中国组织工程研究

中国组织工程研究 ›› 2010, Vol. 14 ›› Issue (20): 3615-3619.doi: 10.3969/j.issn.1673-8225.2010.20.002

• 软骨组织构建 cartilage tissue construction • 上一篇    下一篇

脉冲磁场与前软骨干细胞向成熟软骨细胞的增殖与分化：与Ihh/PTHrP信号通路活性有关吗？

丁  然，游洪波，李  锋，孙  凯   

  1. 华中科技大学附属同济医院矫形外科，湖北省武汉市  430030
  • 出版日期:2010-05-14 发布日期:2010-05-14
  • 通讯作者: 游洪波，副教授，华中科技大学附属同济医院矫形外科，湖北省武汉市 430030 youhongbo003@yahoo.com.cn
  • 作者简介:丁 然，男， 1979年生，湖北省武汉市人，汉族，在读博士，主要从事脊柱关节研究。 dana_doctor@hotmail.com

Effects of pulsed electromagnetic fields on proliferation and differentiation of precartilaginous stem cells into mature chondrocytes: Whether do the effects correlate with the activity of Ihh-PTHrp signal pathway?

Ding Ran, You Hong-bo, Li Feng, Sun Kai   

  1. Department of Orthopaedic Surgery, Tongji Hospital, Huazhong University of Science and Technology, Wuhan   430030, Hubei Province, China
  • Online:2010-05-14 Published:2010-05-14
  • Contact: You Hong-bo, Associate professor, Department of Orthopaedic Surgery, Tongji Hospital, Huazhong University of Science and Technology, Wuhan 430030, Hubei Province, China youhongbo003@yahoo.com.cn
  • About author:Ding Ran, Studying for doctorate, Department of Orthopaedic Surgery, Tongji Hospital, Huazhong University of Science and Technology, Wuhan 430030, Hubei Province, China dana_doctor@hotmail.com

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摘要：

背景：前期研究证实脉冲电磁场对前软骨干细胞增殖具有促进作用，但其作用机制不明确。

目的：进一步探讨脉冲电磁场对前软骨干细胞增殖分化的影响，并观察其对前软骨干细胞Ihh/PTHrP信号通路活性的影响。

方法：采用免疫磁珠分选法获取并纯化SD大鼠前软骨干细胞，使用频率为50 MHz，电场强度为1 mT的脉冲电磁场进行干预，0.5 h/次，2次/d，干预2，4，6 d后收集细胞，以不给予磁场刺激的为空白对照。通过RT-PCR法检测细胞Ⅱ型胶原和聚集蛋白聚糖的基因表达水平，再通过Western-blot法测定各组细胞印度刺猬蛋白、甲状旁腺激素相关肽蛋白的表达水平。

结果与结论：RT-PCR结果显示，Ⅱ型胶原和聚集蛋白聚糖的表达随磁场刺激时间延长而增高，且经磁场刺激前软骨干细胞的聚集蛋白聚糖明显高于空白对照组(P < 0.05，P < 0.01)。Western-blot结果显示，印度豪猪蛋白及甲状旁腺激素相关肽蛋白表达均随磁场刺激时间的延长增加，不同时间磁场刺激组印度刺猬蛋白表达高于空白对照组 (P < 0.01)；甲状旁腺激素相关肽的表达量也随之上升，但当磁场刺激4，6 d后的甲状旁腺激素相关肽蛋白表达与空白对照组差异有显著性意义(P < 0.05，P < 0.01)。提示强度为1 mT，频率为50 MHz的脉冲电磁场能促进前软骨干细胞向成熟软骨细胞增殖分化，其作用机制可能与改变Ihh/PTHrP信号通路活性有关。

关键词: 前软骨干细胞, 种子细胞, ihh-PTHrp信号通路, 软骨组织工程, 脉冲磁场

Abstract:

BACKGROUND: Our early study has found that the effect of pulsed electromagnetic fields (PEMF) can promote the proliferation of the precartilaginous stem cells (PSCs). However, the mechanism remains unclear.

OBJECTIVE: To investigate the effect of PEMF on proliferation and differentiation of PSCs, and to explore the effect on activity of Ihh-PTHrP signal pathway.

METHODS: PSCs were obtained and purified using the method of magnetic-activated cell sorting, and the cells were then exposed in PEMF (1 mT, 50 MHz) for 30 minutes twice a day and the cells were continued to stimulate for 2, 4 and 6 days. Control group did not receive the stimulation of PEMF. Gene expression levels of type II collagen and aggrecan were detected by the method of RT-PCR and protein expression levels of Ihh and PTHrp were detected using Western blot method.

RESULTS AND CONCLUSION: The results of RT-PCR showed that with the time prolonging, the gene expression levels of type II collagen and aggrecan were increased. Comparing with the control group, aggrecan of cartilage stem cells were significantly increased before PEMF (P < 0.05, P < 0.01). Western blot indicated that protein expression levels of Ihh and PTHrp were increased with the time prolonging of PEMF stimulation, while Ihh protein expression in PEMF group was significantly higher than in the control group (P < 0.01). Additionally, PTHrp expression was also increased, but there was significant difference between PEMF and control group at days 4 and 6 after PEMF stimulation (P < 0.05, P < 0.01). This suggested that PEMF (1 mT, 50 MHz) could promote proliferation and differentiation from PSCs into mature chondrocytes, and the mechanism of this effect could probably correlate with the changes of Ihh-PTHrP signal pathway.

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