脂肪干细胞转染Bmp2后成骨能力变化的研究

Study in the changes of osteogenic capacity of adipose stem cells after transfection with Bmp2

目的探究过表达Bmp2对脂肪干细胞成骨基因表达及成骨能力的影响。方法使用脂质体转染过表达Bmp2质粒至大鼠脂肪干细胞,以相应空载质粒作为对照,分为空载对照组(Vector组)、过表达Bmp2组。采用Western Blot、免疫荧光对比两组成骨相关基因表达情况,使用茜素红与碱性磷酸酶染色鉴定脂肪干细胞矿化程度。结果茜素红染色结果显示,与Vector组相比,过表达Bmp2组细胞周围钙盐沉积明显增多,而碱性磷酸酶染色结果显示Bmp2组脂肪干细胞硫化钴沉淀增多,碱性磷酸酶活性强;Western Blot结果显示,与Vector组相比,过表达组细胞Bmp2、p-Smad1/5、Dlx2、OPN的表达显著提高(P<0.05),而smad1/5的表达则无明显统计学差异;免疫荧光标记OPN发现,与Vector组相比,OPN荧光强度明显增强(P<0.05)。结论过表达Bmp2促进了脂肪干细胞中相应成骨基因的表达以及其成骨作用,可为将来临床骨缺损修复提供潜力治疗手段。

Objective To explore the effect of overexpression of Bmp2 on osteogenic gene expression and osteogenic capacity of adipose stem cells.Methods Bmp2 was transfected into rat adipose stem cells with liposomes,and the corresponding null plasmids were used as controls(Vector group),divided into null control and overexpression Bmp2 groups.Western Blot and immunofluorescence were used to compare the expression of bone-related genes in both groups and the degree of adipose stem cell mineralization was identified using alizarin red and alkaline phosphatase staining.Results The results of alizarin red staining showed a significant increase in calcium salt deposition around the cells in the Bmp2 group compared with the control group,while the results of alkaline phosphatase staining showed an increase in cobalt sulfide precipitation and strong alkaline phosphatase activity in the Bmp2 group;Western Blot results showed that the expression of Bmp2,p-Smad1/5,Dlx2,and OPN expression were significantly increased in the Bmp2 group compared with the control group(P<0.05),while the expression of smad1/5 was not statistically different;immunofluorescence labeling of OPN revealed that the fluorescence intensity of OPN was significantly enhanced compared with the control group(P<0.05).Conclusion Overexpression of Bmp2 promotes the expression of corresponding osteogenic genes in adipose stem cells and their osteogenic effects,which may provide potential therapeutic strategy for future clinical bone defect repair.