Ascl1介导Wnt/beta-catenin通路在TLE海马硬化中反应性Astrocytes异常增生的作用及调控机制

Reactive astrogliosis is the main pathological changes in hippocampal sclerosis of Temporal lobe epilepsy (TLE). In clinic, the part of hippocampal sclerosis was often removed by surgery, but which was no much better treatment effect. Although recently studies revealed that the reactive astrogliosis, accompanied by the opening of ion channels and activation of signaling molecules on astrocytes, those can trigger super synchronization of neurons discharge, how they lead to epileptogenesis still remains unknown.Our previous studies have shown that wnt/beta-catenin signaling pathway plays an important role in hippocampal neurogenesis. Preliminary experimental results also found the expression of Wnts signaling molecules was increased significantly in hippocampi of epileptic rats, which indicates the activated Wnt/beta-catenin signaling pathway could involve and aggravate in epileptogenesis. Our present study is intend to comprehensively study the role of Ascl1-regulated wnt/beta-catenin signaling pathway and its molecular mechinism in epileptogenesis via gene transduction, EEG monitoring and CoIP techniques coupling with cell culture, animal model and brain slices.We attempt to discover the novel strategy of targeting intervention reactive astrogliosis for seizures and provide the key theoretical evidence for reconstructing hippocampal sclerosis.

反应性Astrocytes增生是颞叶癫痫（TLE）海马硬化的主要病理特征。临床上应用外科手术切除TLE患者部分病变的海马，但疗效甚微。近年研究发现癫痫发作中反应性Astrocytes异常增生，常伴有离子通道的开放和信号通路的激活，且能够触发癫痫病灶内神经元超同步化放电，其具体机制不明。我们前期研究证实Wnt/beta-catenin通路在海马神经发生中具有重要作用。预实验结果发现Wnt信号分子在癫痫病灶内表达增高，提示其可能参与癫痫发生。本项目拟在细胞水平，动物整体和组织脑片多个水平上，结合基因转染、EEG脑电监测和CoIP免疫共沉淀等技术手段，系统研究癫痫发生中Ascl1调控 Wnt/beta-catenin的作用及分子机制。通过该研究，我们试图探索癫痫发作中靶向干预反应性Astrocytes异常增生转分化为功能性神经元的新策略，为实现海马硬化的重塑，提供重要的实验依据。

海马硬化和反应性Astrocytes增生是颞叶癫痫（TLE）主要的两大病理改变。临床上应用外科手术切除TLE患者部分病变海马，但疗效甚微。近年研究发现癫痫发作中反应性Astrocytes异常增生，常伴有离子通道的开放和信号通路的激活，能够触发癫痫病灶内神经元超同步化放电，其具体机制不明。本项目拟在细胞水平，动物整体和组织脑片多个水平上，结合基因转染、EEG脑电监测和CoIP免疫共沉淀等技术手段：首先研究高表达ASICL1促进海马神经胶质细胞的转分化效应。探明Wnts/beta-catenin信号通路在多种细胞，如原代培养的神经胶质细胞，海马神经干细胞，C6胶质瘤干细胞以及HT22海马神经元细胞系上多了Wnts分子的阳性表达情况；同时，应用Wnt受体阻断剂，进一步验证Wnts分子的阳性表达实验研究。且进行Wnt/β-catenin 信号途径的交互作用调控机制的体内外实验研究。另外，在体内外拓展实验中，应用Astrocytes/Microglia共培养体系，研究PD-1/PDL-1基因缺失后，神经胶质细胞上多个炎性分子的阳性表达实验研究，以及PD-1基因敲除小鼠帕金森模型上小胶质细胞的炎性激活致中脑黑质功能失衡的实验研究。预期阐明癫痫发生中Ascl1介导Wnts/beta-catenin信号参与调控细胞转分化的分子机制。通过该研究，我们试图探索癫痫发作中靶向干预反应性Astrocytes异常增生转分化为功能性神经元的新策略，为实现海马硬化的重塑，提供重要的实验依据。