Emdogain increase osteoclast formation and RANKL/OPG expression in osteoblasts

The level of RANKL/OPG mRNA expression has been shown to be important in bone formation and bone resorption. Several studies have been attempted to clarify the mechanism of Emdogain (EMD) in bone remodeling. However, the mechanism is still unknown. Objectives: Investigate the effects of EMD on RANKL and OPG mRNA expression in osteoblasts and on osteoclast formation. Methods: Osteoblast like-cells MG-63 were grown in DMEM at the amount of 2x10⁵ cells/dish in 60-mm petri dishes. Three different media conditions were tested: DMEM without FBS, DMEM with 10% FBS, and DMEM with EMD 100 µg/ml. MG-63 cells were cultured in these conditions for 48 hours. Total RNA was isolated and RT-PCR was performed using primers specific for RANKL and OPG. The RANKL/OPG ratio was analyzed using ANOVA. For osteoclast formation study, effects of EMD were evaluated 14 days after co-cultured of MG-63 with PBMCs (peripheral blood mononuclear cells) using TRAP staining. In brief, MG-63 were cultured at the amount of 6x10⁶ cells/well in 24 well-plates and maintained in DMEM media for 24 hours then further co-cultured with 1x10⁶ cells/well of PBMCs. 10^-8M 1 α, 25 dihydroxycholecalciferol and10^-7M dexamethasone were added to DMEM co-culture media. EMD 100 µg/ml was added only to the experimental groups for 14 days. TRAP staining was then performed on day 15. TRAP+ cells with three or more nuclei were considered as osteoclasts. Data analyses were carried out using Student's t-test. Results: RANKL/OPG mRNA expression was 33% (P < 0.05) higher in EMD treated groups and EMD increased osteoclast formation by 36% (P < 0.05) compared to controls. Conclusion: EMD stimulated osteoclast formation in co-culture of MG63 and PBMCs. In addition, EMD acted on MG-63 by increasing RANKL/OPG ratio. (This study was approved and supported by Prince of Songkla University grant number DEN49541).