脐血来源富血小板血浆对脐血间充质干细胞增殖的影响

Effect on umbilical cord blood platelet- rich plasma promoting proliferation of umbilical cord blood mesenchymal stem cells

目的：初步探讨脐血来源富血小板血浆促进脐血间充质干细胞增殖及增殖最佳浓度。方法：收集足月健康剖宫产新生儿脐带血,采用组织块培养法进行脐血间充质干细胞的分离培养;采用二次离心法提取脐血富血小板血浆。用ELISA方法检测富血小板血浆中转化生长因子（transforming growth factor,TGF）-β1。富血小板血浆联合10%胎牛血清培养脐血间充质干细胞,根据富血小板血浆中的TGF-β1浓度将实验分为6组：2 000 pg/ml组、1 000pg/ml组、750 pg/ml组、500 pg/ml组、250 pg/ml组、10%胎牛血清组。脐血间充质干细胞接种在96孔板,连续培养7d,分别在1、3、5、7 d用CCK8试剂盒测定不同浓度富血小板血浆对脐血间充质干细胞的增殖影响,统计分析,选出最佳浓度。结果：不同浓度富血小板血浆联合10%胎牛血清培养脐血间充质干细胞呈现不同的增殖速度,500~1 000pg/ml组明显优于其他浓度组,第5天开始差异有统计学意义（P〈0.05）。结论：富血小板血浆可提高脐血间充质干细胞的增殖活性,且呈剂量依赖性。

Objective To explore the best concentration of umbilical cord blood derived platelet rich plasma for promoting the proliferation and proliferation of umbilical cord blood mesenchymal stem cells. Method Umbilical cord blood was collected in term health cesarean selection newborn. Separation of umbilical cord blood mesenchymal stem cells was performed by using the tissue pieces culture. Platelet- rich plasma from umbilical cord blood was extracted with the use of secondary centrifugation. Transforming growth factor- beta 1 in platelet- rich plasma was detected with the method of ELISA. In this experiment,platelet rich plasma combined 10% fetal bovine serum was used to cultivate umbilical cord blood mesenchymal stem cells. According to the concentration of TGF- beta 1 in platelet- rich plasma,the experiment was divided into 6 groups：2 000 pg / ml,1 000 pg / ml,750 pg / ml,500 pg / ml,250 pg / ml,10% fetal bovine serum group. Umbilical cord blood mesenchymal stem cells were incubated in 96- well plates,and cultured for 7 days. After 1,3,5,and 7 days later,CCK8 kit was used to determinate the proliferation effect of mesenchymal stem cells. Meanwhile,statistical analysis was performed to select the best concentration. Results Different concentrations of platelet- rich plasma combined 10% fetal bovine serum resulted in varied proliferation rate from umbilical cord blood mesenchymal stem cells. The findings suggested that the proliferation rate in 500 ~1 000 pg / ml concentration groups was superior to that of other groups. It was not 5- day cultured until there was statistically significant（ P〈0. 05）. Conclusion Platelet- rich plasma can improve proliferation of the umbilical cord blood mesenchymal stem cells. Furthermore,the activity shows a dose dependent.