人骨髓间充质干细胞定向内皮分化前后miRNA的表达差异分析  

Differential Expression of miRNA before and after Directional Endothelial Differentiation of Human Bone Marrow Mesenchymal Stem Cell

作　　者：罗雪兰 周炜潜 杨鹏 陈伟 李龙春 欧和生 LUO Xuelan;ZHOU Weiqian;YANG Peng(Guangxi International Zhuang Medicine Hospital,Guangxi 530201,China)

机构地区：[1]广西国际壮医医院壮瑶药制剂中心,南宁530201 [2]广西中医药大学研究生院,南宁530200 [3]广西国际壮医医院科技部,南宁530201 [4]济南市妇幼保健院药剂科,250001

出　　处：《医学研究杂志》2023年第10期71-75,共5页Journal of Medical Research

基　　金：国家自然科学基金地区基金资助项目(82060655);广西高校中青年教师科研基础能力提升项目(2019KY0330)。

摘　　要：目的探讨人骨髓间充质干细胞(human bone marrow mesenchymal stem cell,hBMSC)定向内皮分化前后miRNA的表达差异分析,为进一步研究其调控hBMSC内皮分化的分子机制奠定理论基础。方法根据hBMSC定向血管内皮诱导分化培养方法培养细胞,应用电子显微镜观察细胞分化前后形态的变化,应用细胞免疫荧光法检测内皮型一氧化氮合酶(endothelial nitric oxide synthase,eNOS)的蛋白质表达情况,应用高通量测序分析检测细胞分化前后miRNA表达谱的改变,应用qRT-PCR验证两组中表达差异明显的miRNA分子。结果与未分化组hBMSC比较,分化后的细胞形态发生类内皮细胞样改变;eNOS蛋白的表达明显增加;分化后hBMSC共筛选41个差异性表达的miRNA,其中表达上调的有27个,表达下调的有14个;分化组细胞miR-31-5p上调近21倍,miR-200a-3p下调近15倍。结论hBMSC定向内皮分化前后miRNA表达发生差异性改变,其改变可能影响干细胞的分化程度,可为进一步研究干细胞在血管内皮功能异常所致的血管性疾病异常机制提供研究方向和理论支持。Objective To explore the differential expression of miRNA before and after endothelial differentiation of human bone marrow mesenchymal stem cell(hBMSC),and to lay a theoretical foundation for further study on the molecular mechanism of miRNA regulating the differentiation of hBMSC.Methods The cells were cultured according to the method of inducing differentiation of hBMSC into vascular endothelium.The morphological changes of cells before and after differentiation were observed with an electron microscope.The protein expression level of endothelial nitric oxide synthase(eNOS)was detected by immunofluorescence.The difference in miRNA expression profiles before and after differentiation was detected by high-throughput sequencing analysis.Results Compared with the undifferentiated hBMSC group,the cell morphology was significantly changed after differentiation.The expression of eNOS protein increased significantly.A total of 41differentially expressed miRNA were screened,of which 27 were up-regulated and 14 were down-regulated.In the differentiation group,miR-31-5p was up-regulated nearly 21 times,and miR-200a-3p was down-regulated nearly 15 times.Conclusion The expression of miRNA changed differently before and after the differentiation of hBMSC into endothelial cells,which may affect the degree of differentiation of stem cells,and provide research direction and theoretical support for further study on the abnormal mechanism of vascular diseases caused by abnormal function of stem cells in vascular endothelium.

关 键 词：人骨髓间充质干细胞 内皮分化 MIRNA ENOS 

分 类 号：R318[医药卫生—生物医学工程]