人脐带间充质干细胞体外分离培养及其生物学特性的研究  

The study of isolation and culture in vitro of human umbilical cord mesenchymal stem cells and their biological properties

作　　者：韩潇[1,2] 白海[2] 赵强[2] 杨柯[2] 欧剑锋[2] 

机构地区：[1]兰州大学第二医院血液内科,兰州730030 [2]兰州军区兰州总医院血液科/全军血液病中心,兰州730050

出　　处：《重庆医学》2016年第7期876-879,共4页Chongqing medicine

基　　金：甘肃省科技重大专项(1102FKDA005)

摘　　要：目的 研究人脐带间充质干细胞（MSCs）详细的生物学特性,包括其细胞形态、免疫表型、纯度及增殖能力,从而建立稳定的MSCs体外分离培养体系。方法 将剔除动静脉及内膜的新鲜人脐带组织剪切成1mm3小块,用含10%胎牛血清的DMEM/F12培养,得到贴壁细胞。观察贴壁细胞形态,利用CCK-8试剂盒测定生长曲线,流式细胞术研究特殊细胞表面抗原CD29、CD73、CD90、CD105、CD31、CD14、CD34、CD45、CD11b、HLA-DR表达,并检测细胞周期。结果 体外培养7~10d后,可见细胞从组织块中游出;细胞主要呈纺锤体样、成纤维细胞样形态;生长曲线显示其增殖能力强;特殊表面抗原CD29、CD73、CD90、CD105表达强阳性,造血性抗原标志CD31、CD14、CD34、CD45、CD11b、HLA-DR表达均呈阴性,流式细胞仪周期检测结果显示G0/G1期细胞超过80%。结论 利用组织块法可有效获得人脐带MSCs,具有高纯度、低成本优点,并在体外较易培养、扩增。Objective To identify a detailed biological characterization of mesenchymal stem cells（MSCs）isolated from human umbilical cord（UC）tissue regarding their morphology,immunophenotype,purity and proliferative capacity and establish a reasonably cultured and amplified system.Methods After stripping off arteries and veins,the remaining parts of umbilical cord were cut into 1mm3 small sections and cultured with DMEM/F12 containing 10%fetal bovine serum.Adhere cells were obtained and the morphology of the cells was observed under inverted phase contrast microscope.The growth curves of them were drawn by CCK-8and the cell cycle and surface antigens（CD29,CD73,CD90,CD105,CD31,CD14,CD34,CD45,CD11 b,HLA-DR）were detected by flow cytometry.Results Seven to ten days after primary culture,adhere cells came out of fragments.The MSCs harvested were a high purity and mainly presented as a fibroblast-like morphology.UC-MSCs had a strong ability of proliferation through the cell growth curve.The special surface antigens CD29,CD73,CD90,CD105 were positive expression,while CD31,CD14,CD34,CD45,CD11 b,HLA-DR were negative.More than 80%cells of MSCs were found at G0/G1 phase.Conclusion Human UC-MSCs could be cultured and proliferated in vitro.

关 键 词：流式细胞术 人脐带间充质干细胞 培养 分离 组织工程 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]