Tumor necrosis factor α (TNF-α)-induced prostaglandin E_2 release is mediated by the activation of cyclooxygenase-2 (COX-2) transcription via NFκB in human gingival fibroblasts

摘要

Nuclear factor kappaB (NFκB) 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 NFκB. TNF-α is a pro--inflammatory cytokine. In this paper, we investigated the involvement of NFκB on TNF-α-mediated prostaglandin E_2 (PGE_2) release and COX-2 gene expression in human gingival fibroblasts (HGF). TNF-α-induced PGE_2 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～+59) ligated to a luciferase reporter gene indicated that TNF-α stimulated the transcriptionalo activity ～1.4-fold. Gel mobility shift assays with a radiolabelled COX-2-NFκB oli-gonucleotide (nts - 223 to -214) revealed an increase in the binding of nuclear proteins from TNF-α-stimulated HGF. The COX-2-NFκB 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-α-stimulated PGE_2 release. These results suggest that NFκB transcription factor is key regulator of COX02 expression in TNF-α-induced PGE_2 production, which is mediated through a tyrosine kinase pathway in HGF.