Methods: Culture supernatants of P.gingivalis W50 were grown anaerobically for 6 days. Ascending bacterial protein concentrations of P.gingivalis culture supernatant up to 5µg/ml were used to challenge W20-17 cell cultures for up to 48h. The levels of RANKL, OPG and PGE2 were analysed by ELISA. The involvement of PGE2 in RANKL and OPG regulation was investigated by treatment of the cells with indomethacin(10-5M). To investigate the involvement of P.gingivalis proteinaceous components, cysteine proteinases, or lipopolysaccharide (LPS), the P.gingivalis culture supernatants were heat-inactivated, TLCK-treated (1mM), or passed through a polymyxin-B column, respectively.
Results: P.gingivalis supernatant increased RANKL protein production and PGE2 secretion, whereas OPG production was decreased, in a concentration dependent manner. Controls vs 5µg/ml P.gingivalis at 48 h; RANKL: 22±14 vs 273±105 ng/ml; PGE2: 776±773 vs 6446±1540 ng/ml; OPG: 97747±35989 vs 17100±4357 ng/ml. Pre-treatment of the cells with indomethacin decreased P.gingivalis-stimulated PGE2 secretion by 97 % and RANKL production by 98%, but did not affect OPG production. Heat- and TLCK-treatment of P.gingivalis had no effects on RANKL and PGE2 production, but polymyxin-B treatment reduced both by 98% and 78% respectively.
Conclusion: Porphyromonas gingivalis up-regulates RANKL and PGE2, but down-regulates OPG in bone marrow stromal cells. The bacterial effects on RANKL induction appear to be mediated through PGE2 stimulation and these events are triggered by a non-proteolytic and non-proteinaceous component of P.gingivalis supernatants, most probably LPS. This activity may contribute to the bone loss characteristic of destructive periodontal disease.