左、右归丸对大鼠骨髓间充质干细胞成骨诱导中转化生长因子β1及Smad2/3的影响

Effect of Zuoguiwan and Youguiwan on transforming growth factor beta1 and its signal transduction protein Smad2/3 in osteogenic induction of rat bone marrow mesenchymal stem cells

背景:滋补肾阴左归丸、温补肾阳右归丸均可协同诱导剂诱导骨髓间充质干细胞向成骨细胞分化,但其机制尚有待探讨。目的:观察左、右归丸含药血清对体外培养的大鼠骨髓间充质干细胞向成骨细胞分化过程中转化生长因子β1及其信号转导蛋白Smad2/3表达的影响。方法:运用全骨髓贴壁法分离和培养大鼠骨髓间充质干细胞;分别以空白血清对照组,左归丸、右归丸、阳性对照药补佳乐制备的大鼠含药血清加诱导剂(地塞米松、维生素C、β-甘油磷酸钠),以及诱导剂组为对照共5组对骨髓间充质干细胞进行干预。采用Western blot法检测Ⅰ型胶原蛋白表达,用免疫组化法检测转化生长因子和Smad2/3在细胞内的表达。结果与结论:左归丸、右归丸含药血清诱导骨髓间充质干细胞成骨后Ⅰ型胶原蛋白、转化生长因子β1、Smad2/3蛋白表达高于空白组、诱导剂组,差异有显著性意义(P<0.05),且左归丸优于右归丸(P<0.05)。提示左、右归丸含药血清能通过转化生长因子β1调节Smad2/3信号通路促进骨髓间充质干细胞成骨分化,且左归丸组效果更好,表明中医"滋肾阴"法更能有效的促进骨髓间充质干细胞的成骨分化。

BACKGROUND：Bone marrow mesenchymal stem cells can differentiate into osteoblasts under inducing condition that Zouguiwan and Youguiwan coordinate inducers, but the mechanism remains to be discussed. OBJECTIVE：To observe the effects of serum containing Zuoguiwan and Youguiwan on transforming growth factorβ1 and its signal transduction protein Smad2/3 message expression during the osteogenic differentiation of bone marrow mesenchymal stem cells. METHODS：A whole bone marrow adherence method was adopted to isolate and cultivate bone marrow mesenchymal stem cells from rats. The cellcultivation was processed in five groups：bone marrow mesenchymal stem cells were respectively cultured with blank serum, serum containing Zouguiwan, serum containing＆amp;nbsp;Youguiwan, positive serum containing progynova＋inducer （dexamethasone, vitamin C, andβ-glycerophosphate）, and inducer. Western blot was applied to detect the expression of type I col agen. The immunohistochemical assay was utilized to test transforming growth factorβ1 and Smad2/3 expression in the osteoblasts. RESULTS AND CONCLUSION：It was apparently more significant for serum containing Zuoguiwan and Youguiwan on type I col agen, transforming growth factorβ1 and Smad2/3 expression, compared with blank serum group and inducer group （P＆lt;0.05）;moreover, serum containing Zuoguiwan was better than serum containing Youguiwan （P＆lt;0.05）. Both of serum containing Zuoguiwan and Youguiwan are able to promote osteogenic differentiation of bone marrow mesenchymal stem cells. Moreover, Zuoguiwan is much more effective indicating that this method of traditional Chinese medicine about nourishing kidneys can be better to promote osteogenic induction of bone marrow mesenchymal stem cells.