Figure 4

Involvement of MAPK (p38, MEK1/2, JNK/AP-1) signalling pathways in 25-HC-augmented IL-8 release. (A) Cells were treated with 10 μM 25-HC or vehicle. At various time points, the nuclear fraction of the cell lysates was obtained. Effect of 25-HC on translocation of phosphorylated c-Jun into the nucleus. Translocation of the phosphorylated c-Jun into the nucleus was evaluated by immunoblotting. Each band intensity was assessed by densitometry. Relative intensity was calculated by dividing the phosphorylated c-Jun band intensity by each appropriate lamin A/C band intensity. The data are expressed as mean values ± SEM for three to seven separate experiments with HBEpC from three donors. (B-D) Effect of c-Jun N-terminal kinase (JNK), p38 Mitogen-activated protein kinase (MAPK) or mitogenic extracellular kinase (MEK)1/2 inhibitor on 25-HC-augmented IL-8 release in HBEpC. Cells were treated with 10 μM 25-HC or vehicle in the presence of a JNK inhibitory peptide, L-JNKi1 (B), a p38 MAPK inhibitor, SB203580 (C) or a MEK1/2 inhibitor, U0126 (D). After 24 h, the supernatants were assayed for IL-8 by ELISA. *p < 0.05, **p < 0.01, compared with the values of control at 0 min or the values of vehicle-treated cells. +++p < 0.01, compared with the values of 25-HC-treated control cells. The data are expressed as mean values ± SEM for three to four separate experiments with HBEpC from two donors. pcJun = phosphorylated c-Jun. N.S. = not significant.