Fig. 3

CYP1A1-derived 12(S)-HETE production intensifies JNK/AP-1 activation. a CYP1A1/RAW and NC/RAW, as well as PMs transfected with CYP1A1 siRNA or scramble siRNA were treated with vehicle or LPS (10 μg/ml) for 12 h, and supernatants were then assayed for 12(S)-HETE concentration and CYP1A1 hydroxylase activity (AHH; presented as %) by ELISA and a standard AHH activity protocol, respectively. b, c RAW cells were treated with vehicle or LPS and 12(S)-HETE (500 nM) for 2 h (b) and 12 h (c). b The cell lysates were used for western blotting analysis of JNK/AP-1 phosphorylation levels. c TNF-α and IL-6 protein levels were assessed by ELISA. d-f CYP1A1/RAW and NC/RAW were administrated 12(S)-HETE antibody (3 μg/ml) for 12 h and then LPS for 2 h (western blotting and EMSA) and 12 h (ELISA). d Treated cells were lysed for measuring JNK/AP-1 phosphorylation level by western blotting. e Nuclear-extract proteins of treated cells were incubated with AP-1-binding site probe and binding activity measured using EMSA. f TNF-α and IL-6 protein levels were assessed by ELISA. Data are shown as mean ± SEM of three independent experiments. Results were compared by one-way ANOVA. *p < 0.05