持续激活ErbB信号通路诱导病理性激活星型胶质细胞转分化为神经元的机制和应用研究

The hardship of neuronal regeneration after their death is a bottleneck for therapies of neurological disorders. Inducing reactive astrocytes to reprogram into neurons is one of the important directions for development of therapeutic approaches. We have revealed that constitutively activated ErbB signaling promotes efficient reprogramming of reactive astrocytes into neurons. Based on our preliminary results, we propose here to investigate the steps of reprogramming astrocytes and the properties of induced neurons by combining multiple in vivo approaches including genetic lineage tracing, morphological analysis, and molecular marker verification, as well as to investigate the cellular reprogramming mechanisms by in vitro approaches including primary astrocyte culture and neuronal induction. Meanwhile, we will explore the molecular mechanisms for ErbB signaling to induce reactive astrocytes into neurons by inhibition of different ErbB downstream signaling with small chemical compound inhibitors. To assess the translational potential, we will constitutively activate ErbB signaling in reactive astrocytes induced in CNS injury mouse models, and examine the incorporation of induced neurons into the existed system, and the survival time of induced neurons. All the results will provide fundamental base for development of therapeutic medicine to induce neuronal regeneration by reprogramming reactive astrocytes in various neurological disorders.

神经元缺失后无法再生是治疗神经系统疾病的瓶颈，诱导体内病理性激活星形胶质细胞重编程并分化成神经元是开发新疗法的重要方向。我们前期研究发现：持续激活ErbB信号通路能高效促进病理性激活星型胶质细胞转分化为神经元。本项目将利用遗传学细胞系追踪手段，结合细胞形态、分子标记物检测等指标，通过体内和离体实验，追踪星形胶质细胞的重编程过程，研究诱导神经元的种类，检测诱导神经元的突触形成情况，阐明该过程的细胞机制；同时利用ErbB信号通路的胞内下游各通路的特异小分子抑制剂，深究ErbB信号通路诱导星型胶质细胞转分化的分子机制；构建成年小鼠脑损伤模型，通过电生理实验检测诱导神经元与损伤部位局部环路的整合程度，并追踪诱导的神经元存活时间，探讨ErbB信号通路诱导星形胶质细胞转分化神经元修复中枢损伤的应用。研究结果将为开发靶向病理性激活星形胶质细胞诱导神经元再生来治疗神经系统疾病的药物提供必要的理论依据。

本研究包含以下三部分：.1神经元退行引发星形胶质细胞病理性激活，抑制其活化能够延缓病情，发掘其细胞和分子机制利于开发治疗靶点。本项目利用可诱导ErbB信号通路亢奋/抑制的转基因小鼠模型，发现ErbB信号通路激活诱导星形胶质细胞病理性活化，而在物理损伤的大脑皮层中，抑制该通路能够减缓星形胶质细胞的活化特征及胶质瘢痕面积。研究结果深入解析了星形胶质细胞活化的分子机制，为神经元退行疾病疗法研发提供了潜在分子靶点。.2神经元退行疾病新疗法研发中，诱导活化的星形胶质细胞转化成神经元是一个潜在的研发方向，利用遗传学细胞系追踪手段，我们通过体内和体外研究，探讨ErbB信号通路持续活化能否诱导星形胶质细胞转分化，然而实验结果否定了我们的假设。.3 已报到的体外研究发现，星形胶质细胞参与神经细胞间扩散Aβ和prion等变异蛋白，可能促进神经元退行疾病的恶化过程，本项目发现：在小鼠大脑皮层内注射携带EGFP基因的AAV后，Layer Ⅰ-Ⅲ中感染的星形胶质细胞特异表达的EGFP，通过纳米级细胞连接通道-TNT，运输到Layer Ⅴ的神经元内。本项目为神经元退行疾病病理研究，提供了实验动物模型。