兔骨髓间充质干细胞的制备及诱导成骨的实验研究

目的:在体外原代培养兔骨髓间充质干细胞,观察兔骨髓间充质干细胞的体外生长特性,为用于组织工程学的种子细胞培养提供实验基础。方法:自兔股骨、胫骨取骨髓,分离骨髓间充质干细胞,分别采用常规培养液及含矿化液的培养液两组进行培养,对第3代细胞进行酶动力学法碱性磷酸酶(ALP)活性检测及钙结节茜素红染色,观察细胞在体外长期培养时钙结节形成情况。结果:两组细胞其培养液中的ALP含量不断升高,经过两独立样本t检验,矿化液诱导组第2,6,10天的ALP水平较正常对照组均明显升高(P<0.05)。细胞表现出成骨细胞特性,在体外长期不传代培养时可形成钙结节。结论:兔骨髓间充质干细胞在体外适当的培养条件下,增殖能力很强,可以向成骨细胞方向分化,可作为组织工程学的种子细胞。

Study on induced bone formation of rabbit marrow mesenchymal stem cells

Objective: To establish primary bone marrow mesenchymal stem cells(BMSCs) of rabbit in vitro and observe its growth characteristics,so as to provide experimental framework to cultivate various seed cells for tissue engineering.Methods: Bone marrow was extracted from the femoral and shin bone of rabbit,BMSCs were separated from bone marrow and cultivated in two groups,that is DMEM media group and DMEM with mineralized liquid group respectively.To monitor the activity of alkaline phosphatase(ALP) of passage 3 BMSCs by means of enzyme kinetics and to observe BMSC-derived calcification nodule by staining with alizarin bordeaux.Results: ALP level in both groups increased gradually.Independent samples t test showed the ALP activity in the DMEM with mineralized liquid group on the 2nd,6th,10th day was significantly higher than that in DMEM media group(P<0.05).BMSCs had the characteristics of osteoblast and could form calcification nodule.Conclusion: Rabbit BMSCs can proliferate well under suitable nutrient condition,and can be induced to differentiate into osteoblasts in vitro as well,therefore they can be used as the seed cells for the bone tissue engineering.