猕猴骨髓基质干细胞体外培养及其生物学特性的研究  

Study on the culture and biological characteristics of bone marrow stromal cells form Macaca mulatta in vitro

作　　者：胡军[1] 刘小林[1] 朱家恺[1] 邓宇斌[2] 向剑平[1] 王建云[1] 邹一丰[2] 

机构地区：[1]中山大学附属第一医院骨科-显微外科医学部,广州市510080 [2]中山大学中山医学院病理生理教研室

出　　处：《中华显微外科杂志》2004年第2期113-116,i003,共5页Chinese Journal of Microsurgery

基　　金：国家自然科学基金 (30 1 70 962 );广州市科技攻关重大项目 (2 0 0 2Z1 E0 0 35)

摘　　要：目的　通过体外分离、培养猕猴骨髓基质干细胞 (BMSCs) ,研究其生物学特性、表面标志和遗传学性能 ,为BMSCs用作组织工程化神经种子细胞提供研究基础。　方法　通过密度梯度离心分离猕猴BMSCs ,体外培养、扩增 ,用倒置相差显微镜进行形态学的观察 ,通过绘制生长曲线、计算克隆形成率来研究细胞的活力和增殖能力 ,流式细胞仪检测表面标志 ,核型分析反应遗传学性能。　结果　密度梯度离心的方法能获得较纯的猕猴BMSCs ,第 7代以前的细胞有较强的活力和增殖能力 ,具有与人的BMSCs相似的形态和表面标志 ,体外培养第 10代细胞仍稳定为二倍体核型。　结论　用本实验方法能较容易地获得大量的猕猴BMSCs ,具有与人的BMSCs相似的形状和表面标志 ,第 3～ 6代的细胞可满足以猕猴为动物模型的组织工程化神经种子细胞的研究要求。Objective To investigate the proper conditions of isolation and culture of bone marrow stromal cells(BMSCs) from macaca mulatta,and study on the biological characteristics of them to further application in tissue engineered peripheral nerve(TEPN). Methods BMSCs were isolated from marrow suspension by discontinuous gradient centrifugation on Ficoll-paque at a density of 1.073 g/ml,cultured in Dulbecco's Modified Eagle's Medium/low glucose(L-DMEM) containing 100 ml·L -1 fetal bovine serum. The morphology was inspected by inverted phase contrast microscope,and the antigentic phenotyper were analysized by FACScan. Results The isolated and cultured BMSCs from macaca mulatta were morphologically like fibroblast cells,and the cells of pre-passage 7 had a strong self-renewal capacity. BMSCs were positive for CD29,CD105 and CD166,and negative for CD34,CD80,CD86,CD71 and CD44. The cells of passage 10 still had 21 pairs of chromosomes,which were normal karyotype. Conclusions BMSCs from macaca mulatta had been easily obtained and proliferated by our experiment methods. They had a similar morphology to human MBSCs,and displayed an array of cell-surface antigens similar to BMSCs from rats and humans,and the cells of passage 3～6 were suitable for the study on the seed cells of tissue engineered peripheral nerve.

关 键 词：猕猴 骨髓 基质干细胞 体外培养 生物学 细胞培养 神经缺损 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]