Fig. 5.

TNF-α and EGF-induced COX-2 expression requires EGFR tyrosine kinase activity. Cultures of 18Co cells were pretreated for 1 h with the EGFR inhibitor 1 μM AG1478 for 1 h before exposure to 5 ng/ml EGF for 4 h in the presence or absence of an 18-h incubation with 10 ng/ml TNF-α. Cell lysates were analyzed by SDS-PAGE and Western blot using anti-COX-2 antibody and an antibody that detects p/42/44 MAPK phosphorylation. Similar results were obtained in ≥3 independent experiments for each condition. Results are means ± SE and are expressed as percentage of the maximum level of COX-2 expression and as a percentage of the maximum level of p42/44 MAPK phosphorylation, depicted in graphical form at bottom. Equal protein loading was verified using an antibody that detects α-SMA.