Enhancement of Osteogenic Differentiation by Combination Treatment with 5-azacytidine and Thyroid-Stimulating Hormone in Human Osteoblast Cells

ABSTRACT

BACKGROUND AND OBJECTIVES: The role of thyroid-stimulating hormone (TSH) signaling on osteoblastic differentiation is still undetermined. The aim of this study was to investigate the effects of 5-aza-2′-deoxycytidine (5-azacytidine) on TSH-mediated regulations of osteoblasts. MATERIALS AND METHODS: MG63, a human osteoblastic cell-line, was treated with 5-azacytidine before inducing osteogenic differentiation using osteogenic medium (OM) containing L-ascorbic acid and β-glyceophosphate. Bovine TSH or monoclonal TSH receptor stimulating antibody (TSAb) was treated. Quantitative real-time PCR analyses or measurement of alkaline phosphatase activities were performed for evaluating osteoblastic differentiation. RESULTS: Studies for osteogenic-related genes or alkaline phosphatase activity demonstrated that treatment of TSH or TSAb alone had no effects on osteoblastic differentiation in MG63 cells. However, treatment of 5-azacytidine, per se, significantly increased osteoblastic differentiation and combination treatment of 5-azacytidine and TSH or TSAb in the condition of OM showed further significant increase of osteoblastic differentiation. CONCLUSION: Stimulating TSH signaling has little effects on osteoblastic differentiation in vitro. However, in the condition of epigenetic modification using inhibitor of DNA methylation, TSH signaling positively affects osteoblastic differentiation in human osteoblasts.