颞下颌关节滑膜间充质干细胞成骨潜能的实验研究

Osteogenic Differentiation of Synovial Mesenchymal Stem Cells in vitro

目的研究滑膜间充质干细胞的成骨潜能。方法用2g/L的Ⅰ型胶原酶消化获得滑膜细胞,待细胞汇合后用有限稀释法进行单细胞克隆,筛选出滑膜间充质干细胞(SMSC)。取第3代SMSC进行成骨诱导培养,每天观察细胞形态,并于培养7d后检测ALP活性和骨桥蛋白的表达,RT_PCR检测核心结合因子al(cbfal)mRNA的转录;培养30d后作VonKossa′s染色,检测成骨化程度。结果SMSC在体外培养形成葵花样细胞集落,胞浆突起明显,相互连接成网状;成骨化诱导剂可诱导SMSC定向分化为多形性成骨细胞,细胞ALP染色阳性,骨桥蛋白强阳性,电泳显示有cbfal的特异性条带,矿化区经VonKossa′s染色呈阳性反应。结论经体外纯化的SMSC可定向诱导分化为成骨细胞,符合成骨细胞的生物学特征。

Objective To investigate the potential of synovial mesenchymal stem cells(SMSC) in osteogenic differentiation.Methods SMSC were obtained by limited dilution method and expanded to culture in 25-milliliter flasks. The attached cells were treated with inductive medium containing dexamethasone, glycerophosphate and vitamin C at 3rd passage SMSC. The mineralized nodule was stained by Von Kossa method. The expression of ALP and osteopontin were detected by histochemical, immunohistological staining technique, respectively, while the expression of cbfa1 mRNA by RT-PCR.Results Pure SMSC which were of spindle shape and star shape, uniform in size, could be induced to pleomorphism osteoblast in vitro, which were intensive positive in ALP and osteopontin. The expression of cbfa1 mRNA were also verified by RT-PCR and the polygonal cells formed nodular structure at 4 weeks. All these were coincident with the characters of osteoblast.Conclusion SMSC can be purified and induced into osteoblast in vitro.