人脐血间充质干细胞的分离培养及成骨诱导活性  

Isolation of mesenchymal stem cells from human umbilical cord blood and its bone inducing activity in vitro

作　　者：张赫[1] 李昂[2] 饶国洲[2] 周洪[1] 

机构地区：[1]西安交通大学口腔医学院正畸科,陕西西安710004 [2]西安交通大学口腔医学研究中心,陕西西安710004

出　　处：《上海口腔医学》2010年第3期290-295,共6页Shanghai Journal of Stomatology

基　　金：陕西省"13115"科技创新工程重大科技专项计划项目(2008ZDKG-65)

摘　　要：目的:研究人脐血来源的间充质干细胞(HUCB-MSCs)的部分生物学特性,并考察不同血清浓度的培养基对原代细胞生长状况的影响和传代细胞向成骨样细胞分化的能力。方法:分离人脐血单个核细胞,倒置显微镜下观察原代细胞的生长状况和形态特点,并进行不同血清浓度培养的比较研究,免疫组化法测定表型;传代细胞定向诱导并分析细胞的分化情况。实验数据采用SPSS16.0软件包进行处理,方法为重复测量随机区组设计的方差分析。结果:原代细胞培养呈现4种形态;免疫组化染色显示,所有形态的细胞CD34、CD45均呈阴性,CD44、CD105呈阳性;当培养基中血清含量为15%时,体外培养HUCB-MSCs较为适宜;传代细胞可以在体外诱导为成骨样细胞,可见钙化结节及ALP活性水平的变化。结论:HUCB-MSCs原代细胞存活能力较强,但增殖活性略低,能达到稳定增殖的时间较长;4种形态细胞的部分免疫表型具有一致性,可能为HUCB-MSCs的4种亚型;血清浓度控制对HUCB-MSCs原代培养有着重要影响;传代细胞在一定条件下可向成骨样细胞分化。PURPOSE： To investigate the biological characters of mesenchymal stem cells obtained from human umbilical cord blood（HUCB-MSCs）, and detect the influence of different serum concentrations of culture medium to primary HUCB-MSCs’ growth kinetics and the osteogenic differentiation capacity of the passage cells. METHODS： The mononuclear cells were isolated from umbilical cord blood （UCB）, the growth characters and morphology of the primary MSCs were observed. Primary cells were cultured in α -MEM with different serum concentrations, and immunophenotype was investigated. P3 cells were induced to osteoblast-like cells and the differentiation results were analysed. SPSS 16.0 software package was used for statistical analysis through repeated measures analysis of variances. RESULTS： Four subpopulations with different morphology could be distinguished in primary HUCB-MSCs. All the subpopulations were positive for CD44 and CD105, and negative for CD34 and CD45. HUCB-MSCs displayed faster and stable growth when cultured in α-MEM containing 15% serum. Passage cells of HUCB-MSCs could be induced into osteoblast-like cells, the formation of mineralized extracellular matrix and the positive staining of expression of ALP could be observed. CONCLUSIONS： HUCB-MSCs are high in viability but low in proliferation. HUCB-MSCs contain 4 subpopulations with different morphology. The appropriate concentration of serum in culture medium should be 15%. HUCB-MSCs could be induced into osteoblast-like cells in vitro. Supported by ＂13115＂ Technological Innovation Project of Shanxi Province （Grant No.2008ZDKG-65）.

关 键 词：脐血 间充质干细胞 细胞形态 成骨样细胞 体外培养 

分 类 号：R329[医药卫生—人体解剖和组织胚胎学]