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| Role of Notch Signaling Pathway in Bone Morphogenetic Protein 4-induced Osteogenic Differentiation of Marrow-derived Mesenchymal Stem Cells and Its Mechanism |
| CAO Jun-jie, LI Ai-fang, WEI Ya-lin, LIAN Jing, TANG Min |
| Key Laboratory of Laboratory Medical Diagnostics of Ministry of Education, Chongqing Medical University, Chongqing 400016, China |
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Abstract Objective:To study the role of Notch signaling pathway in bone morphogenetic protein 4 (BMP4) induced osteogenic differentiation of mesenchymal stem cells and its mechanism. Method:(1) After treatment MEFs cells with DAPT or AddnNotch1 and BMP4-CM, the early osteogenic marker alkaline phosphatase (ALP) activity was detected by quantitative assay and staining assay; (2) Later osteogenic marker calcium deposition was determined by Alizarin Red S staining; (3) The expression of essential osteogenic factors (ALP, Runx2, Col1a1) induced by BMP4 were detected by RT-PCR; (4) The protein expression of p-Smad1/5/8 was detected by Immunohistochemistry (ICC); (5) The cell proliferation,cycle were determined with crystal violet staining and flow cytometry. Results:(1) BMP4-induced early osteogenic marker ALP activity was significantly inhibited by DAPT in a dose-dependent manner; (2) After transfecting AdDLL1 and treating MEFs cell with BMP4-CM, the early osteogenic differentiation marker ALP activity and ALP staining and later osteogenic differentiation marker calcium deposition were enhanced. However, after treating MEFs cell with DAPT or AddnNotch1 and BMP4-conditioned medium (BMP4-CM), the early osteogenic differentiation marker ALP activity and ALP staining were inhibited; (3) The expression of essential osteogenic factors (ALP, Runx2, Col1a1) induced by BMP4 were enhanced by DLL1, but were inhibited by DAPT; (4) The protein expression of p-Smad1/5/8 in cell nucleus were enhanced by DLL1, but were inhibited by DAPT; (5) The cell proliferation were promoted by DLL1, but were inhibited by DAPT. Conclusion:Notch signaling promotes BMP4-induced osteogenic differentiation of MSCs by BMP/Smads pathway. At the same time, Notch also enhances cell proliferation during BMP4-induced osteogenic differentiation.
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Received: 12 August 2016
Published: 25 April 2017
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