星形胶质细胞通过CX47促进少突胶质前体细胞增殖  

Astrocyte Promotes Oligodendrocyte Precursor Cell Proliferating through Connexion CX47

作　　者：李真[1] 刘兆雨 徐丹[2] 陈婷[1] 孟赞[1] 唐勇[1] 彭彦[1] 

机构地区：[1]重庆医科大学基础医学院,重庆400016 [2]重庆医科大学第一附属医院,重庆400016

出　　处：《中国生物工程杂志》2015年第12期21-29,共9页China Biotechnology

基　　金：国家自然科学基金(81470057);重庆市首批"百名学术科学领军人才"培养计划资助项目

摘　　要：目的:研究星形胶质细胞(AST)对少突胶质前体细胞(OPC)生长分化的影响及作用机制。方法:用P3SD乳鼠,建立星形胶质细胞与少突胶质前体细胞原代培养体系。实验分两部分,第一部分为常规OPC培养组、AST分泌因子组、AST与OPC直接接触式培养组;第二部分为AST与OPC直接接触式培养组、缝隙连接干扰对照组、缝隙连接干扰组。免疫荧光鉴定OPC与AST细胞纯度,光镜观察细胞生长状态,流式细胞术检测细胞周期,转录组测序分析不同状态下OPC中缝隙连接蛋白CX47差异表达,PCR和Western blot检测鉴定转录组测序CX47差异表达,EDU检测CX47干扰后OPC增殖能力,流式细胞术测干扰CX47后细胞周期变化。结果:光镜与流式检测发现,与AST接触培养的OPC增殖能力最强、新生细胞数目最多。转录组测序分析和PCR验证显示,AST接触调控OPC增殖的主要差异表达蛋白为CX47,干扰CX47后细胞周期阻滞在G1期,OPC增殖能力明显下降。结论;AST可通过缝隙连接蛋白CX47调控细胞周期,促进OPC增殖。Objective：To explore the effect and mechanics of growth and differentiation on oligodendrocyte precursor cell （OPC）influences by astrocyte （AST）, and to provide theoretical and experimental basis for the treatment of demyelinating diseases. Methods： OPC and AST were isolated from SD Rats about 1 ~ 3 days. The study is divided in two parts, The first part includes three groups ： OPC conventionally cultured group （ OPC ）, AST and OPC co-cultured in Transwell room group （CO-OPC）, AST and OPC co-cultured through directly contacted group （AST-OPC）. The second part a/so includes three parts： AST and OPC co-cultured through directly contacted group, connexion interference normally control group, connexion interference group. Growth state was observed by optical microscope and the cell cycle distribution was measured by flow cytometry. The dominant factor promoting proliferation and differention of OPC analyzed by transcriptorne sequencing, and the relative expression of connexion was measured by reverse transcription-polymerase chain reaction. Western blot was used to detect the expression of connexion 47. The proliferation of OPC was measured by EDU assay and the cell cycle distribution was determined by flow cytometry after connexion 47 interferenced. Result ： Cell purity of OPC and ast were more than 95 % labeled by PDGFoR and GFAP. In the group of OPC and AST co-cultured through directly contact, OPC has greater proliferative ability than control group, newly horned OPCs were more than conventionally cultured and transwell cultured. The differential expression of connexions was CX47 measured by transcriptome sequencing, and verified by reverse transcription-polymerase. The viability of OPC was obviously decreased and the cell cycle distribution of OPC was hold in G0/G1 phase after connexion 47 interferenced. Conclusion; The proliferation of OPC was promoted by AST through directly contact, and the underlying mechanism may be related to the upregulation of connexion 47.

关 键 词：少突胶质前体细胞 缝隙连接 髓鞘再生 转录组测序 

分 类 号：Q421[生物学—神经生物学]