Fig. 2

CYP1A1 is involved in LPS-induced macrophages activation. a, d CYP1A1/RAW and NC/RAW were treated with LPS (10 μg/ml). TNF-α and IL-6 mRNA levels were detected by qRT-PCR after 4 h LPS stimulation. TNF-α and IL-6 protein levels were measured by ELISA after 12 h LPS stimulation. JNK/AP-1 (c-fos and c-jun) activities were assessed by western blotting after 2 h LPS stimulation. b, d PMs transfected with CYP1A1 siRNA or scramble siRNA were stimulated with LPS. TNF-α and IL-6 mRNA (LPS 4 h) and protein (LPS 12 h) levels and JNK/AP-1 activities (LPS 2 h) were determined respectively. c CYP1A1 siRNA or scramble siRNA were transfected into WT and AhR^−/− PMs respectively. Transfected cells were stimulated with LPS for 12 h. Supernatants were collected for analysis of TNF-α and IL-6 protein release using ELISA. e JNK and AP-1 protein levels were detected in CYP1A1/RAW and NC/RAW in which JNK and AP-1 were knocked out. f, g JNK-knockout CYP1A1/RAW and NC/RAW were treated with vehicle or LPS for 2 h. f The cells were lysed for western blotting analysis of phosphorylated c-fos and c-jun. g The nuclear extract proteins of treated cells were incubated with AP-1-binding site probe and binding activity measured by EMSA. h JNK-knockout CYP1A1/RAW and NC/RAW, as well as AP-1 knockout-CYP1A1/RAW and NC/RAW were stimulated with LPS for 12 h and supernatants were collected for analysis of TNF-α and IL-6 protein release using ELISA. Data are mean ± SEM of three independent experiments. Results were compared by one-way ANOVA. *p < 0.05