扩增对骨髓间充质干细胞向成骨细胞分化的影响

Effect of extensive subcultivation on differentiation of bone marrow osteobiast in vitro

目的 :评价体外扩增对兔骨髓间充质干细胞向成骨细胞分化的影响 ,为骨组织工程提供功能活跃的种子细胞。方法 :分离、培养新生兔骨髓间充质干细胞 ,将扩增至2、6、10代 (P2、P6、P10)骨髓间充质干细胞分别用含地塞米松、β_甘油磷酸钠和维生素C的条件培养液进行培养 ,观察它们经诱导培养后的增殖能力、碱性磷酸酶活性和钙化结节形成量的变化。结果 :低代细胞 (P2、P6)间的细胞增殖率、碱性磷酸酶活性、钙化结节形成量无显著不同 ;高代细胞 (P10)与低代细胞比较 ,其增殖率明显降低 (P<0.05) ,碱性磷酸酶活性及钙化结节量也有降低趋势 (P>0.05)。结论 :骨髓间充质干细胞在10代内扩增 ,均能诱导分化为成骨细胞 ,可作为骨组织工程种子细胞。

Objective:To evaluate the effect of extensive subcultivation on osteoblastic defferentiation of rabbit bone marrow mesenchymal stem cells in vitro and provide active seeding_cells for bone tissue engineering.Methods:The mesenchymal stem cells were isolated from the thighbone marrow of new born rabbit and cultured and subcultured.The subcultured cells of 2,6 and 10 passage were respectively assayed for both the growing rate and osteogenic potedtial in respose to an osteoinductive medium containing dexamethasone(10 nmol.L-1),ascorbic acid (50mg.L-1)and β-glycerophosphate(1mmol.L-1).The osteogenic potentiall of cells was evidenced by the activity of alkaline phosphates(ALP) and formation of mineralized node were no statistical defferent between the P2,and P6 cells of low passages.Compared with the low passages,the rate of growth in p10 of high passage was decreased (P<0.05),the activity of ALP and numbers of mineralized node became decreased potential but was no statistical different(P>0.05).Conclusion:With 10 passages process of extensive subcultivation,the cells were remained ability of progressing through the osteogenic lineage and could be provided seeding cells for bone tissue engineering,but the activity of high passage cells was receded with different grade.