The role of Hedgehog signaling pathway in the simvastatin-induced osteogenic differentiation of bone marrow mesenchymal stem cells
10.13220/j.cnki.jipr.2019.11.005
- Author:
Zhi-Bin GUO
1
Author Information
1. Linxi Hospital of Kailuan General Hospital
- Publication Type:Journal Article
- Keywords:
Bone marrow mesenchymal stem cells;
Gli1;
Hedgehog signaling pathway;
Osteogenic differentiation;
Simvastatin
- From:
Journal of International Pharmaceutical Research
2019;46(11):832-839
- CountryChina
- Language:Chinese
-
Abstract:
Objective: To investigate the osteogenic differentiation of bone marrow mesenchymal stem cells (BMSC)induced by simvastatin(SIM)in vitro and then further investigate whether the Hedgehog signaling pathway is involved in the SIM-induced osteogenic differentiation of BMSC using the Hedgehog signaling pathway blocking agent cyclopamine(Cpn). Methods: The rat BMSC was extracted by the whole-bone marrow adherent culture method and cul- tured in the induction medium(control medium,CM)and the induction medium containing SIM or SIM+Cpn. The expression of alkaline phosphatase(ALP)in induced cells was detected by the ALP staining. Immunofluorescence stain- ing was performed to detect the espressions of Gli1 and osteocalcin(OCN)in the induced cells. The Gli1,ALP,colla- gen type(COL)and OCN mRNA expression was detected by real-time quantitative PCR(RT-PCR). Western blot (WB)was used for assessing the expression level of Gli1,Runx2,COLand OCN proteins. The cell calcium nodule for- mation and matrix mineralization ability were detected by alizarin red staining. Results: The ALP expression was signifi- cantly higher in SIM group than in CM group(P<0.05),while the ALP expression was lower in SIM+Cpn group than in the SIM group(P<0.05)but higher than in the CM group. Immunofluorescence assay showed that SIM could promote the expression of Gli1 and OCN in the conditions with or without Cpn. On the 10th and 18th day of intervention,the mRNA expression level of Gli1,ALP,OCN and COLwas significantly higher in the SIM group than in the CM group(P<0.05), and the higher mRNA expression in the SIM group could be completely blocked by Cpn. Further,on the 10th and 18th day of intervention,the expressed Gli1,Runx2,COLand OCN protein level was significantly higher in the SIM group in the CM group(P<0.05),while the level of these expressed proteins in the SIM+Cpn group was significantly lower than in the SIM group(P<0.05). The alizarin red staining showed that SIM group had stronger matrix mineralization ability than the other groups(P<0.05). Conclusion: SIM could up-regulate the expression of Gli1 and the osteogenesis markers ALP,Runx2,COLand OCN to induce osteogenic differentiation of BMSC,and Cpn could not completely block the SIM-induced differentiation. Further study is needed to uncover the underlying mechanism.
