摘要
[目的]观察冲击波(shock waves)与地塞米松对自体血清培养的人骨髓间充质干细胞(hum an m esenchy-m al stem cells,hMSCs)体外成骨分化的影响比较。[方法]选择10名健康志愿者,每名健康志愿者抽取骨髓60 m l,抽取外周静脉血100 m l,然后分离出自体血清。把每名志愿者的骨髓分别用10%AS和10%FBS对hMSCs进行体外培养,通过相差倒置显微镜观察细胞形态、检测24 h细胞贴壁率、细胞倍增时间、检测细胞表面标记物、细胞周期及免疫细胞化学染色检测比较两组hMSCs的增殖状况。然后把AS培养的hMSCs两组分别用体外冲击波疗法(extracor-poreal shock waves therapy,ESWT)[0.36 m j/(mm2.100次)]和传统的诱导成骨培养基(10 nmol/L地塞米松、50umol/L抗坏血酸、10 mmol/Lβ-甘油磷酸钠)分别对hMSCs进行成骨诱导,对诱导好的细胞进行ALP定量检测、ALP染色、茜素红染色及RT-PCR测成骨基因(碱性磷酸酶、骨钙素、骨桥蛋白、骨结合素、I型胶原)表达比较两组成骨分化情况。[结果]在细胞增殖上,两组细胞各代在细胞形态、细胞贴壁率、细胞周期以及免疫细胞化学染色检测上无明显差异,但在细胞倍增时间、细胞增殖速率上,自体血清明显优于胎牛血清。在细胞分化方面,两组在ALP定量检测、ALP染色、茜素红染色及RT-PCR测成骨基因上,冲击波作用于人骨髓间充质干细胞的成骨效应明显优于地塞米松。[结论]冲击波与地塞米松相比较,具有更良好促hMSCs成骨分化作用。
[Objective]To compare effects on the osteoblasts of human mesenchymal stem cells（hMSCs） in autologous serum by extracorporeal shock waves versus dexamethasone.[Methods]60 ml iliac crest bone marrow and 100 ml peripheral blood from 10 healthy volunteers were collected and the autologous serum were from peripheral blood.The iliac crest bone marrow from each volunteer were cultured for hMSCs with 10% AS（the experiment group） and 10% FBS（the control group）,respectively.The cell morphology,the cell adhesion rate for 24-hour,the cell doubling time were observed by inverted microscope.The cellular surface antigen and the generation cycle were detected by flow cytometry and immunocytochemical staining,the hMSCs cultured by AS medium were induced to osteoblasts by method of ESWT（extracorporeal shock waves therapy） and the traditional method of dexamethasone（10 nM）,ascorbic acid（50 uM） and sodium glycerophosphate（10 mM）,respectively.The ALP staining and Alizarin Red staining were used to detect the activity of ALP and the the expression differences of oteogenic genes（including ALP,OP,OC,ON,Col I） were analyzed by RT-PCR after inducion.[Results]The two groups of hMSCs had no significant differences in cell morphology,cell adhesion rate,cell cycle and immunocytochemical staining during cell proliferation in AS or FBS medium.But the cell doubling time and cell proliferation rate of the hMSCs in AS medium was better than that of the hMSCs in FBS medium.The results of ALP quantitative,ALP staining,alizarin red staining and RT-PCR of oteogenic genes showed that the ESWT method was superior to the traditional method.[Conclusion]The ESWT is a better method to promote the osteoblastic differentiation of hMSCs than the traditional dexamethasone method.
出处
《中国矫形外科杂志》
CAS
CSCD
北大核心
2011年第9期758-764,共7页
Orthopedic Journal of China
基金
国家自然科学基金资助项目(编号:30371430)
关键词
人骨髓间充质干细胞
体外冲击波
地塞米松
自体血清
成骨分化
human mesenchymal stem cells
extracorporeal shock waves
dexamethasone
autologous serum
osteogenic differentiation
