Wnt/β-catenin通路介导海马神经发生在CIHH改善癫痫鼠认知功能中的机制研究

Increasing evidence showed that spontaneous recurrent seizures (SRS) occurring in status epilepticus (SE) frequently lead to debiliatting cognitive impairment in patients, prompting research on the potential delay of chronic SRS after SE to rescue cognitive function. We previously found that chronic intermittent hypobaric hypoxia (CIHH) pretreatment effectively decreased SRS, rescued neuronal damage, increased the number of synapses, and induced hippocampal neurogenesis after SE. While the exact underlying mechanisms of CIHH remain unclear, the Wnt/β-catenin pathway, mediating hippocampal neurogenesis, has been proposed as a main mechanism. On the basis of previous research, we will include neurogenesis as an outcome measure and observe the effect of CIHH treatment in rats after SE. To elucidate the effect of CIHH on hippocampal neurogenesis, synaptic plasticity, and cognitive function, a retroviral vector carrying the nestin-TK gene will be used to inhibit neurogenesis in the rat hippocampus. We will also utilize a specific inhibitor of the Wnt/β-catenin pathway to verify whether CIHH treatment is able to activate this pathway, to induce hippocampal neurogenesis, to increase synaptic plasticity to repair the injured neuronal network, and to rescue SE-impaired cognitive function in both in vitro cultured hippocampal neurons and in vivo experiments. The study will contribute to provide a new target for epilepsy treatment and intervention.

癫痫持续状态（SE）常发展为慢性期自发性反复发作（SRS），易致患者出现明显的认知损害，如何减缓SE后SRS从而阻止认知损害一直是治疗的难点。前期我们发现：慢性间歇性低压低氧（CIHH）预处理能有效减缓SE后大鼠SRS，减缓海马神经元损伤,并增加突触数量及诱导神经发生,但机制不明。研究证实Wnt/β-catenin通路为介导海马神经发生的主要通路。本项目拟在前期研究基础上通过CIHH干预，从神经发生角度出发，构建逆转录病毒载体nestin-TK基因抑制神经干细胞，观察CIHH对SE后大鼠海马的神经发生、突触可塑性及认知功能的影响；并通过离体原代海马神经干细胞培养和在体实验，特异性抑制Wnt/β-catenin通路,验证CIHH是否可通过激活该通路介导的海马神经发生来修复SE后损伤的神经元，进而增强海马突触可塑性以改善其认知功能这一假说。本项目旨在为癫痫的治疗提供一种新的靶点和干预手段。

癫痫持续状态（SE）常发展为慢性期自发性反复发作（SRS），易致患者出现明显的认知功能损害，如何减缓SE后SRS从而有效阻止认知损害一直是临床中治疗的难点。前期我们研究发现：慢性间歇性低压低氧（CIHH）预处理能有效减缓SE后大鼠SRS，减轻海马神经元损伤，并可诱导海马神经发生，但机制不明。近期大量研究证实Wnt/β-catenin信号通路为介导成年海马神经发生的主要通路。本项目在前期研究基础上通过CIHH干预，从神经发生的角度出发，制备国际常用的SE动物模型，分别通过Morris水迷宫和新物体识别试验进行行为学检测，Nissl染色、免疫荧光共定位及透射电镜扫描等方法进行神经发生相关的形态学检测，同时应用Western blot等方法检测相关神经生长因子的表达变化，进一步应用在体电生理技术记录大鼠海马的场电位变化情况，明确了CIHH可通过诱导内源性海马神经发生来改善SE后慢性期大鼠的认知功能。随后通过离体原代海马神经干细胞培养和在体动物实验，给予Wnt/β-catenin信号通路特异性抑制DKK-1后，进一步验证CIHH可通过激活该通路介导的海马神经发生来修复SE后损伤的神经元，进而改善其认知功能这一假说。本项目的研究结果旨在为临床SE患者的慢性进行性认知损害提供一种有效的防治手段和新的干预靶点。