Cyclooxygenase-2 and synthesis of PGE(2) in human bronchial smooth-muscle cells

The purpose of this study was to determine the mechanism of enhanced prostaglandin synthesis in cultured human bronchial smooth-muscle cells challenged with interleukin-1 beta (IL-1 beta). Cells were incubated with IL-1 beta (10 to 50 U/ml) for 0 to 24 h. Prostaglandin E(2) (PGE(2)) production was evaluated through the conversion of exogenous (C-14)-arachidonic acid and specific enzyme immunoassay of endogenous products. IL-1 beta enhanced PGE(2) formation in a concentration- and time-dependent manner, reaching its peak at 6 to 8 h and fading at 18 to 24 h. Immunoblot analysis showed that the inducible cyclooxygenase enzyme (COX-2) was expressed only in IL-1 beta treated cells, whereas the constitutive isoform of cyclooxygenase (COX-1) remained unaltered. COX-2 expression and PGE(2) formation were inhibited by dexamethasone (2 mu M), cycloheximide (10 mu M), and IL-1-receptor antagonist (IL-1ra) (250 ng/ml), independently. PGE(2) synthesis was significantly reduced by compound SC-58125, a specific COX-2 inhibitor. The close parallelism between the kinetics of COX-2 protein expression and PGE(2) accumulation, as well as the constitutive nature of COX-1 isoform, indicate that IL-1 beta-driven PGE(2) formation in human bronchial smooth-muscle cells is mediated by de novo expression of COX-2 enzyme.