兔骨髓基质细胞的体外培养

目的 :比较用全骨髓法和离心法培养兔原代骨髓基质细胞 (BMSC)的差异 ,同时观察BMSC在体外培养时的生长特征及体外诱导为成骨细胞的可能性。方法 :抽取新西兰大白兔骨髓 ,分别用全骨髓法和密度梯度离心法进行原代培养 ,比较第 12d收获细胞的数量。传代后观察 1～ 6代细胞的生长特征 ,绘制生长曲线 ,测定分裂指数和贴壁率 ;同时将部分第 3代细胞进行诱导培养 ,第 16d计算碱性磷酸酶 (ALP)阳性细胞率。结果 :全骨髓法较离心法收获细胞数量少 (P <0 .0 5 ) ,1～ 6代细胞的生长特征相似 ,增殖能力强。第 3代细胞被成功地诱导为成骨细胞 ,第 16dALP阳性细胞率为 80 %。结论 :离心法较全骨髓法培养BMSC可得到较多的细胞 ,两种方法得到的细胞前 6代细胞均有较强的增殖能力 ,并可以诱导为成骨细胞 ,可作为骨组织工程用的种子细胞

The Culture of Bone Marrow Stromal Cell in Vitro

Objective: To compare the difference of all bone marrow culture (ABMC) and isolated bone marrow culture (IBMC) in bone marrow stromal cell(BMSC) primary culture,and at the same time to observe the growth characteristics and the proberbility of BMSC being induced into the ostoblasts. Methods: Bone marrow was aspirated from the New Zerland white rabbits, part of BM were cultured directly and the other part were isolated from the nucleated cells fraction by density gradient centrifuge, then cultured in the same medium. The cell number was counted after 12 days primary culture. Subculture was done for 5 passage. The following data were obtained,growth cure,division rate and adhensive rate. Part of the third passage were cultured in inducing medium. The rate of alkaline phosphetase positive cells was counted after 16 days. Results: The BMSC was less in ABMC than IBMC(P<0.05). The growth cures of passage 1 to 6 were almost similar. BMSC showed active proliferative capacity in vitro culture. The third passage cells were induced into ostoblasts succesfully. The rate of ALP positive cell got to 80% after inducing culture. Conclusion: We can get more BMSC through IBMC than ABMC. BMSC obtained from two methods both shows high ability of proliferation before 6 passage in vitro culture and can be used as the seeded cells in the bone tissue engineering.