BMP-7对大鼠骨髓间充质干细胞膜片成骨分化的影响

Effect of BMP-7 on osteogenic differentiation of rat bone marrow mesenchymal stem cells sheet

目的:探讨骨髓间充质干细胞(bone marrow mesenchymal stem cells,BMMSCs)膜片的生物学性能以及骨形成蛋白7(bone morphogenetic protein 7,BMP-7)对BMMSCs膜片成骨分化的影响。方法:分离培养SD大鼠原代BMMSCs,采用维生素C诱导构建大鼠BMMSCs膜片,实验分为自然分化组、添加BMP-7的骨诱导组以及标准骨诱导组,采用HE染色观察,CCK-8检测膜片细胞增殖情况以及RT-PCR检测细胞膜片成骨分化水平。结果:分离大鼠BMMSCs诱导10~14 d后获得白色膜状细胞膜片。镜下观察细胞呈梭形重叠生长,细胞间紧密连接并有大量细胞外基质(ECM)分泌。CCK-8显示膜片细胞密度随时间递增。与BMMSCs膜片相比,BMP-7诱导的细胞膜片高表达ALP、Osteocalcin等成骨基因。结论:BMP-7诱导的大鼠BMMSCs细胞膜片成骨分化能力强,在骨组织工程中优势显著。

Objective： To probe the biological properties of the contracted rat bone marrow mesenchymal stem cell（ BMMSCs） sheets and the effect of rh BMP-7 on osteogenic differentiation of BMMSCs sheets. Methods： Three groups were setted in the study including spontaneous differentiation group,bone induction group and BMP-7 suplied bone induction group. BMMSCs was harvested from SD rats and used to construct the rat BMMSCs sheets with vitamin C inducing,inverted microscope and HE staining was employed for morphological observation,CCK-8 assay and osteogenesis inducetion was examined to the proliferation and differentiation of the cell sheets. Related osteogenesis gene mRNA expression was examined by RT-PCR. Results： After 10 ~ 14 days culture of BMMSCs in vitro,a white membranaceous sheet was formed and lifted by aseptic forceps. BMMSCs grew in multilayer and retained their spindle shape. HE staining showed abundant extracellular matrix（ ECM）. The density of cells was increased by time. Compared with BMMSCs sheet,gene expression of ALP and OCN were significantly up-regulated in the cell sheets induced by rh BMP-7. Conclusion： The osteogenesis ability of rat BMMSCs sheet induced by rh BMP-7 is favorable,more advantages in the bone tissue engineering.