健骨二仙丸介导间充质干细胞的成骨细胞定向诱导及其成骨活性

Effects of "JianGuErXianWan" on Osteoblasts Induced from the Human Umbilical Cord Blood Mesenchymal Stem Cells and its Osteogenic Potential

目的 :研究健骨二仙丸对间充质干细胞 (Mesenchymal Stem Cells,MSCs)成骨细胞定向分化及其成骨活牲的影响。方法 :采用标准 Ficoll-Hypaque技术分离人脐血 MSCs,以地塞米松、β-磷酸甘油和维生素 C为辅剂定向诱导成骨细胞 ,碱性磷酸酶、骨矿化结节、骨钙素及上清钙含量与 型胶原作为成骨细胞鉴定与活性评价的指标。另以健骨二仙丸或健骨二仙丸合地寒米松、β-磷酸甘油与维生素 C为辅剂诱导 ,用正常培养液作为空白对照 ,1 5 d后观测对比上述相关指标。结果 :健骨二仙丸不能单独诱导 MSCs向成骨细胞转化 ,对 MSCs成骨细胞诱导前后的细胞形态无任何影响。定向成骨细胞诱导后 ,健骨二仙丸能显著提高其标志性产物碱性磷酸酶、骨矿化结节和上清钙、骨钙素以及 型胶原的表达。结论 :健骨二仙丸作为补肾益气药能促进 MSCs成骨细胞定向转化 ,提高其成骨活性。

Objective:Mesenchymal stem cells (MSCs) are adult stem cells which contribute to the regeneration of mesenchymal tissues such as bone,cartilage,muscle,et al.Due to the Multilineage cells potential and self-renewal capacity,the cells can be applied in many fields,such as the therapeutic agents for repairing critical-size bone defects and gene therapy.Here we report that the osteoblasts which were induced from human umbilical cord blood MSCs and the effects of the Traditional Chinese Medicine,JianGuErXianWan(JGEXW) which is a formula of invigorating the kidney,on the processes of inducing and the cell's osteogenic potential.Methods:The mononuclear fraction was isolated using standard Ficoll-Hypaque techniques.The cells were purified,proliferated and cultured in the same culture medium of L-DMEM.In order to induce the osteoblast cells,we applied JGEXW as the agent for the first group,the Dexamethasone(Dex),β-glycerophosphate(β-GP) and ascorbic acid-2-phosphate (VitC) for the second group,and JGEXW combine with Dex,β-GP and VitC for the third group,and the normal medium for the control group.Alkaline Phospatase(ALP),type Ⅰ Collagen,Osteocalin and in vitro mineralization were applied to identify the osteoblasts and evaluate the activity of them.Results:The osteoblsats induced from MSCs have a positive expressing for known markers:ALP and in vitro mineralization,and also stable expressing of ALP,osteocalcin and typeⅠ Collagen.JGEXW can not induce osteoblast independently,but it can improve the efficiency of Dex,β=GP and VitC on MSCs and the osteogenic potential of oseoblasts.Conclusion:MSCs can be induced into osteoblasts in a suitable condition,JGEXW activate the effect of Dex,β-GP and VitC on MSCs and the osteogenic potential of oseoblasts.