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目的探讨大鼠骨髓间充质干细胞(BMSCs)的体外分离培养方法并鉴定其生物学特性。方法采用全骨髓贴壁细胞培养法进行BMSCs的分离培养,倒置相差显微镜下观察细胞形态及生长特性,绘制细胞生长曲线,并检测细胞在不同诱导条件下向成脂肪细胞和成骨细胞分化的能力。结果原代大鼠BMSCs培养24h后镜下观察大部分细胞已经贴壁,传至第三代,细胞形态均一,呈梭形、螺旋样排列。经过约2d的适应期,3～7d进入对数生长期,7d后进入平台期;成骨细胞诱导18d,茜素红染色可见明显钙结节,成脂肪细胞诱导2周后油红O染色可见明显脂质沉淀。结论全骨髓贴壁细胞培养法操作简单、快速,可以获取形态均一、纯度较高的BMSCs。
Objective To investigate the isolation and culture methods of rat bone marrow mesenchymal stem cells (BMSCs) in vitro and to identify their biological characteristics. Methods BMSCs were isolated and cultured by whole bone marrow adherent cell culture. The morphology and growth characteristics of BMSCs were observed under inverted phase contrast microscope. The cell growth curve was drawn and the ability of cells to differentiate into adipocytes and osteoblasts under different inducing conditions . RESULTS: Primary cultured rat BMSCs were observed under microscope for 24 hours. Most of the cells were adherent to the third generation. The cells were uniform in shape and spindle-shaped and arranged spirally. After about 2 days of acclimation, 3 ~ 7d into the logarithmic growth phase, into the plateau after 7d; osteoblasts induced 18d, alizarin red staining visible calcium nodules, adipose tissue 2 weeks after induction oil red O staining can be seen Obvious lipid precipitation. Conclusion Whole bone marrow adherent cell culture method is simple and rapid, and can obtain uniform morphology and high purity of BMSCs.