骨形态发生蛋白9诱导间充质干细胞骨向分化与Wnt11的关系

Relationship between Wnt11 and BMP9-induced osteogenic differentiation in mesenchymal stem cells

目的 探讨骨形态发生蛋白9(BMP9)诱导干细胞骨向分化与Wnt11的关系及可能的分子机制。方法通过qPCR、组织化学染色及Western blot等方法,检测成骨分化相关标志物,以及BMP/Smad和p38 MAPK信号的变化;利用荧光素酶报告质粒,检测BMP/Smad信号的活性改变。结果 BMP9增加C3H10T1/2细胞中碱性磷酸酶(ALP)活性、钙盐沉积、骨桥素(OPN)和Wnt11表达。Wnt11在C3H10T1/2、MEFs、MC3T3-E1和C2C12细胞中均有表达。在C3H10T1/2细胞中,Wnt11增强BMP9促进ALP活性、钙盐沉积、Runx-2和OPN表达的作用,以及BMP9对BMP/Smad报告质粒转录活性和Smad1/5/8磷酸化的促进作用;Wnt11还增强BMP9诱导p38 MAPK磷酸化的作用;抑制p38 MAPK则减弱BMP9诱导ALP活性、钙盐沉积及OPN表达的作用,但该效应能被Wnt11部分逆转。结论 BMP9在MSCs中能上调Wnt11表达。Wnt11能促进BMP9的骨向分化诱导作用,该作用可能与其增加BMP/Smad和p38 MAPK信号的活性有关。

Aim To investigate the relationship between Wnt11 and the BMP9-induced osteogenic differentiation,and the possible molecular mechanism underlying this process as well.Methods qPCR,histochemical staining and Western blot were used to detect the changes of osteogenic differentiation markers and activities of BMP/Smad and p38 MAPK signal.Luciferase reporter assay was used to detect the activity of BMP/Smad signal.Results BMP9 increased the alkaline phosphatase(ALP) activity,mineralization,expression of osteopontin (OPN) and Wnt11 in C3H10T1/2 cells.Wnt11 was detectable in C3H10T1/2,MEFs,MC3T3-E1 and C2C12 cells.In C3H10T1/2,Wnt11 promoted the effect of BMP9 to increase ALP activity,mineralization,and the expression of Runx-2 and OPN.Wnt11 enhanced the effect of BMP9 on increasing BMP/Smad reporter plasmid transcriptional activity and phosphorylation of Smadl/5/8.Wnt11 also increased the BMP9-induced phosphorylation of p38 MAPK.Inhibition of p38 MAPK attenuated the BMP9-induced ALP activity,mineralization and expression of OPN,but these effects were partially reversed by Wnt11.Conclusions BMP9 can up-regulate Wnt11 in MSCs.Wnt11 can promote the BMP9-induced osteogenic differentiation,which may be mediated by increasing the activity of BMP/Smad and p38 MAPK signaling.