Zhang Xiao-Qiang, Li Xu, Wu Tao, Li Jian-Wei, DU Hao, Pei Guo-Xian
Department of Orthopedics and Traumatology, Nanfang Hospital. Southern Medical University, Guangzhou, China.
Nan Fang Yi Ke Da Xue Xue Bao. 2009 Mar;29(3):419-22.
To explore the isolation, in vitro culture and chondrogenic differentiation of goat bone marrow mesenchymal stem cells (BMSCs).
Bone marrow was harvested from a 10-month-old Chinese goat for adherent culture of the BMSCs in vitro. Flow cytometry was performed to detect the cell surface markers of the BMSCs of the fourth generation. The induction medium (containing 10% fetal bovine serum, high-glucose DMEM, 6.25 microg/ml insulin, 6.25 microg/ml transferrin, 50 microg/ml vitamin C, 100 nmol/L DXM and 10 ng/ml transforming growth factor-beta1) was then applied for chondrogenic differentiation. Cytochemical staining, RT-PCR and Western blotting were performed to detect the expressions of type II collagen and aggrecan in the cells at the time points of 0, 1, 2 and 4 weeks.
The goat BMSCs grow well in vitro with a high purity in the fourth generation. The expression of chondrocyte phenotypes were observed at 1, 2 and 4 weeks, which became more obvious as the culture prolonged. The mRNA and protein expression of type II collagen and aggrecan in the BMSCs increased obvious after the induction and had reached a satisfactory level by 2 weeks.
Goat BMSCs have the potential to differentiate into chondrocytes in vitro, and the results of this study provide the experimental basis for application of goat BMSCs in bone and cartilage tissue engineering in vivo.
探讨山羊骨髓间充质干细胞（BMSCs）的分离、体外培养及成软骨分化。
从10月龄中国山羊采集骨髓，进行BMSCs的体外贴壁培养。采用流式细胞术检测第4代BMSCs的细胞表面标志物。然后应用诱导培养基（含10%胎牛血清、高糖DMEM、6.25μg/ml胰岛素、6.25μg/ml转铁蛋白、50μg/ml维生素C、100nmol/L地塞米松和10ng/ml转化生长因子-β1）进行成软骨分化。在0、1、2和4周时间点进行细胞化学染色、RT-PCR和蛋白质印迹法检测细胞中Ⅱ型胶原和聚集蛋白聚糖的表达。
山羊BMSCs在体外生长良好，第4代纯度较高。在1、2和4周时观察到软骨细胞表型表达，且随着培养时间延长而更加明显。诱导后BMSCs中Ⅱ型胶原和聚集蛋白聚糖的mRNA和蛋白表达明显增加，到2周时达到满意水平。
山羊BMSCs在体外具有向软骨细胞分化的潜能，本研究结果为山羊BMSCs在体内骨和软骨组织工程中的应用提供了实验依据。
