The regulation of osteopontin in stress-induced human periodontal ligament cells

Abstract

Mechanical stress such as orthodontic forces can produce mechanical damage and inflammatory reaction in the periodontium. Osteopontin (OPN) is a multifunctional cytokine, function as a chemotactic factor and enhance the spreading and the attachment of osteoclasts to the bone surface. This study aimed to examine the influence of mechanical stress on the expression and regulation of OPN as well as the signaling pathway involved in human periodontal ligament (HPDL) cells. The in vitro mechanical stress was generated by continuous compressive force to HPDL cell culture, the expression of OPN mRNA and protein was examined by reverse transcription-polymerase chain reaction and Western analysis, respectively. The application of inhibitors was used to investigate the involved mechanism. The results of this study demonstrated the increased of OPN in a force dependent manner in both mRNA and protein levels. Interestingly, the increased of receptor activator of nuclear factor-kappa B ligand (RANKL) was also observed. Application of indomethacin could abolish the induction of RANKL but not that of OPN, suggesting the cyclooxygenase-independent mechanism for stress-induced OPN expression. The upregulation of OPN was diminished by Rho kinase inhibitor suggesting of the involvement of Rho kinase pathway involved in this mechanism. The condition media (CM) collected from stress-induced HPDL cell also increased OPN expression similar to the direct stress application. Application of suramin and NF449, a purinergic P2 receptor inhibitor, abolished both stress-induced and CM-induced OPN expression, indicating the involvement of nucleotides and P2 receptor in the inductive mechanism. In addition, the increase of adenosine 5'-triphosphate (ATP), the universal nucleotide ligand of P2 receptor family, was also found in CM after stress stimulation and the application of exogenous ATP also induced OPN expression via Rho kinase pathway. In the conclusion, mechanical stress affects OPN expression in HPDL cells by stimulating the releasd of ATP into the medium, subsequently, ATP acts through P2 receptor located at cell surface, and then activated Rho kinase pathway. The increase of OPN participates in clinical alveolar bone resorption and remodeling by inducing the migration and the attachment of osteoclasts.