Tumor necrosis factor alpha (TNF-alpha)-induced prostaglandin E2 release is mediated by the activation of cyclooxygenase-2 (COX-2) transcription via NFkappaB in human gingival fibroblasts.
Nuclear factor kappaB (NFkappaB) is a transcription factor and plays a key role in the expression of several genes involved in the inflammatory process. Cyclooxygenase (COX) is the key regulatory enzyme of the prostaglandin/eicosanoid synthetic pathway. COX-2 is a highly inducible enzyme by proinflammatory cytokines, of which gene expression is regulated by NFkappaB. TNF-alpha is a pro-inflammatory cytokine. In this paper, we investigated the involvement of NFkappaB on TNF-alpha-mediated prostaglandin E2 (PGE2) release and COX-2 gene expression in human gingival fibroblasts (HGF). TNF-alpha-induced PGE2 release and COX-2 mRNA accumulation in a time- and concentration-dependent manner in HGF. The results of transient transfection assays using a chimeric construct of the human COX-2 promoter (nts -1432 approximately +59) ligated to a luciferase reporter gene indicated that TNF-a stimulated the transcriptional activity approximately 1.4-fold. Gel mobility shift assays with a radiolabelled COX-2-NFkappaB oligonucleotide (nts -223 to -214) revealed an increase in the binding of nuclear proteins from TNF-alpha-stimulated HGF. The COX-2-NFKB DNA-protein complex disappeared after treatment with pyrrolidine dithiocarbamate (PDTC; an antioxidant) or herbimycin A (a tyrosine kinase inhibitor). PDTC and herbimycin A attenuated TNF-alpha-stimulated PGE2 release. These results suggest that NFkappaB transcription factor is a key regulator of COX-2 expression in TNF-alpha-induced PGE2 production, which is mediated through a tyrosine kinase pathway in HGF.
D000818:Animals; D000975:Antioxidants / Q000494:pharmacology; D001665:Binding Sites; D002417:Cattle; D002478:Cells, Cultured; D051546:Cyclooxygenase 2; D015232:Dinoprostone / Q000378:metabolism*; D024202:Electrophoretic Mobility Shift Assay; D004790:Enzyme Induction / Q000187:drug effects*; D005347:Fibroblasts / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism; D005881:Gingiva / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D006801:Humans; D007527:Isoenzymes / Q000235:genetics*; D008565:Membrane Proteins; D016328:NF-kappa B / Q000378:metabolism*; D011451:Prostaglandin-Endoperoxide Synthases / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D013997:Time Factors; D014158:Transcription, Genetic / Q000187:drug effects*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
D000975:Antioxidants; D007527:Isoenzymes; D008565:Membrane Proteins; D016328:NF-kappa B; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D051546:Cyclooxygenase 2; C496540:PTGS2 protein, human; D011451:Prostaglandin-Endoperoxide Synthases; D015232:Dinoprostone