar; it also significantly improves motor and cognitive functions following brain injury. The proposed studies are intended to address the following hypotheses: (1) TBI up-regulates key cell cycle constituents at both the mRNA and protein levels in neurons, astrocytes, and microglia; (2) such an up-regulation promotes apoptosis in neurons and proliferation of astrocytes; (3) up-regulation of cell cycle proteins contributes to microglial activation and subsequent release of associated inflammatory factors; and (4) treatment with cell cycle inhibitors is neuroprotective, through mechanisms that include inhibition of the intrinsic caspase pathway in neurons, as well as reduced glial activation and diminished release of microglial mediated inflammatory factors. Specific aims are to demonstrate that: (1) a. TBI causes increased expression of a number of critical cell cycle related genes/proteins, including cyclin D1, CDK4, CDK5 and Rb in both neurons and glia; b. increased protein expression is associated with caspase-dependent apoptosis in neurons, proliferation of astroglia, activation of microglia and facilitated release of microglia-related inflammatory factors; c. cyclin D1 knockout mice show less intense injury-induced pathobiology including neuronal apoptosis, brain lesion, astroglial scar formation, release of microglial associated inflammatory factors, and post-traumatic neurological deficits; (2) a. structurally different cell cycle inhibitors in dose-dependent manner reduce lesion volumes and improve cognitive as well as motor function in two pathobiologically different TBI models in the rat and mouse; b. cell cycle inhibitors decrease cell cycle activation after TBI, thereby reducing subsequent neuronal cell death, reactive gliosis and microglial activation; c. delayed systemic administration of a cell cycle inhibitor, a more clinically relevant paradigm, is neuroprotective.