Involvement of ROS, ASK1, and JNK in denbinobin-induced c-Jun phosphorylation in A549 cells. Cells were pretreated with 1 mM NAC, 100 μM GSH (A), or 10 μM SP600125 (B), or transiently transfected with pcDNA, ASK1DN, JNK1DN, or JNK2DN for 6 h (C). After treatment, cells were treated with vehicle or 20 μM denbinobin for 30 min. Cells were then harvested for immunoblotting to assess the level of c-Jun phosphorylation as described in "Materials and methods". Equal loading in each lane is reflected by approximately similar intensities of the c-Jun bands. Compiled results are shown in the lower panel. Each column represents the mean ± S.E.M. of at least three independent experiments. * p < 0.05, compared to the denbinobin group. * p < 0.05, compared to the control group (A and B) or the pcDNA (mock) group (C) in the presence of denbinobin.