诱导骨髓间充质干细胞向成骨细胞和成软骨细胞分化

Osteoge nic and chondrogenic differentiation of mesenchymal stem cells from swine bone m arrow

目的探讨小香猪骨髓间充质干细胞(MSC)向成骨细胞和成软骨细胞诱导分化的能力。方法选用早期贴壁培养的小香猪骨髓MSC,在传代两三次后加入条件培养基,观察了细胞的形态学变化,同时检测碱性磷酸酶、钙盐沉积以及细胞外基质和II型胶原的表达。结果MSC在骨形成蛋白(BMP)、地塞米松和β-磷酸甘油钠的作用下,两三天细胞从梭形变为圆形或类圆形,细胞体积增大,12～15d时细胞集落的表面可见有颗粒状物质分泌,碱性磷酸酶染色为阳性,20～25d时,分泌颗粒更加明显,核固红染色为阳性,表现出成骨细胞的特点;在培养体系中加入地塞米松和转化生长因子-β1后两三天,可见细胞由类成纤维状变为圆形或椭圆形状,有的呈肾性,体积增大,10d后可见细胞周围有基质分泌,II型胶原染色为阳性,表现为软骨细胞分化的特点。结论在一定培养条件下成功诱导小香猪骨髓MSC向成骨细胞和成软骨细胞分化,为我们进一步的研究打下基础。

Objective To explore osteogenic and chondrogenic potential of mesenc hy mal stem ce lls(MSC)derived from swine bone marrow.Methods MSC which derived from swine bone marrow with early adherent methods were induced osteogenic and chondrogeni c dif-ferentiation under conditional cul ture medium and were obser ved the change of morphology,alkaline phosphatase activity,matrix mineral-iza tion and expression of type II collagen and extracellular matrix.Results MSC be came more flattened and round morphology at 3days in the presence of bone morph ogenetic protein-2(BMP-2),dexamethasone,a-glycerophoshpate disodium sa lt-p entahydrate.By day 15and 25,his-tological analysis of the cells rev e aled strong staining with alkaline p hosphatase method and alizarin red c onfi rmed that the cells had osteogen ic potential.Culture cells also diffe rentiate d into chondrocytes after e xposure to dexamethasone and TGF-a 1 ,the cells uniformly spindle shape bacame round and ellipse shape at day2a nd 3,immunohistochemistry revealed expression of II collagens and in dicated a n abundance of glycosaminoglycans within the extr acellular matrix.Conclusion W e successfully induce swine mesenchymal stem cells derived from bone marrow diff erentiate into osteobla st like cells and chondrocytes.