Figure 2. The effect of PCNA knockdown on RANKL-induced differentiation of osteoclast. (A) PCNA was successfully knocked down using its siRNA. RAW264.7 cells were transfected with siRNA targeting PCNA for 48 hours. Then, the cells were treated with RANKL (100 ng/mL) for 3 days and harvested for western blot assay using the indicated antibodies. (B) The effect of knockdown of PCNA on osteoclast differentiation. RAW264.7 cells were transfected with PCNA siRNA for 48 hours, then the cells were treated with RANKL for 3 days and stained with TRAP. (C) The number of TRAP-positive cells stained in (B). n=5, **: p <0.01, ***: p<0.001. Then, the expression of osteoclast differentiation marker NFAFc1 was detected in control and RANKL-induced RAW264.7 cells, with or without PCNA knockdown (D, E). *: p<0.05, ***: p<0.001.