TNF-α-induced cyclooxygenase-2 expression in human lung epithelial cells:: Involvement of the phospholipase C-γ2, protein kinase C-α, tyrosine kinase, NF-κB-inducing kinase, and I-κB kinase 1/2 pathway

TNF-ru induced a dose- and time-dependent increase in cyclooxygenase-2 (COX-2) expression and PGE, formation in human NCI-H292 epithelial cells. Immunofluorescence staining demonstrated that COX-2 was expressed in cytosol and nuclear envelope. Tyrosine kinase inhibitors (genistein or herbimycin) or phosphoinositide-specific phospholipase C inhibitor (U73122) blocked TNF-alpha -induced COX-2 expression, TNF-alpha also stimulated phosphatidylinositol hydrolysis and protein kinase C (PKC) activity, and both were abolished by genistein or U73122, The PKC inhibitor, staurosporine, also inhibited TNF-alpha -induced response. The 12-O-tetradecanoylphorbol 13-acetate (TPA), a PRC activator, also stimulated COX-2 expression, this effect being inhibited by genistein or herbimycin, NF-kappaB DNA-protein binding and COX-2 promoter activity were enhanced by TNF-alpha, and these effects were inhibited by genistein, U73122, staurosporine, or pyrolidine dithiocarbamate, TPA stimulated both NF-kappaB DNA-protein binding and COX-2 promoter activity, these effects being inhibited by genistein, herbimycin, or pyrolidine dithiocarbamate. The TNF-alpha -induced, but not the TPA-induced, COX-2 promoter activity was inhibited by phospholipase C-gamma2 mutants, and the COX-2 promoter activity induced by either agent was attenuated by dominant-negative mutants of PKC-alpha, NF-kappaB-inducing kinase, or I-kappaB (inhibitory protein that dissociates from NF-kappaB) kinase (IKK)1 or 2, IKK activity was stimulated by both TNF-alpha and TPA, and these effects were inhibited by staurosporine or herbimycin, These results suggest that, in NCI-H292 epithelial cells, TNF-1 gamma might activate phospholipase C-gamma2 via an upstream tyrosine kinase to induce activation of PKC-alpha and protein tyrosine kinase, resulting In the activation of NF-kappaB-inducing kinase and IKK1/2, and NF-kappaB in the COX-2 promoter, then initiation of COX-2 expression and PGE(2) release.