To identify whether the mouse papilla-derived MDPC-23 cells could spontaneously differentiate into odontoclast-like cells and function as odontoclasts.

MDPC-23 cells and RAW 264.7 cells stimulated with RANKL were cultured for 6 and 4 days respectively, then the cells were stained for TRAP. The TRAP-positive multinuclear cells induced by MDPC-23 cells were observed and compared with osteoclasts by RAW 264.7 cells. The resorption pits and morphology of MDPC-23 cells on the dentin discs were further determined by metallurgical microscope and scanning electron microscope (SEM). F-actin rings stained with phalloidin were photographed using a confocal microscope. Western blot analysis or immunostaining was used to investigate the protein expression level of odontoclast-specific markers TRAP and Cathepsin K as well as the key protein in odontoclastgenesis—RANKL, RANK and TRAF6. The secretion of RANKL was measured by ELISA in the supernatant harvested from MDPC-23 cells. Two independent samples t test was used to statistically analyze the Cathepsin K and TRAP expression. Statistical analysis of the secretion of RANKL on day 0, 2, 4, 6 was performed using one-way ANOVA.

MDPC-23 cells could spontaneously differentiate into a small amount of TRAP-positive multinuclear cells on day 6, which were morphologically different from osteoclasts induced in RAW 264.7 by RANKL. The resorption pits by these multinuclear cells on the dentin discs were quite shallow. Immunostaining results showed the ringlike structure of F-actin in odontoclast-like cells. Western blot, immunofluorescent staining and ELISA revealed that MDPC-23 cells express RANK and TRAF6 protein and secrete RANKL. The differnce among the level of secreted RANKL from 0 to 6 days was statistically significant (F = 5.373, P = 0.026) . In addition, RANKL release levels were upregulated on day 2 when compared with day 0 (P = 0.007) . Protein expression of TRAP and Cathepsin K were upregulated in MDPC-23 cells on day 6 (t_TRAP = 5.033, P_TRAP = 0.0024; t_{Cathepsin K} = 12.95, P_{Cathepsin K} = 0.0002) .

Papilla-derived MDPC-23 cells could spontaneously differentiate into odontoclast-like cells, but with weak resorption capacity. RANKL, RANK and TRAF6 may play an important role in the regulation of odontoclastgenesis.