Fig. 4. Inhibition of Rankl induced osteoclasts differentiation by Nampt enzyme inhibitors MC4, FK866 with or without MTX in RAW 264.7 and BMM.

a RAW cells were pretreated with 2 nM MC4 or FK866 or 50 nM MTX for 3 h and then stimulated with Rankl (100 ng/ml) for 5 days. TRAP staining was performed using a TRACP and ALP double staining kit. b The frequency of TRAP-positive cells expressed as mean of triplicate determinations ± SD. c BMM were isolated from wild-type mice pretreated with 2 nM MC4 or FK866 or 50 nM MTX for 3 h and then stimulated with Rankl (100 ng/ml) for 5 days. TRAP staining was performed using a TRACP and ALP double staining kit. d The frequency of TRAP-positive cells expressed as mean of triplicate determinations ± SD. *P < 0.05; **P < 0.005; ***P < 0.001