Induction of cyclooxygenase-2 by platelet-derived growth factor (PDGF) and its inhibition by dexamethasone are independent of NF-κB/IκB transcription factors

Gucocorticoids are potent inhibitors of cyclooxygenase-2 (prostaglandin G/H synthase-2, COX-2) expression. The focus of this work was to investigate the molecular mechanisms, by which glucocorticoids interfere with platelet-derived growth factor (PDGF)-mediated induction of COX-2 with special emphasis on the role of the transcription factors NF-kappa B/I kappa B alpha. In rat renal mesangial cells. PDGF induced a rapid and transient increase of COX-2 mRNA and protein, which reached maximal levels after 1-2 and 4 h, respectively. The in vivo half-life of COX-2 mRNA, which was estimated to be less than 1 h, was reduced by dexamethasone. Kinetic studies and COX-2 promoter activity assays indicated that dexamethasone also interfered with COX-2 transcription. Inhibition of COX-2 induction by dexamethasone was abrogated by cycloheximide, an inhibitor of translation, indicating dependence on de novo protein synthesis. As a possible mediator of dexamethasone action, the NF-kappa B/I kappa B alpha system of transcription factors was investigated. Dexamethasone doubled I kappa B alpha protein levels within 1 h anti reduced complex formation of nuclear NF-kappa B proteins with DNA. Newly synthesized I kappa B alpha may thus bind to NF-kappa B and interfere with gene activation. PDGF-induced signalling, however, barely affected the NF-kappa B/I kappa B alpha s\i stem: I kappa B alpha protein remained unaltered for 30 min after treatment of mesangial cells with PDGF and was only reduced by 30% after h. Concomitantly, binding of NF-kappa B proteins to DNA, detected by electrophoretic mobility shift assays, was slightly increased by 30%. Furthermore, stably transfected COX-2 promoter constructs with and without the NF-kappa B binding site were comparably activated by PDGF (2.5-fold increase of luciferase activity). Taken together, these data indicate that although dexamethasone interferes with the NF-kappa B/I kappa B alpha system of transcription factors, this mechanism is not essential for the inhibition of PDGF-induced COX-2 expression.