Effects of metformin on osteoclasts differentiation in vitro
10.3760/cma.j.issn.0253-2352.2011.05.021
- VernacularTitle:二甲双胍对破骨细胞体外分化的影响
- Author:
Ming LU
;
Song XU
;
Qiguang MAI
;
Rongping ZHOU
;
Zhongmin ZHANG
;
Liang WANG
;
Minjun HUANG
;
Xiaokai WANG
;
Dadi JIN
- Publication Type:Journal Article
- Keywords:
Osteoclast;
Mefformin;
Differentiation
- From:
Chinese Journal of Orthopaedics
2011;31(5):535-541
- CountryChina
- Language:Chinese
-
Abstract:
Objective To investigate the effects of mefformin on the differentiation of osteoclastas well as relative mechanism.Methods Raw264.7 cells from the murine macrophage cell line was used.Receptor activator of NF-κB ligand (RANKL) was used to stimulate osteoclast differentiation from Raw264.7 cells.Osteoclast differentiation was assessed by tartrate-resistant acid phosphatase (TRAP) and actin fluorescence staining and counting the TRAP-positive cells after exposure to different concentrations of mefformin (0 μmol/L,400 μmol/L,800 μmol/L and 1000 μmol/L) or rapamicin (100 nmol/L) in the presence of 50 ng/ml RANKL for 5 days.Bone-resorbing activity was evaluated by BD BioCoatTM OsteologicTM Bone Cell Culture System.The expression of osteoclast-specific genes like TRAP,capthesin K,calcitonin receptor (CTR) and matrix metalloproteinase (MMP-9) was evaluated by RT-PCR.The expression of tumor necrosis factor-α(TNF-ct) S6K1Thr389,S6 Ser235/236,4E-BP1Thr37/46 and c-Fos protein was evaluated by ELISA kit and Western blot analysis,respectively.Results Mefformin dose-dependently inhibited RANKL-stimulated osteoclasts differentiation in Raw264.7 cell culture,as manifested by decrease of TRAP-positive multinucleated cells and pit erosion area,down-regulation of TRAP,cathepsin K,CTR and MMP-9 mRNA and reduction of TNF-α and c-Fos protein expression.Further study revealed that RANKL activated mTOR complex 1(mTORC1) signaling,while mefformin impaired RANKL-stimulated mTORC1 signaling.Rapamycin,an mTORCl-specific inhibitor and immunosuppressive macrolides could also prevent RANKL-induced osteoclast differentiation and bone resorption in vitro.Conclusion Mefformin inhibits osteoclastogenesis in vitro,which may due to reduction of TNF-α and c-Fos protein expression,and mTORC1 signaling is involved in this process.
