基于Cx43蛋白泛素化探讨缺血性脑中风发生后星形胶质细胞极化介导的神经系统保护作用及机制

Astrocytes are abundant cells in the CNS that can both hinder and support CNS recovery. It has been reported that the A2 subtype of reactive astrocytes have protective effects on the CNS, while the mechanism has not been clear and many questions remain about their functions. Our previous study showed that in the A2 subtype of reactive astrocytes, there was a high plasma-membrane shifting level of connexins43, which provides protective support for neurons. By utilizing the protein ubiquitination analysis technology, we found the key ubiquitin E3 ligases, Smurf2, that mediates the ubiquitination of Cx43 in the A2 subtype of reactive astrocytes. The further study demonstrated that HIF-1α-related signaling pathway regulated the function of Smurf2-RNF11 on the ubiquitination of Cx43. Based on the research results, here we plan to establish the connection between the downstream regulation of HIF-1α and the ubiquitination regulation of Cx43 in the A2 subtype of reactive astrocytes. In this project, we intend to clarify the CNS protection mechanism of reactive astrocytes in cerebral ischemia, which would provide new strategies for the diagnosis and therapy of cerebral ischemia.

星形胶质细胞在神经系统中发挥着损伤和保护的双重作用。研究表明，缺氧诱导下星形胶质细胞发生A2极化并产生神经保护作用，然而其具体作用机制尚不明晰。本项目前期研究发现 A2型星形胶质细胞内缝隙连接蛋白Cx43泛素化水平降低从而发生明显质膜转移，产生神经元保护作用。通过蛋白泛素化分析我们发现了A2型星形胶质细胞内介导Cx43泛素化的关键泛素E3连接酶Smurf2。进一步功能学研究表明缺氧诱导因子HIF-1α在A2型星形胶质细胞内高度表达，其下游基因的转录调控可影响RNF11参与的Smurf2对Cx43的泛素化调节。基于上述研究，本项目主要思路是将HIF-1α作为抓手，深入探究HIF-1α所介导的下游信号通路，将其与Cx43泛素化机制建立联系，为探究缺氧诱导下星形胶质细胞A2极化介导的神经元保护作用的分子作用网络明确关键联结位点，从而为靶向新型药物研发及临床诊断治疗缺血性脑中风提供新的思路和方法。

星形胶质细胞在神经系统中发挥着损伤和保护的双重作用。研究表明，缺氧诱导下星形胶质细胞发生A2极化并产生神经保护作用，然而其具体作用机制尚不明晰。本项目前期研究发现A2型星形胶质细胞内缝隙连接蛋白Cx43泛素化水平降低从而发生明显质膜转移，产生神经元保护作用。通过蛋白泛素化分析我们发现了A2型星形胶质细胞内介导Cx43泛素化的关键泛素E3连接酶Smurf2。进一步功能学研究表明缺氧诱导因子HIF-1α在A2型星形胶质细胞内高度表达，其下游基因的转录调控可影响RNF11参与的Smurf2对Cx43的泛素化调节。据此，课题组提出科学假说：缺氧条件下A2型极化的星形胶质细胞HIF-1α通路被激活，调控Cx43的泛素化，促进其质膜转移，进而发挥抗脑缺血保护作用。通过借助小鼠脑缺血模型，星形胶质细胞-神经元细胞共培养等技术，探究缺氧诱导下星形胶质细胞A2极化介导的神经元保护作用的分子作用网络明确关键联结位点。实验结果揭示，在缺氧诱导下星形胶质细胞A2极化介导的神经元保护作用依赖于缝隙连接蛋白Cx43的质膜转移，缺氧状态下Cx43的质膜转移依赖于HIF-1α的转录后调控机制，并在体内水平明确了星形胶质细胞A2极化介导的神经元保护作用在缺血性脑中风治疗中的作用。本研究为靶向星形胶质细胞的新型抗脑缺血药物研发和临床诊断/治疗提供了新的思路和方法。