The Effects Of1,25-dihydroxyvitamin D3on The Adipogenesis And Osteogenesis In Human Bone Mesenchymal Stem Cells

Objective1)To investigate the effects of1,25-dihydroxyvitamin D3on PPAR-γ in hBMSCs adipogenic differentiation.2)To investigate the effects of1,25-dihydroxyvitamin D3on Hedgehog/Glil、 Wnt/β-catenin、TGF-β/BMP2and Notchl in hBMSCs osteoblast differentiation.3)To investigate the single drug1,25-dihydroxyvitamin D3enhancing hBMSCs to the calcium nodus and the effects of1,25-dihydroxyvitamin D3on the serum bone metabolic parameters.Method1.Human bone mesenchymal stem cells were cultured in adipogenic medium alone or in combination with different concentrations of1,25-dihy droxyvitamin D3.The expression of PPARymRNA were measured by quantitative PCR, and the expression of PPARy protein were measured by Western blot five days later.Comparisons between groups were tested by One-Way ANOVA analysis and t test, or nonparametric tests.2.Human bone mesenchymal stem cells were cultured in osteogenic medium alone or in combination with different concentrations of1,25-dihydroxyvitamin D3.The expressions of Glil、beta-catenin、BMP2and Notch1mRNA were measured by quantitative PCR and the expression of Gli1、beta-catenin、BMP and Notchl protein were measured by Western blot seven days later.Comparisons between groups were tested by One-Way ANOVA analysis and t test,or nonparametric tests.3.The effects of vitamin D3on calcium nodules and serum bone metabolism:1)Human bone mesenchymal stem cells were cultured in common medium or in combination with different concentrations of1,25-dihydroxy vitamin D3,final calcium nodules was identified by Alizarin red stain21days later.2)Patients with underlying risk factors for osteoporosis were selected for study.All patients had oral vitamin D3treatment, ranging from2-6months time,then the changes in the serum25(OH)D3and bone metabolism before and after treatment were tested.Paired samples non-parametric test was used to compare each index before and after medication.Result1.The effects of1,25-dihydroxy vitamin D3on the adipogenesis of hBMSC:1)The expression of PPARy-mRNA was significantly reduced in the10-8mol/L VitaminD3group compared with the adipogenic induction group(P value was0.042); the expression of PPARy protein was significantly reduced in the10-8mol/L VitaminD3group compared with the adipogenic induction group(P value was0.032).2.The effects of1,25-dihydroxyvitamin D3on the osteogenesis of hBMSCs:1)The expression of Glil-mRNA and Notchl-mRNA was significantly increased in the10-5mol/L and10"7mol/L VitaminD3group compared with the osteogenic group(P value was respectively0.018,0.028and0.006、0.016);The expression of β-catenin-mRNA and BMP2mRNA was significantly increased in the 10-5mol/L VitaminD3group compared with the osteogenic group(P value was0.035and0.009).2)The expression of Glil protein was significantly increased in the10-5mol/L VitaminD3group compared with the osteogenic group(P value was0.009);The expression of P-catenin protein BMP2protein、Notchl protein was significantly increased in the10-5mol/L and10-7mol/L VitaminD3group compared with the osteogenic group(P value was0.014and0.03/0.003and0.015/.007and0.043).3.The effects of vitamin D3on calcium nodules and serum bone metabolism:1)Alizarin red staining--after the treatment for21days, the calcium nodules can be prominently observed in the VitD3used group but not in the common medium group.2)The levels of25(OH)D3was significantly increased after vitamin D3treatment,and β-CT was significantly decreased after vitamin D3treatment.Conclusion1)Effective concentrations of1,25-dihydroxyvitamin D3can inhibit the expression of PPARy which in turn lead to the suppression of hBMSCs to the adipocytes.2)Effective concentrations of1,25-dihydroxyvitamin D3can enhance the expression of Hedgehog/Gli1、Wnt/p-catenin、TGF-β/BMP and Notch1which in turn lead to the increase of hBMSC to the osteoblast.3)Use of single drug1,25-dihydroxyvitamin D3can enhance hBMSCs to the calcium nodus,and use of vitamin D3can reduce β-CT thus reducing bone resorption.