Rapid and transient induction of cyclo-oxygenase 2 by epidermal growth factor in human amnion-derived WISH cells

Abstract

The central enzyme in the prostaglandin (PG) biosynthetic cascade is PGH2 synthase or cyclo-oxygenase (COX). At present, two distinct isoforms of PGH2 synthase/COX have been identified: COX-1 and COX-2. In many systems, COX-1 is a constitutively expressed isoform that is responsible for normal physiological production of PGs, whereas COX-2 is an inducible isoform that responds to cytokines, endotoxin and growth factors by producing high levels of PGs. The regulation of COX-2 mRNA and protein, and the subsequent production of PGE2, were therefore examined in amnion-derived WISH cells stimulated with epidermal growth factor (EGF). Treatment of WISH cells with EGF (0.01Ő100 ng/ml) elicited dose-dependent synthesis of COX-2 mRNA and protein de novo. In addition, stimulation of WISH cells with EGF (10 ng/ml) induced steady-state levels of COX-2 mRNA and protein that appeared within 30 min and then declined rapidly to near baseline levels within 2Ő4 h. In contrast, COX-1 protein was unchanged in response to treatment with EGF. PGE2 production was also rapid and transient. Preincubation of cells with the novel COX-2 enzymic inhibitor NS-398 (10-5Ő10-10 M) completely prevented PGE2 formation in a dose-dependent manner. Preincubation of cells in dexamethasone (Dex; 0.1 ƁM), however, resulted in only a 31% decrease in PGE2 formation in response to EGF (10 ng/ml) while completely attenuating PGE2 biosynthesis in tumour necrosis factor α (TNF-α)-stimulated cells. In addition, Dex (0.1 ƁM) was only partly effective at preventing EGF-induced COX-2 mRNA and protein expression de novo, whereas Dex completely inhibited TNF-α-promoted COX-2 mRNA and protein expression. Thus the results presented here demonstrate that EGF induces the rapid but transient expression of COX-2 mRNA and protein and the subsequent production of PGE2 in WISH cells.