As a crucial virulence factor of Porphyromonas gingivalis, gingipains play an 
important role in periodontal destruction. This study aimed to investigate the 
effect of gingipains on osteoclastogenesis. We used RAW264.7 cells as osteoclast 
precursors in our study. In experimental groups, cells were treated with 
gingipains and/or receptor activator of nuclear factor-κB ligand (RANKL). 
Tartrate-resistant acid phosphatase (TRAP) activity staining assay showed 
osteoclast precursors and RANKL-induced mature osteoclasts were increased in a 
gingipains dose-dependent manner. Real-time reverse transcription polymerase 
chain reaction analysis demonstrated that gingipains upregulated osteoclastic 
genes including the protease cathepsin K (Ctsk), matrix metalloprotein 9 (Mmp9), 
nuclear factor of activated T cells 1 (Nfatc1) and acid phosphatase 5, tartrate 
resistant (Acp5) in a time-dependent manner. Western blotting assays presented 
upregulated expressions of TNF receptor-activating factor 6 (TRAF6) and integrin 
β3 induced by gingipains and RANKL compared to RANKL alone. Enhanced 
integrin-related signaling was also demonstrated by elevated phosphorylations of 
FAK and paxillin compared to control. Moreover, the pit resorption assays showed 
that gingipains augmented bone resorptive function of osteoclasts induced by 
RANKL. When we used Cilengitide to block integrin αvβ3, gingipains reversed the 
reduction of formation and resorptive function in RANKL-induced osteoclasts, as 
they enhanced integrin αvβ3 levels more than RANKL treatment alone. In 
conclusion, our data suggest that gingipains augmented the differentiation and 
function of mature osteoclasts induced by RANKL through the increase in integrin 
αvβ3.