Cyclic Pressure Affects Cytokine Production in Human Gingival Fibroblasts

Mucasal overload areas due to the chronic stress and movement of an ill-fitting denture will often contribute to the alveolar bone resorption. However, involvement of oral mucosa cells in the mechanical stress-induced bone resorption has not been understood. Objectives: To investigate the effect of cyclic pressure on biological responses in human gingival fibroblasts (hGFb). Methods: hGFb isolated form gingival tissues were subjected to cyclic pressure using a custom-designed pressure system. During experiments, hFb placed in the sealed, 37C°, humidified pressure chamber in an atmosphere of 5% CO₂ were exposed to cyclic pressure regimes characterized by pulse pressure (5-50 kPa above atmospheric), frequency (1.67 mHz), and duration of exposure (6 days). Pressurized 5% CO₂ gas was injected into, and expelled from the pressure chamber through computer-control. Cellular cytotoxicity and proliferation were measured by CytoTox-Glo^TM Cytotoxicity Assay (Promega) and WST-1 Assay (Dojin Laboratory). Expressions of mRNAs encoding interleukin (IL)-1β, IL-6, IL-8, osteoprotegrin, and receptor activator for nuclear factor κ B ligand (RANKL) under the cyclic pressure condition were determined by reverse transcription (RT)-PCR analysis. Results: hGFb cultured for 2-6 days responded to cyclic pressure in a magnitude- and time- dependent manner. Increases in pressure magnitude resulted in significant decrease in cellular proliferation. Cellular cytotoxicity remained unchanged even under the maximum cyclic pressure condition (50 kPa, 1.67 mHz) over the course of the culture period. Cyclic pressure of 50 kPa for 6 days induced expression of IL-1β, IL-8 and RANKL, while the expression levels of IL-6 and OPG mRNA remained unchanged. hGFb clearly increased expression of IL-8 mRNA in response to the cyclic pressure within 6 hours, as determined by real-time RT-PCR analysis. Conclusion: These results suggest that a harmless level of cyclic pressure stimulates early induction of IL-8 and late induction of RANKL mRNA expression in hGFb.