Thrombin induces osteoprotegerin synthesis via phosphatidylinositol 3′-kinase/mammalian target of rapamycin pathway in human periodontal ligament cells

Background and Objective: Thrombin influences the biological behavior of periodontal ligament cells and plays multiple roles in the early stages of bone healing. Osteoprotegerin (OPG) is one of the key molecules that regulate bone homeostasis and prevent osteoclastogenesis. The purpose of this study was to evaluate the biological effects of thrombin on OPG synthesis in human periodontal ligament (HPDL) cells in vitro. Material and Methods: Cells were treated with various concentrations (0.001, 0.01 and 0.1 U/mL) of thrombin. The mRNA expression and protein synthesis of OPG, as well as of receptor activator of nuclear factor kappa B ligand (RANKL), were determined by reverse transcriptase-polymerase chain reaction (RT-PCR) and Western blot analysis, respectively. The influence of thrombin on OPG synthesis and its signaling pathway were investigated using inhibitors. Results: Thrombin profoundly induces protein synthesis of OPG at 0.1 U/mL. The inductive effect was inhibited by cycloheximide, but not by indomethacin. The phosphatidylinositol 3'-kinase (PI3K) inhibitor, LY294002, and the mammalian target of rapamycin (mTOR) inhibitor, rapamycin, exerted an inhibitory effect on the thrombin-induced OPG synthesis. In addition, the effect was inhibited by protease-activated receptor (PAR)-1 antagonist. Activation of phospho-Akt (p-Akt) was observed and the effect was abolished by LY294002. Conclusion: Thrombin induces OPG synthesis in HPDL cells post-transcriptionally, possibly through PAR-1. The regulation was through the PI3K/Akt and mTOR pathway. This finding suggests that thrombin may play a significant role in alveolar bone repair and homeostasis of periodontal tissue, partly through the OPG/RANKL system.