rhBMP-7对小鼠骨髓间充质干细胞向成骨细胞分化的影响

Effects of rhBMP-7 on osteoblast differentiation of murine bone marrow mesenchymal stem cells in vitro

目的:观察重组人骨形成蛋白-7(rhBMP-7)对小鼠骨髓间充质干细胞(BMSCs)向成骨细胞分化的影响。方法:采用密度梯度离心法分离骨髓,体外培养扩增,获得小鼠骨髓间充质干细胞,贴壁培养至第3代,向培养液中加入rhBMP-7,培养5 d后进行碱性磷酸酶(ALP)染色,ALP活性与骨钙素(OC)含量检测,并用RT-PCR法分析成骨细胞标志基因骨钙素(OC)与Ⅰ型胶原(Col-Ⅰ)表达情况,Western blot法检测Ⅰ型胶原(Col-Ⅰ)蛋白的分泌量。结果:rhBMP-7诱导组的骨髓间充质干细胞的ALP染色强度、ALP活性及OC含量明显升高,OC与Col-Ⅰ的mRNA表达水平以及Col-Ⅰ的蛋白表达水平均明显提高。结论:rhBMP-7可促进体外培养的小鼠BMSCs向成骨细胞方向分化。

AIM： To study the effects of recombinant human bone morphogenetic protein-7 （rhBMP-7） on osteo- blast differentiation of murine bone marrow mesenchymal stem cells （BMSCs）. METHODS： The BMSCs were collected from murine bone marrow by density gradient centrifugation. Cells were adherent cultured and expanded in vitro. RhBMP-7 was added into the culture medium of the 3rd passage BMSCs. Five days later, we performed alka- line phosphatase （ALP） staining and detected ALP activity and osteocalcin （OC） content. Osteoblast differentiation marker gene including OC and collage Ⅰ （Col-Ⅰ ） were assayed by RT-PCR. Total protein was isolated and the secretion of Col- Ⅰ in the cells was measured by Western blotting. RESULTS： The staining intensity of ALP in the group with rhBMP-7 was stronger than that in the negative control group. ALP activity and OC content of rhBMP-？ group were obviously higher than that of the negative control group. The mRNA of OC and Col-Ⅰ and the protein of Col-Ⅰwere highly expressed in the group induced by rhBMP-7. CONCLUSION： RhBMP-7 can induce murine BMSCs to differentiate into osteoblast in vitro.