摘要
目的建立大鼠骨髓间充质干细胞(bone marrow-derived mesenchymal stem cells,BMSCs)体外分离培养方法。方法采用全骨髓贴壁法分离培养BMSCs,进行形态学观察,绘制生长曲线,流式细胞仪分析细胞周期、检测细胞表面抗原标志物,并进行成脂、成骨分化诱导。结果获取的BMSCs形态呈均一成纤维细胞样,并呈集落样生长。生长曲线呈S形,细胞周期显示86.02%P3代细胞为G0/G1期,保持活跃的扩增能力。BMSCs高表达CD44、CD90、低表达CD34、CD45。成脂诱导21 d后,可见细胞的胞浆内出现大量红染脂滴。成骨诱导21 d后,可见大量橘红色矿化结节形成。结论全骨髓贴壁培养法可成功有效地分离培养BMSCs。
Objective To establish a simple method for isolation and expansion of rat BMSCs in vitro. Methods The morphology of cells that isolated by bone marrow adhesion separation,was observed with phase contrast microscope,and the growth curve was obtained depend on the cell number during expansion. Cell cycle and surface antigen markers were confirmed by Flow cytometry instrument. BMSCs could be induced toward osteoblast and adipocyte in special medium. Results The cultured BMSCs showed the typical fibroblast-like morphology and colony. Growth curve of cells resembled S shape.The passage 3 cells in G0/G1 accounted for 86.02%. Cells kept a high potential of multiplication. Ceils were positive for CD44 and CD90,but negative for CD34 and CD45.Following 21 days of adipogenic induction,oil red O staining showed that red lipid droplet existed in adipocytes. Following 21 days of osteogenic induction, cell alizarin red staining showed that alizarin red was positive in osteoblasts. Conclusion BMSCs can be separated and pu- rified effectively by bone marrow adherence culture.
出处
《中国现代医生》
2014年第1期7-10,共4页
China Modern Doctor
基金
浙江省科技计划项目(2011C13029-1)
关键词
骨髓间充质干细胞
全骨髓贴壁法
鉴定
[Key words] BMSCs
Bone marrow adherence culture
Identification
