JNK promotes DCLF/IFN-mediated phosphorylation of STAT-1 at Ser 727 via activation of ERK. HepG2 cells were treated with VEH (0.1% DMSO) or SP600125 (20 μM) and simultaneously treated with sterile water (Control) or DCLF (250 µM) alone or in combination with TNF and/or IFN. Whole cell lysates were collected 18 h after treatment. (A) pSTAT-1 (Tyr 701), pSTAT-1 (Ser 727), α-tubulin and (B) pERK and α-tubulin levels were detected via western analysis. a, significantly different from corresponding bar in VEH group. b, significantly different from corresponding bar in TNF group. c, significantly different from Control within a cytokine group. d, significantly different from DCLF within a cytokine group. e, significantly different from SP600125 within a cytokine group. Western analysis of proteins from cells treated with and without SP600125 was performed simultaneously. Data are represented as mean ± SEM of at least 3 experiments. Abbreviations: VEH, vehicle; DCLF, diclofenac; pSTAT-1, phosphorylated signal transducer and activator of transcription-1; Tyr, tyrosine; Ser, serine; pERK, phosphorylated extracellular signal-regulated kinase.