中国组织工程研究

中国组织工程研究 ›› 2012, Vol. 16 ›› Issue (34): 6313-6316.doi: 10.3969/j.issn. 2095-4344.2012.34. 009

• 药物控释材料 drug delivery materials • 上一篇    下一篇

壳聚糖载体耦联骨形态发生蛋白2目的基因转染MC3T3-E1细胞后细胞生物学变化

赵 刚，高 雪，莫宏兵，平玉卓，刘可俗   

  1. 赵刚★，男，1975年生，黑龙江省依兰县人，汉族，2005年佳木斯大学毕业，硕士，副教授，主要从事错颌畸形的预防、诊断及矫治研究。
    zhaogangkq@ 126.com
  • 收稿日期:2012-01-05 修回日期:2012-03-10 出版日期:2012-08-19 发布日期:2012-08-19
  • 作者简介:赵刚★，男，1975年生，黑龙江省依兰县人，汉族，2005年佳木斯大学毕业，硕士，副教授，主要从事错颌畸形的预防、诊断及矫治研究。 zhaogangkq@ 126.com

Biological changes of mesenchymal cells 3T3-E1 transfected by chitosan coupled bone morphogenetic protein-2 target gene

Zhao Gang, Gao Xue, Mo Hong-bing, Ping Yu-zhuo, Liu Ke-su   

  1. Stomatology Medical College of Jiamusi University, Jiamusi 154002, Heilongjiang Province, China
  • Received:2012-01-05 Revised:2012-03-10 Online:2012-08-19 Published:2012-08-19
  • About author:Zhao Gang★, Master, Associate professor, Stomatology Medical College of Jiamusi University, Jiamusi 154002, Heilongjiang Province, China zhaogangkq@ 126.com

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299

被引次数

2

摘要：

背景：目前应用生长因子进行基因治疗成为最有前途的研究领域之一。骨形态发生蛋白具有诱导间充质细胞向成骨细胞方向分化，进而诱导新骨形成的能力，在骨愈合过程中发挥着重要作用。
目的：选用MC3T3-E1细胞进行体外转染，以壳聚糖为输送载体将骨形态发生蛋白2导入MC3T3-E1细胞，以实现骨诱导蛋白的局部持续表达，促进细胞的增殖与分化。
方法：采用复凝聚法制备壳聚糖-骨形态发生蛋白2复合体，琼脂糖凝胶电泳检测壳聚糖结合骨形态发生蛋白2的能力。采用壳聚糖纳米颗粒载体转染技术，将携带有骨形态发生蛋白2基因的表达质粒导入成骨细胞中，评价转染效率，同时运用四氮唑盐法检测细胞增殖情况以及细胞内碱性磷酸酶活性。
结果与结论：电泳图显示壳聚糖能很好地结合骨形态发生蛋白2质粒；壳聚糖可成功将骨形态发生蛋白2目的基因导入成骨细胞，且转染率达35%左右；四氮唑盐法实验结果提示转染后可促进成骨细胞增殖；碱性磷酸酶结果显示细胞碱性磷酸酶活性增强。提示壳聚糖纳米粒载体系统成功耦联骨形态发生蛋白2目的基因并有效转染MC3T3-E1细胞，同时促进MC3T3-E1细胞的增殖与分化。

关键词: 壳聚糖, MC3T3-E1细胞, 骨形态发生蛋白2基因, 转染, 增殖

Abstract:

BACKGROUND: It has become one of the most promising fields for the application of growth factor in gene treatment. Bone morphogenetic protein (BMP) can induce mesenchymal cells into osteoblasts and new bone formation, which plays an important role in bone healing.
OBJECTIVE: To realize the continuous expression of BMP partly and to promote cell proliferation and differentiation through transfecting mesenchymal 3T3-E1 cells (MC3T3-E1) in vitro with BMP-2 via the chitosan-based carrier.
METHODS: Chitosan-BMP-2 complex was prepared by complex coacervation method. The ability of chitosan combined with BMP-2 was tested by agarose gel electrophoresis. The expression plasmid carrying BMP-2 gene was induced into osteoblasts by using chitosan nanoparticle vector transfection technique. The transfection efficiency was evaluated. Cell proliferation and intracellular alkaline phosphatase activity were tested by MTT assay.
RESULTS AND CONCLUSION: Electrophoresis diagram showed that chitosan could well combined with BMP-2 plasmid. BMP-2 target gene could be induced into osteoblasts successfully by chitosan, and the tranfection efficiency was about 35%. MTT assay results exhibited that alkaline phosphatase activity was increased. These findings suggest that chitosan nanoparticles carrier system can successfully coupled BMP-2 target gene and effectively transfected MC3T3-E1, as well as promote MC3T3-E1 proliferation and differentiation.

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