马鹿茸间充质干细胞软骨分化过程表面标志物的特征性变化
Analysis of Cell Surface Markers in Differentiation of Antler Mesenchymal Stem Cells into Chondrocytes in Vitro

利用阿利新蓝、甲苯胺蓝、茜素红染色和免疫组化法鉴定鹿茸间充质干细胞（MSCs）不同分化阶段蛋白多糖、Ca2+和Ⅱ型胶原蛋白3种软骨细胞表面标志物的分泌量。结果表明，诱导第14~21 天细胞蛋白多糖分泌逐渐增加，阿利新蓝和甲苯胺蓝染色结果逐渐达强阳性，至第28 天时蛋白多糖胞外积累量开始下降；茜素红染色与免疫组化结果显示，诱导第14~35 天，细胞阳性结果均逐渐上升，第35 天茜素红染色与免疫组化检测阳性结果显著高于前3个阶段。表明：体外诱导鹿茸MSCs至第14 天软骨细胞开始出现，并分泌蛋白多糖和少量Ⅱ型胶原蛋白，第21 天软骨细胞大量出现，开始分泌少量Ca2+，至第28 天时软骨开始向骨分化，蛋白多糖分泌下降，Ca2+在胞外积累，第35 天软骨细胞完成终末分化进入钙化期。
Used four methods of alcian blue,toluidine blue,alizarin red staining and immunohistochemistry,three cells’ surface markers,including Proteoglycans,Ca2+ and Col Ⅱ,were identified on the 14th,21st,28th and 35th d mesenchymal stem cells(MSCs) by stimulating using growth factor beta1 (TGF-β1,10 ng/mL) in vitro. Alcian blue staining and toluidne blue staining revealed heavy positive staining in the cellular matrix from the 21th d,while its accumulations decreased after 28th d. By two methods of alizarin red staining and immunohistochemical staining,the amount of positive cells increased gradually from the 14th d to 28th d in the cartilage differentiation process. While results of the other stages showed significant difference. In conclusion,during the cartilage differentiation process,antler MSCs began to differentiate to secrete protein polysaccharide and a small amount of Col Ⅱ after induction for 14th d. After the 28th d osteogenic differentiation of antler cartilage cells began to be detect when protein polysaccharide decreased. After 21th d the cartilage cells began to secrete a small amount of Ca2+,amount of Ca2+ has accumulated in the extracellular along with the induction time. Antler MSCs were induced to terminally differentiate after 35th d.