不同传代倍数成人间充质干细胞成骨分化能力的研究

目的:观察不同传代倍数成人骨髓间充质干细胞(MSC)定向诱导分化为成骨细胞的能力,为自体MSC修复骨组织损伤的临床应用提供部分参数。方法:采用全髓直接接种法分离培养胸科手术取下肋骨骨髓,贴壁细胞达90%以上融合时消化传代。传代细胞部分以1×106/ml的细胞密度接种于含100ml/L胎牛血消(FBS)的L-DMEM的培养基中继续培养,传代;部分在培养基中添加成骨诱导剂地塞米松,β-甘油磷酸钠及抗坏血酸向成骨细胞诱导分化。如此连续传10代。结果:5代以内成人MSC成骨分化能力无显著差异(P>0.05),以第3代最强,第6代后MSC成骨分化能力逐渐减弱。结论:不同代次的MSC诱导分化为成骨细胞的能力不同,6代以后明显减弱。做为组织工程的种子细胞,成人MSC体外培养不宜超过5代。

Study for the capability of osteogenic differentiation from distinct transfer of culture power adult mesenchyme stem cell

Objective: To observe the capability that succeeds the osteoblast orientation inductive differentiation from the aniso-transfer of culture power adult mesenchyme stem cell,and to supply some part test parameter of clinical application to self MSC plerosis in the bone tissue damage. Methods: With holo-marrow direct inoculation methods, the fresh rib bone marrow that was taken off by the chest surgical department operation was isolated and cultured. When the adherence cells were confluence above 90%,, then those were digested and went down to posterity. All passage cells were separated into 1 ~ 106/ml cell density, and then were inoculated in conventional culture that was L-DMEM to be contained 100ml/L fetus ox blood serum (FBS) and went on cultivation and passage. The portion was added to osteogenic inducer Desamethasone, β-Sodium Glycerophosphate and Ascorbicacid in the culture medium, and all along succeeded osteoblast inductive differentiation. Such in this way was continuously passed on 10 generation times. Results: In 5 generation times the capability of osteogenic differentiation from human MSC there were no significant difference (P>0.05). It was the mightiest with 3rd generation, The capability of osteogenic differentiation from human MSC after 8th generation were to weaken one by one. Conclusion: The capability of osteogenic differentiation is changed with the different osteoblast human MSC's generation times, which is obviously to abate after 8th generation. As the seed cell of the constitution engineering, MSCs in vitro cultivation is not suitable for more than 5 generations.