Role of Notch Signaling Pathway in Bone Morphogenetic Protein 4-induced Osteogenic Differentiation of Marrow-derived Mesenchymal Stem Cells and Its Mechanism

Objective: To study the role of Notch signaling pathway in bone morphogentic protein 9(BMP9) induced osteogenic diffierentiaion of marrow-derived mesenchymal stem cells and its mechanism.Method: After treatment C2C12 cells with recombinant adenovirus GFP and BMP9(AdGFP/AdBMP9),the early osteogenic marker Alkaline phosphatase(ALP) activity was detected by staining assay,later osteogenic marker calcium deposition was determined by Alizarin Red S staining.Effects of Notch signaling pathway on ostegenic differentiation was detected by cell-density assay and Notch signaling pathway inhibitor(DAPT).The receptor of BMP9,Smad1/5/8 and some transcription factors were detected in order to study the its mechanism.Results: AdBMP9 is not only increased the activity of ALP on days 3,5,7 matrix mineralization on 7d and 14d in a dose-dependent manner.Notch specific inhibitor DAPT decreased ALP activity and the expression of OCN.Furthermore,DAPT markedly change expression level of ALK2 and ostegenic differentiation marker Runx2 and Col1a-α induced by BMP9 of C3H10T1/2.DAPT did not change total protein level of Smad1/5/8,however,DAPT increased the phosphorlated form of Smad1/5/8.Conclusion: Notch signaling pathway can enhance BMP9-induced osteogenic differenitiation of marrow-derived mesenchymal stem cells possibly by up-regulating the expression of its ligands and receptors through BMP9,Notch signaling pathway feedback up-regulates BMP9 signaling,thus leading to osteogensis.