摘要
目的探讨转化生长因子β1（TGF-β1）在骨髓间质细胞分化中的作用及与纤维连接蛋白在诱导分化中表达的量效关系。方法抽取Wister大鼠骨髓于DMEM培养液中单层培养，扩增。用2，3传代细胞按一定比例种植于24孔板内，加入TGF-β1 1，5，10ng／mL，分别于不同时间点取材固定，用于甲苯胺兰组织学染色及抗纤维连接蛋白抗体免疫组化染色。用不同剂量的纤维连接蛋白抗体加入诱导体系中观察细胞的聚集变化。结果传代的骨髓间质细胞在TGF-β1的作用下于第4天产生形态变化，由梭型变为圆形，甲苯胺兰染色呈阳性，TGF-β1为5ng／mL时效果显著；纤维连接蛋白的表达主要以48～72h明显，当加入其抗体时，细胞结合逐渐松散，当剂量为200μL／孔时，不再发生细胞聚集现象。结论TGF-β1在诱导骨髓间质细胞转化中发挥重要作用，其最佳剂量为5ng／mL，同时它可促进纤维连接蛋白的合成分泌，二者呈正相关。本研究结果为细胞外基质蛋白纤维连接蛋白可能参与TGF-β1的信号传导通路提供了有力依据。
Objective To investigate the role of TGF-β1 on chondrogenesis of Mesenchymal stem cells and the relationship with the expression of extracellular matrice Fibronectin during cellular differentiation. Methods The wistar rabbit＇s MSCs were harvested and expanded in monolayer culture in six-pore plates and the 2nd or 3rd passaged cells were induced by different dose of TGF-β1. The coverslip with differentiated cells were fixed at different timepoints and used for toludine blue or inmunohistochemical dying Meanwhile, different doses of antibody to Fibronectin were added to observe aggregating phenomenon. Results The passaged cells induced by TGF-β1 showed morphological change at 4 days from spindle shape to the round . The differentiated cells were positive to toludine blue and the optimal dose was 5 ng/mL. The expression of fibronectin was evident at 48-72 hours . Perterbation of antibody to Fibronectin could influence the cell＇s aggregating, for example, when dose of 200 μL/pore was added , no aggregation was occurred. Conclusion TGF-β1 superfamily could play an important role during chondrogenesis of MSCs and the optimal dose is 5 ng/mL. It can promote Fibronetin secretion, and shows positive-related tendency . Our result maybe prove further that the extracellular matrice Fibronectin should be one of the TGF-β1 transduction pathways.
出处
《江西医学院学报》
2006年第2期31-33,共3页
Acta Academiae Medicinae Jiangxi
