补肾活血汤促骨折端间充质干细胞体外迁移及CCR2表达的研究

Research of Bushenhuoxue Decoction on Promoting the Migration of Mesenchymal Stem Cells in the Fracture Site and Expression of CCR2

目的:探讨补肾活血汤对骨折端间充质干细胞(Mesenchymal Stem Cells,MSCs)体外迁移及表面受体CC类趋化因子受体2(C-C motif chemokine receptor-2,CCR2)表达的影响。方法:补肾活血汤灌胃治疗大鼠股骨中段骨折模型7、14、21d,分别取骨折端MSCs进行培养,传至第3代采用Transwell体外迁移实验检测补肾活血汤对骨折端MSCs体外迁移的影响,Western Blot、qPCR检测CCR2的表达。结果:补肾活血汤治疗后第7、14天,骨折端MSCs体外迁移能力明显加强,与模型组、对照组比较,差异有统计学意义(P<0.05)。第21天,骨折端MSCs体外迁移能力明显减弱,与模型组比较,差异无统计学意义(P>0.05)。Western blot、qPCR结果显示,补肾活血汤治疗第7天,治疗组CCR2的表达明显高于模型组、对照组(P<0.05)。第14天,CCR2的表达略有上升,与治疗后7d比较,差异无统计学意义(P>0.05),但治疗组仍显著高于模型组、对照组(P<0.05)。第21天,CCR2的表达有所下降,与治疗后7d比较,差异有统计学意义(P<0.05)。但治疗组与模型组比较,差异无统计学意义(P>0.05)。实验结果还表明,对照组CCR2的表达均处于低水平,7、14、21d比较,差异无统计学意义(P>0.05)。结论:补肾活血汤可以促进大鼠骨折愈合过程中MSCs的体外迁移,其相关机制可能与上调CCR2的表达、激活MCP-1/CCR2信号轴有关。

Objective：To explore the effect of Bushenhuoxue decoction （BSHXD）on promoting the migration of mesenchymal stem cells （MSCs）in the fracture site and the expression of C-C motif chemokine receptor-2 （CCR2）. Methods：The rat femur fracture model was established and treated with BSHXD. Then the MSCs in the fracture site was isolated and cultured at 7th, 14th,and 21st day,respectively. At the third passage,Transwell assay was applied to examine the migration of MSCs influenced by BSHXD. The expression of CCR2 was detected by Western blot and qPCR. Results：BSHXD could promote significantly more migration of MSCs at 7th and 14th day, but not at 21st day, compared with the model group and control group （P〈0.05）. The results of Western blot and qPCR showed that the expression of CCR2 in the treatment group at 7th day was significantly higher than the model group and control group （P〈0.05）. At 14th day, the expression of CCR2 increased slightly compared with 7th day,with no statistically significant （P〉0.05）, but it was still significantly higher than the model group and control group （P〈0.05）. At 21st day, the expression of CCR2 was declined significantly compared with the expression at 7th day （P〈0.05）, which was not statistically different between the treatment group and the model group （P〉0.05）. The results also show that the expression of CCR2 in the control group were at a low level, there was no significant difference among 7 th, 14th, and 21 st day （P〉 0.05）. Conclusion： BSHXD could promote migration of MSCs during fracture healing, and its mechanism may be associated with the up-regulation of CCR2 and activation of MCP-1/CCR2 signaling axis.