Journal
MOLECULAR AND CELLULAR BIOCHEMISTRY
Volume 209, Issue 1-2, Pages 113-118Publisher
KLUWER ACADEMIC PUBL
DOI: 10.1023/A:1007155525020
Keywords
COX-2; gene regulation; IL-1 beta; NF kappa B; tyrosine phosphorylation; human gingival fibroblasts
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The immediate-early cyclooxygenase-2 (COX-2) gene encodes an inducible prostaglandin synthase enzyme which is implicated in inflammatory and proliferative diseases. COX-2 is highly induced during cell activation by various factors, including mitogens, hormones and cytokines. Since pro-inflammatory cytokine IL-1 beta has been shown to induce prostaglandin E-2 (PGE(2)) release in human gingival fibroblasts (HGF), here we analyzed the effect of IL-1 beta on the expression of COX-2 and the activation of NF kappa B in HGF. Northern hybridization analysis revealed that IL-1 beta (200 pg/ml) increased the expression of COX-2 mRNA in HGF. The effect of IL-1 beta was abrogated by herbimycin A, a protein tyrosine kinase inhibitor, and enhanced by orthovanadate, a protein tyrosine phosphatase inhibitor. IL-1 beta-induced PGE(2) release was blocked by the tyrosine kinase inhibitor and increased by the tyrosine phosphatase inhibitor. The results of transient transfection assays using chimeric constructs of the human COX-2 promoter (nt -1432 +59) ligated to a luciferase reporter gene indicated that IL-1 beta stimulated the transcriptional activity 1.5-fold. Gel mobility shift assays with a radiolabelled COX-2-NF kappa B oligonucleotide (nts -223 to -214) revealed an increase in the binding of nuclear proteins from IL-1 beta-stimulated HGF. This increase of DNA-protein complex formation induced by IL-1 beta was blocked by herbimycin A and another tyrosine kinase inhibitor, genistein. These results suggest that NF kappa B is an important transcription factor for IL-1 beta-induced COX-2 gene expression, and is involved in inducing COX-2 gene transcription through tyrosine phosphorylation in HGF.
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