Connexin43调控脑出血后星形胶质细胞表型转化的机制及其在EMSC移植治疗中的应用

Reactive astrocytes play both beneficial and detrimental roles in the pathological process after intracerebral hemorrhage (ICH). It was reported on Nature that there are two phenotypes of reactive astrocytes. A1 reactive astrocytes lose most normal astrocytic functions but gain a new neurotoxic function by up-regulating many classical complement cascade genes, rapidly killing neurons and mature differentiated oligodendrocytes. A2 reactive astrocytes up-regulate many neurotrophic factors thus are neuroprotective. Previously we reported that Connexin 43 (Cx43) downregulation triggered YAP nuclear translocation and promoted astroglial-mesenchymal transition (AMT) during the early stage of ICH-induced astrocytic activation. By AMT, the resting astrocytes (A0) switch to mesenchymal phenotype (Am). We believe that Am is an intermediate phenotype formed during astrocytic activation to meet the requirement of proliferation and migration. Eventually, Am will differentiate into A1 or A2 astrocytes. The phenotypic transition path is A0→Am→A1/A2. Referring to literatures, we found that Cx43 has multiple function modes such as gap junctional intercellular communication, hemichannel function and non-channel related regulational function. Based on our previous findings that Cx43 mediates astrocytic activation and ectomesenchymal stem cell (EMSC) is rich in Cx43 and can regulate astrocytic activation. We propose that Cx43 regulates the phenotypic transition of astrocytes in a multi-target mode. Then we verify the hypothesis by in-vivo and in-vitro experiments. On this basis, we will further study the regulational effect and mechanisms of EMSC on the biological behavior of reactive astrocytes. Our study will provide the experimental evidence for future ICH treatment based on regulating astrocytic phenotype switching and EMSC transplantation.

脑出血（ICH）后星形胶质细胞（ASC）活化是一把“双刃剑”。研究发现，活化的ASC可分为A1（神经毒性）和A2（神经保护）两种表型。我们前期研究发现Cx43调控了ASC由静息态（A0）向间充质表型（Am）转化，我们认为Am是A0增殖、迁移并分化为A1或A2过程中形成的中间表型，ASC活化的表型转化路径是A0→Am→A1/A2。本课题组通过调研大量文献，发现Cx43具有缝隙连接通道功能、半通道功能和非通道功能等多种作用模式，基于Cx43参与调控ASC活化以及外胚间充质干细胞（ectomesenchymal stem cell，EMSC）高表达Cx43并可驯化ASC的前期研究发现，我们提出Cx43调控ASC表型转化的多靶点作用模式假说并进行验证。在此基础上进一步研究EMSC调控ASC生物学行为的作用机制，为今后以Cx43为分子靶点调控ASC表型转化和EMSC移植修复ICH后神经损伤提供依据。

星形胶质细胞活化在脑出血后的继发性损伤中扮演重要角色，探讨其活化及表型转化机制对于脑出血后的神经保护治疗具有重要意义。本项目按计划完成以下工作：（1）对缝隙连接蛋白Connexin43（Cx43）介导的通道功能以及非通道相关的基因表达调控功能在脑出血诱导的铁死亡和细胞凋亡中的调控作用进行了深入研究；（2）通过单细胞转录组测序对脑出血后不同时间点小鼠血肿周边脑组织进行检测，通过拟时序分析描绘出星形胶质细胞和小胶质细胞在脑出血诱导的活化过程中的表型演变轨迹；（3）探讨了外胚间充质干细胞EMSC基于Cx43对脑出血背景下A1表型星形胶质细胞的逆转调控作用及机制；（4）并通过单细胞转录组测序方法对EMSC进行鉴定，探讨EMSC发挥调控作用的机制，为EMSC移植用于调控脑出血后炎症微环境治疗提供研究基础和理论依据；（5）针对脑出血后氧化应激及炎症反应这两大继发性损伤，采用多孔硒纳米材料Se@SiO2和大蒜素进行治疗，探讨其神经保护作用及机制。并在此基础上，衍生出Cx43介导的ASC表型转化在胶质瘤和脑转移瘤中的作用等课题方向，形成系列研究。