天津医药

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非病毒载体携带Cbfa1诱导成人骨髓间充质干细胞向成骨细胞分化的研究

孙杰1,魏学磊1,张志伟1,马士超1,马宝通1,赵文君2,邢国胜3,张凯1   

  1. 1. 天津市天津医院
    2. 天津市天津医院骨科研究所
    3. 天津医院
  • 收稿日期:2011-03-02 修回日期:2011-04-12 出版日期:2011-08-15 发布日期:2011-08-15
  • 通讯作者: 魏学磊

Osteogenic differentiation of human bone marrow mesenchymal cells induced with Cbfa1 through non-viral vector

  • Received:2011-03-02 Revised:2011-04-12 Published:2011-08-15 Online:2011-08-15

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摘要： 目的：观察核心结合因子a1（core-binding factor a1，Cbfa1）对成人骨髓间充质干细胞（human bone marrow mesenchymal cells，hBMSCs）向成骨细胞定向分化的诱导作用。方法：经密度梯度离心法和贴壁筛选法获得hBMSCs。非病毒载体FuGene HD携带Cbfa1转染hBMSCs后7、14d， real-time RT-PCR分析hBMSCs成骨细胞标志基因表达：Cbfa1、碱性磷酸酶（alkaline phosphase，ALP）、骨钙素（osteocalcin，OC）、骨桥蛋白（osteopontin，OPN）和I型胶原；培养1、2和3周， ALP染色、钙结节染色检测成骨分化。结果： Cbfa1转染hBMSCs表现出与成骨细胞相似的形态，成骨细胞标志基因ALP、OC 、OPN 和I型胶原的表达均明显上调，ALP染色阳性、并且有明显的钙结节形成。结论: 非病毒载体携带Cbfa1转染hBMSCs可诱导其向成骨细胞分化。　

关键词: Cbfa1, hBMSCs, 非病毒载体, FuGene, HD, 成骨分化

Abstract: Objective: To study the effect of core-binding factor a1 (Cbfa1) on the human bone marrow mesenchymal cells(hBMSCs) osteogenic differentiation. Methods: The hBMSCs were harvested through gradient centrifugation and adhering to the plastic culture wall. The third passage cells were used in our study. After the MSCs were infected with Cbfa1 through non-viral vector FuGene HD, the expressions of mRNA for osteoblast gene markers such as Cbfa1, alkaline phosphatase, osteocalcin, osteopontin and type I collagen were analyzed with real-time reverse transcriptase polymerase chain reaction (real-time RT-PCR ) at 7d and 14d. Alkaline phosphatase ( ALP ) activity and expression of osteocalcin were measured at 1, 2, and 3 weeks after transfection. Results: The real-time RT-PCR indicated that all the osteoblast gene markers were up-regulated in the Cbfa1 treatment group. The morphous of hBMSCs modified with Cbfa1 were similar to osteoblasts, and expressed ALP and osteocalcin. Conclusion: FuGene HD-Cbfa1 can directly promote the osteogenic differentiation of hBMSCs.

Key words: Cbfa1, hBMSCs, non-viral vector, FuGene HD, osteogenic differentiation