Activation of Receptor Activator of Nuclear Factor-κB Ligand and Matrix Metalloproteinase Production in Periodontal Fibroblasts by Endothelin Signaling

Background: Periodontitis is a group of inflammatory diseases affecting the tissues supporting the teeth that will progressively cause the loss of alveolar bone and periodontal ligaments and eventually the dentition. Activation of osteoclast activity by receptor activator of nuclear factor-kB ligand (RANKL) and released enzymes such as matrix metalloproteinases (MMPs) are among the factors involved in the breakdown of the periodontium. However, the mechanisms regulating their production in periodontitis are poorly understood. Endothelin signaling via the activation of the endothelin-A receptor (EDNRA) by endothelin-1 may play a role in the disease because the expression of the receptor and ligand is elevated in the periodontal tissues of patients with periodontitis. Methods: Cultured primary human periodontal fibroblasts were treated with 20 and 100 nM endothelin-1 for 6 and 24 hours and then collected to assess MMP and RANKL production by immunoblotting. Inhibitors were used to identify the molecular pathways activated by EDNRA in these cells. Results: Endothelin-1 stimulated the production of MMP1, MMP8, and RANKL in a dose- and time-dependent manner; blocking EDNRA function with the antagonist TBC3214 inhibited the response, although EDNRA activation had no effects on osteoprotegerin production. These mechanistic studies indicate that EDNRA activates phospholipase C, which then 1) increases the MMP1 protein levels through activation of the extracellular signal-regulated kinase mitogen-activated protein kinase-dependent pathway and 2) upregulates RANKL by a different pathway. Conclusion: These results suggest that EDNRA may function in the breakdown of the periodontal tissues associated with periodontitis by promoting the protein expression of MMPs and RANKL via the phospholipase C pathway.