低氧刺激通过HIF-1α直接调节Myocardin表达促进骨髓间充质干细胞向心肌样细胞分化  

Hypoxic stimulation directly regulates Myocardin expression through HIF-1α and promotes the differentiation of bone marrow mesenchymal stem cells into cardiomyocyte-like cells

作　　者：哈艳平 刘晓晓 邵钟铭 邹园 伍彩霞 郭峻莉[2,3] 申志华 揭伟 HA Yan-ping;LIU Xiao-xiao;SHAO Zhong-ming;ZOU Yuan;WU Cai-xia;GUO Jun-li;SHEN Zhi-hua;JIE Wei(Department of Pathology, School of Basic Medicine Sciences, Guangdong Medical University, Zhanjiang 524023, China;Key Laboratory of Emergency and Trauma, Ministry of Education, Hainan Medical University, Haikou 571199, China;Hainan Provincial Key Laboratory for Tropical Cardiovascular Diseases Research, Hainan Medical University, Haikou 571199, China)

机构地区：[1]广东医科大学基础医学院病理学系,湛江524023 [2]海南医学院急救与创伤研究教育部重点实验室,海口571199 [3]海南医学院海南省热带心血管病研究重点实验室,海口571199

出　　处：《临床与实验病理学杂志》2021年第9期1038-1043,共6页Chinese Journal of Clinical and Experimental Pathology

基　　金：国家自然科学基金(81170121)。

摘　　要：目的探讨低氧刺激在骨髓间充质干细胞(bone marrow mesenchymal stem cells,BMSC)向心肌细胞谱系分化过程中对心肌素(Myocardin)表达的影响及机制。方法分离Sprague Dawley大鼠股骨BMSC,磁激活细胞分选法获得c-Kit^(+)细胞亚群。分别用HIF-1α过表达慢病毒感染或物理性低氧(2%O2)刺激c-Kit^(+)BMSC,定量RT-PCR检测HIF-1α、Myocardin和心肌细胞分化基因(Nkx2.5、cTnT)的mRNA水平,免疫荧光检测Myocardin和cTnT蛋白表达,双荧光素酶报告基因法检测HIF-1α对Myocardin启动子活性的影响。结果HIF-1α过表达慢病毒感染和物理性低氧刺激均诱导c-Kit^(+)BMSC中Myocardin和心肌细胞分化标志基因Nkx2.5、cTnT的表达水平明显增高。shRNA抑制Myocardin后一定程度上减弱低氧诱导的c-Kit^(+)BMSC心肌细胞样分化。HIF-1α可直接上调Myocardin启动子活性。结论低氧刺激可通过HIF-1α直接上调Myocardin表达,进而促进c-Kit^(+)BMSC向心肌样细胞分化。Purpose To explore the effect and mechanism of hypoxic stimulation on Myocardin expression during the differentiation of bone marrow mesenchymal stem cells(BMSC)into cardiomyocyte-like cells.Methods Magnetic activated cell sorting method was adopted to isolate c-Kit^(+)subsets derived from Sprague Dawley rat femur BMSC.Hypoxia-inducible factor 1α(HIF-1α)overexpressing lentivirus or physical hypoxia(2%O2)were used to treat c-Kit^(+)BMSC.Then,quantitative RT-PCR and immunofluresent staining were used to detect the expression of HIF-1α,Myocardin and cardiomyocyte differentiation genes(Nkx2.5,cTnT).Dual luciferase reporter gene method was used to detect the impact of HIF-1αon Myocardin promoter activity.Results The expression of Myocardin and cardiomyocyte cell differentiation genes including Nkx2.5 and cTnT were significantly increased after treated c-Kit^(+)BMSC with HIF-1αoverexpression lentivirus or physical hypoxia.Inhibition of Myocardin by shRNA to some extent attenuated the hypoxia-induced cardiomyocyte-like differentiation of BMSC.In the mechanism,HIF-1αcan directly up-regulate Myocardin promoter activity.Conclusion Hypoxic stimulation promotes the differentiation of c-Kit^(+)BMSC into cardiomyocyte-like cells through the up-regulation of Myocardin expression via HIF-1αdirectly mediating Myocardin promoter’s activity.

关 键 词：骨髓间充质干细胞 心肌样细胞分化 低氧 HIF-1Α MYOCARDIN 

分 类 号：Q21[生物学—细胞生物学]