条件培养对骨髓间质干细胞体外成骨潜能的影响

Influence of conditional culture medium on osteogenic potential of mesenchymal stem cells cultured in vitro

目的:骨髓间质干细胞(mesenchymalstemcells,MSCs)有多种分化的潜能,但其分化并非是一个完全自发的过程,探讨使用条件培养基对MSCs体外成骨潜能的影响,为优化骨组织工程的种子细胞提供实验依据。方法:采用条件培养(A组)与完全培养(B组)基两种方法将兔骨髓间质干细胞进行体外培养后,对各组细胞进行形态学观察、组织学检测以及采用碱性磷酸酶(ALP)与骨钙素检测试剂盒进行定量分析,以判断两种培养基对骨髓间质干细胞体外成骨能力的影响。结果:在细胞传代培养中A组细胞增殖速度明显低于B组。A组细胞随培养时间的延长细胞呈多层生长,不发生接触抑制现象。而B组随时间的延长,细胞间逐渐出现接触抑制,细胞分裂停止,逐渐死亡。ALP含量A组为(8.17±0.61)U/L,B组为(5.07±0.49)U/L。骨钙素含量A组为(25.38±1.06)μg/L,B组为(14.37±1.71)μg/L,A,B两组比较差异均有显著性意义(P<0.05)。结论:体外培养的MSCs具有一定的成骨能力,使用含有地塞米松、β-甘油磷酸钠和维生素C的条件培养基可以大大提高其成骨能力。

AIM: Mesenchymal stem cells(MSCs) have many differentiating potentials, but its differentiation was by no means a fully spontaneous process. To study the influence of conditional culture medium on osteogenic potential of cultured MSCs in vitro, which provides the experimental basis for optimizing the seed cell of bone tissue engineering. METHODS: The rabbit MSCs was cultured in vitro with two kinds of culture medium, one is conditioned medium(Group A), the other is pure culture medium(Group B).The cells in each group were examined by morphological and histological observation, analyzed with alkaline phosphatase(ALP) and bone Gla containing protein(BGP) testing kit to estimate the influence of the two mediums on osteogenic potential of MSCs in vitro. RESULTS: The increased speed of the cells in the Group A was significantly lower than that in the Group B in the process of passage culture. The cells in the Group A grew with many layers along with the prolongation of the culture time without contacted inhibition. But the cells in the Group B appeared contacted inhibition between the cells along with the prolongation of the culture time; stopped cell division and began to die gradually.The contents of ALP were (8.17±0.16) U/L and (5.07 ±0.49) U/L in the Group A and Group B respectively. The contents of BGP were (25.38±1.06) μg/L and (14.37 ±1.71) μg/L in the Group A and Group B respectively. There were all significant differences in both groups (P< 0.05). CONCLUSION: Cultured MSCs in vitro have osteogenic potential, which can be improved greatly with conditional culture medium containing dexamethasone,β sodium glycerophosphate and vitamin C.