Effect Of Metformin(MF) On The Proliferation And Collagen ⅠandLRP5mRNA Expression In Cultured Human Osteoblasts

ObjectiveTo observe the effect of the different concentrations of metformin on proliferation and the mRNA expression of Collagen I and LRP5in vitro human osteoblasts,further exploreing the possible mechanisms of effect of metformin in human osteoblasts,provide valuable references for diabetic osteoporosis.Methods1. The human trabecular bone specimen of femoral neck from adult patients were obtained, by enzymatic digestion in vitro cultured human osteoblasts,and observe their morphology and functional identified.2.The human osteoblasts were cultured in vitro, different concentrations of metformin (0,25,50,100,200μmol/L) stimulate human osteoblasts after72hours,cell proliferatio was assessed with CCK-8colorimetric method,and then determine their ODmv.3.Different concentrations of metformin (0,25,50,100,200μ mol/L) stimulate human osteoblasts after72hours,The expression of Collagen Ⅰand LRP5mRNA were examined with RT—PCR.4.All statisticalanalysis is performed with Windows SPSS16.0,data are presented as (?)±s. The differents between the groups are detemined by One-way ANOVE.The differences between random two groups are determined by LSD-T.Differences are considered significant at a value of P〈0.05. Results1.Most of cultured human cancellous bone osteoblasts, observed under a microscope were spindle fusiform or irregular,cell surface has multiple short spike,surrounding cells to protuberant connected. Osteocalcin immunohistochemistry staining cytoplasm brownish yellow.And the calcium alizarin red staining was positive.2.Different concentrations of metformin (0,25,50,100,200μ mol/L) stimulate human osteoblasts after72hours, the OD value are increased, in the concentration dependence manner,it was most remarkable at200μ mol/L,the experimental groups and compared with the control groups were considerd significant at a value of P<0.05.3.The expression of Collagen I and LRP5mRNA were examined with RT—PCR,the result shows that different concentrations of metformin (0,25,50,100,200μ mol/L) can promote Collagen I and LRP5gene mRNA expression,especially at200μ mol/L,in dose-dependent manner. The experimental groups and compared with the control groups were considerd significant at a value of P<0.05.Conclusion1.Metformin can promote proliferation of the cultured human osteoblasts.2.Metformin can promotes expression of Collagen I and LRP5gene mRNA.3. A certain concentration range of metformin probably through the Wnt/LRP5/β-catenin signal path regulate the expression of Collagen Ⅰand LRP5mRNA promote proliferation of the cultured human osteoblasts,thus promoting bone formation.