中国组织工程研究

中国组织工程研究 ›› 2012, Vol. 16 ›› Issue (36): 6651-6657.doi: 10.3969/j.issn.2095-4344.2012.36.001

• 骨髓干细胞 bone marrow stem cells •    下一篇

骨髓间充质干细胞成骨分化中血管内皮生长因子和骨形态发生蛋白2的表达

刘 傥，张湘生，张 庆，雷 霆，熊光仲   

  1. 中南大学湘雅二医院，湖南省长沙市 410011
  • 收稿日期:2012-03-05 修回日期:2012-04-09 出版日期:2012-09-02 发布日期:2012-09-02
  • 通讯作者: 张庆，硕士，主任医生，副主任，中南大学湘雅二医院骨科，湖南省长沙市 410011 zhangqing65230@126.com
  • 作者简介:刘傥☆，男，1981年生，湖南省汉寿市人，汉族，2009年中南大学毕业，博士，主治医师，主要从事骨不连、组织工程及骨肿瘤的临床与基础研究。 liutang1204@126.com

Expression of vascular endothelial growth factor and bone morphogenetic protein 2 during osteogenic induction of rabbit bone marrow mesenchymal stem cells

Liu Tang, Zhang Xiang-Sheng, Zhang Qing, Lei Ting, Xiong Guang-zhong   

  1. The Second Xiangya Hospital of Central South University, Changsha 410011, Hunan Province, China
  • Received:2012-03-05 Revised:2012-04-09 Online:2012-09-02 Published:2012-09-02
  • Contact: Zhang Qing, Master, Chief physician, the Second Xiangya Hospital of Central South University, Changsha 410011, Hunan Province, China zhangqing65230@126.com
  • About author:Liu Tang☆, Doctor, Attending physician, The Second Xiangya Hospital of Central South University, Changsha 410011, Hunan Province, China liutang1204@126.com

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460

被引次数

11

摘要：

背景：有研究表明血管内皮生长因子可促进内皮细胞增殖、分化和迁移，而骨髓间充质干细胞经诱导分化后细胞自身的血管内皮细胞生长因子及骨形态发生蛋白2如何表达，却鲜见报道。
目的：观察兔骨髓间充质干细胞成骨诱导分化过程中血管内皮细胞生长因子和骨形态发生蛋白2的表达。
方法：取第3代新西兰大白兔骨髓间充质干细胞，1 d后将细胞分为两组：对照组不更换培养基；诱导组加入成骨诱导培养基进行成骨诱导，流式细胞仪检测其表面抗原。RT-PCR和Western-Blot检测细胞成骨分化过程中血管内皮细胞生长因子和骨形态发生蛋白2的表达。
结果与结论：兔骨髓间充质干细胞成骨诱导后，细胞形态由长梭形逐渐变为短梭形，多角形，最后成集落层叠生长；在诱导过程中，碱性磷酸酶活性逐渐增高，与对照组比较有显著性差异(P < 0.05)；诱导7 d时Ⅰ型胶原免疫细胞化学染色、14 时骨钙素免疫细胞化学染色、21 d时钙化结节染色阳性；RT-PCR和Western-Blot结果显示两组均有骨形态发生蛋白2和血管内皮细胞生长因子表达，诱导后第7，14，21天表达比较，差异有显著性意义(P < 0.05)，第7天，两者表达最强。说明兔骨髓间充质干细胞分离、提取操作简便易行，且有较强的生长增殖能力，可诱导分化为成骨细胞。其在成骨诱导过程中，骨形态发生蛋白2和血管内皮生长因子表达均先增强后减弱，血管内皮细胞生长因子表达可能对血管生成有促进作用。

关键词: 骨髓间充质干细胞, 组织工程, 骨形态发生蛋白2, 血管内皮细胞生长因子, 成骨分化, 干细胞

Abstract:

BACKGROUND: Studies have shown that vascular endothelial growth factor can promote endothelial cell proliferation, differentiation and migration, but the reports describing how the vascular endothelial cell growth factor and bone morphogenetic protein 2 express in the cells differentiated from bone marrow mesenchymal stem cells are rare.
OBJECTIVE: To observe the expression of vascular endothelial growth factor and bone morphogenetic protein 2 during osteogenic induction of rabbit bone marrow mesenchymal stem cells.
METHODS: Passage 3 New Zealand white rabbits were used for bone marrow mesenchymal stem cells extraction. The cells were divided into two groups at 1 day after extraction: the culture medium was not changed in the control group; the induction group was induced with osteogenic medium, and flow cytometry was used to detect the expression of cell surface antigen. RT-PCR and Western blot were used to detect the expression of vascular endothelial growth factor and bone morphogenetic protein 2 during osteogenic induction.
RESULTS AND CONCLUSION: During osteogenic induction of rabbit bone marrow mesenchymal stem cells, cell morphology was gradually changed from a long fusiform to short fusiform and polygonal, and finally stacked into colony growth. During the induction process, alkaline phosphatase content was gradually increased in the induction group, and there was a significant difference when compared with the control group (P < 0.05). Positive immunocytochemical staining of type Ⅰ collagen, osteocalcin and calcium node could be seen at 7, 14 and 21 days respectively. RT-PCR and Western blot showed that vascular endothelial growth factor and bone morphogenetic protein 2 were expressed in two groups, and there was significant difference at 7, 14 and 21 days after induction (P < 0.05). The expression of vascular endothelial growth factor and bone morphogenetic protein 2 peaked at 7 days after osteogenic induction of bone marrow mesenchymal stem cells. The method of isolation and culture of bone marrow mesenchymal stem cells is simple and feasible. Bone marrow mesenchymal stem cells can be induced to differentiate into osteoblasts with strong growth and proliferation abilities. During osteogenic induction of bone marrow mesenchymal stem cells, vascular endothelial growth factor and bone morphogenetic protein 2 expression first increased and then decreased. The expression of vascular endothelial growth factor may play a role in promoting angiogenesis.

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