大鼠骨髓间充质干细胞培养方法的比较

Comparison of different culture methods for rat bone marrow mesenchymal stem cells

关键词:  细胞培养  大鼠  骨髓间充质干细胞  碱性成纤维细胞生长因子

Key words:  [Key words] 　cell culture  rat  bone marrow mesenchymal stem cells  basic fibroblast growth factor

摘要:

目的　比较4种培养条件下大鼠骨髓间充质干细胞（BMSCs）的生长与增殖性差异。方法　采用密度梯度离心法提取大鼠BMSCs，分别在DMEM-LG培养基、DMEM/F12培养基、DMEM-LG+10 ng/mL碱性成纤维细胞生长因子(bFGF)与DMEM/F12+10 ng/mL bFGF培养条件下贴壁培养，观察各代细胞的形态特征，绘制第4代细胞的生长曲线，利用流式细胞仪对各培养条件下的BMSCs进行细胞表面标志CD45、CD29和CD90的检测。结果　不同培养条件下的BMSCs生长速度不同。加入bFGF培养条件下细胞增殖速度较单用DMEM-LG或DMEM/F12培养条件下更快（P<0.05）。DMEM-LG+10 ng/mL bFGF的培养效果最佳，细胞密度高，生长状态良好。4种条件下培养的细胞均阳性表达CD29及CD90，阴性表达CD45。结论　短期培养过程中加入bFGF不引起BMSCs分化，DMEM-LG+10 ng/mL bFGF是较理想的BMSCs培养条件。

Abstract:

[Abstract] Objective　To study the growth and proliferation differences of bone marrow mesenchymal stem cells (BMSCs) in 4 different culture conditions. Methods　BMSCs were isolated from the bone marrow of rats by density gradient centrifugation, and cultured in DMEM-LG, DMEM/F12, DMEM-LG+10 ng/mL basic fibroblast growth factor (bFGF) and DMEM/F12+10 ng/mL bFGF respectively. The morphologies of each generation of BMSCs, the growth curve of the 4th generations and the expressions of CD45, CD29 and CD90 of BMSCs in each culture conditions were detected. Results　The growths of BMSCs were different among 4 culture conditions. There was statistical difference in the proliferation of BMSCs in the bFGF supplement groups in contrast to DMEM-LG and DMEM/F12 (P<0.05). DMEM-LG+10 ng/mL bFGF was the best culture condition as it achieved the highest cell density and best growth state of BMSCs. Flow cytometry analysis showed that BMSCs cultured in 4 different conditions highly expressed CD29 and CD90, but did not express CD45. Conclusion　Short-term supplement of bFGF did not lead to differentiation of BMSCs. DMEM-LG+10 ng/mL bFGF is an ideal culture condition for BMSCs.