转化生长因子-β2对骨髓间充质干细胞体外向成骨细胞分化的影响

Influence of transforming growth factor-β2 on differentiation of mesenchymal stem cells into osteoblasts

目的 观察转化生长因子（TGF）-β2在体外诱导骨髓间充质干细胞（BMSCs）向成骨细胞分化能力.方法 取第3代BMSCs接种于3.5×103个/孔密度培养板,分为3组：A、B、C组,A组不做任何处理,B、C组培养基中分别添加5 μg/L TGF-β1、5μg/L TGF-β2,每孔200μl,每组4孔.7d对3个实验组行碱性磷酸酶（ALP）活性检测和Western blot检测Collagen Ⅰ表达,第3代BMSCs以5×107/L细胞浓度接种于6孔培养板,每组1板,分组添加方式同前,21d细胞爬片行Von kossa染色.结果 A、B、C组第7天的ALP活力值分别为1.15±0.16、4.26±0.39、4.34±0.14,各组间比较差异有统计学意义（P＜0.05）,ALP的染色面积及深度、Von kossa染色钙化结节C组强于B组和A组,B组强于A组;Collagen Ⅰ表达C组高于B组和A组,B组高于A组（P＜0.05）.结论 在一定条件下,TGF-β2体外诱导BMSCs成骨能力较转化生长因子-β1强.

Objective To investigate the ability of transforming growth factor （TGF）-β2 to differentiate the bone marrow mesenchymal stem cells （BMSCs） into bone cells in vitro.Methods BMSCs of 3rd generation were seeded in 3.5 × 103/pore density culture plate and divided into three groups：A,B and C.Group A received no treatment.Groups B and C received the culture medium containing 5 μg/L TGF-β1 and 5 μg/L TGF-β2 respectively,200 μL per well of each of the four holes.On 7th day,alkaline phosphatase （ALP） activity assay and Western blotting method were applied to detect the expression of collagen Ⅰ.BMSCs of 3rd generation were seeded in six well plates at a density of 5 × 107/L,grouping and treatment principle were performed as described before,and the calcium salts were observed by Von Kassa at the 21st day.Results The ALP activity values in groups A,B and C on 7th day were 1.15 ± 0.16,4.26 ± 0.39 and 4.34 ± 0.14 respectively （P 〈 0.05）.The stained area and depth of ALP and Von kossa staining for calcified nodules were increased in group C as compared with groups A and B,and those in group B were increased as compared with group A.Collagen Ⅰ expression in group C was highest among the groups and lowest in group A.Conclusion Under above mentioned conditions,TGF-β2 induced BMSCs osteogenic capacity significantly stronger than TGF-β1 and the control.