Fig. 1. Inhibition of -secretase increases the positive effect of IL-1 on the biosynthesis pathway of lipid mediators. Serum-starved cells were treated daily for 6, 24 or 48 hours with IL-1 (10 ng/ml) or vehicle (control) and/or with DAPT (0.5 µM). (A) PLA₂ secretion was expressed as a percentage of that of control cells (in pmol/min/µg RNA: 6.7±0.1 at 24 hours; 4.7±0.1 at 48 hours). (B) 15 µg of total proteins were separated by electrophoresis (6.5% SDS PAGE). COX-2 and -actin were immunodetected with appropriate antibodies. The autoradiogram is representative of three independent experiments. The histogram represents values obtained from scanning COX-2 bands normalized to that of -actin and results are expressed as a percentage of those from IL-1-treated cells. (C,D) PLA₂-IIA (C) and COX-2 (D) transcripts were assayed by RT-PCR. Results are expressed as a percentage of the PLA₂-IIA or COX-2 mRNA level of control cells and represent the mean ± s.e. of three independent experiments. (E) PGE₂ secretion was expressed as a percentage of the basal PGE₂ secretion of control cells (in pg/ml: 16.7±2.1 at 24 hours; 19.1±0.5 at 48 hours). For PLA₂ and PGE₂ assays, the data represent the mean ± s.e.m. of five or six independent experiments. C, control; IL, IL-1; D, DAPT; D+IL, DAPT plus IL-1 versus IL-1; ns, non-significant; *, P<0.05; **, P<0.01; ***, P<0.001.