Neuritin转基因鼠TGI模型脑组织差异蛋白分析及其分子机制的研究

Neuritin, a novel neurotrophic factor, plays critical roles in neurite growth and neuronal survival with the function of nerve-injury recovery. However, researches in the field are mostly limited in cellular level, lacking of series of systemic evaluation, and the molecular mechanisms of neuritin-related function are also poorly understood so far. Accordingly, our research is aimed to investigate whether high expression of neuritin could activate neural repair pathway, so as to excavate its potential function in protecting and repairing neural injury in transgenic mice. At the same time, the Transient Globe Ischemic (TGI) model will be constructed by occluding the double common carotid arteries in mice. The differentially expressed proteins between transgenic and non-transgenic TGI mice will be analyzed by means of iTRAQ technique, followed by verification of the interaction of neuritin with the differentially expressed proteins by both in vivo and in vitro approaches. The proposal may further elucidate the molecular mechanism of neuritin in neural repair and nerve regeneration, provide theoretic foundation and technical support for the treatment of the kinds of disease, and offer a new opportunity to address the unmet medical need in the patient population.

Neuritin是一种与神经可塑性密切相关的神经营养因子，具有维持神经元存活、促进神经突起生长的作用，这些特性赋予其具有神经损伤后的修复功能。然而，目前对该领域的研究十分有限且多停留在环境相对简单的细胞水平，缺乏系统水平下的全面评价，对其发挥作用的分子机制更是知之甚少。鉴于此，本研究旨在探索neuritin转基因鼠脑缺血损伤后，过表达的neuritin能否有效发挥神经再生修复功能，从动物个体水平评价neuritin在中枢神经系统损伤中的作用；同时，利用iTRAQ技术筛选转基因与非转基因TGI模型后小鼠脑组织差异表达的蛋白，从在体和细胞水平验证这些差异蛋白与neuritin之间互作关系，阐明neuritin在受损神经再生修复过程中的分子机制，为更好地挖掘neuritin基因的功能，探索其在神经系统疾病治疗方面的作用提供理论依据和技术支撑。

脑缺血再灌注损伤是由多因素、多环节、多途径共同引发的一系列级联复杂的细胞反应，神经营养因子的缺乏是加重再灌注损伤的重要因素，Neuritin作为调节神经系统的发育和可塑性的神经营养因子，使其成为神经损伤性疾病的潜在靶点，而目前欠缺在个体水平综合评价含GPI 锚定位点的全长蛋白对中枢神经系统损伤后的相关数据。本研究通过核显微注射技术构建Neuritin转基因的小鼠，并从基因水平、转录水平及蛋白水平全面筛选过表达的后代用于繁殖建系。并以此为系建立能够稳定表达的Neuritin基因的转基因子代鼠。通过构建小鼠全脑短暂性脑缺血模型，从复杂系统生物学体系下探究成熟Neuritin蛋白对于受损神经的再生与修复作用。与野生组相比，转基因组能抑制脑缺血再灌注后修复关键时间窗Parp1蛋白的降解，减少对缺血敏感的海马区神经元尼氏体的丢失，上调神经再生标志物NF-200、SYN和GAP-43 的表达，促进神经突起的再生和突触可塑性的重建，同时，能够显著提升小鼠学习与记忆能力，加速空间认知功能的恢复。本研究表明Neuritin基因在脑缺血再灌注后可提供维持神经细胞存活和突起再生有利的微环境，这些特性为开发治疗缺血性中风的药物提供新的策略。