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-Induced Cyclooxygenase-2 Expression in Human Lung Epithelial Cells: Involvement of the Phospholipase C-
2, Protein Kinase C-, Tyrosine Kinase, NF-
B-Inducing Kinase, and I-B Kinase 1/2 Pathway1
Institute of Pharmacology, College of Medicine, National Taiwan University, Taipei, Taiwan
TNF-
induced a dose- and time-dependent increase in
cyclooxygenase-2 (COX-2) expression and PGE2 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--induced COX-2 expression. TNF- 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--induced response. The
12-O-tetradecanoylphorbol 13-acetate (TPA), a PKC
activator, also stimulated COX-2 expression, this effect being
inhibited by genistein or herbimycin. NF-
B DNA-protein binding and
COX-2 promoter activity were enhanced by TNF-, and these effects
were inhibited by genistein, U73122, staurosporine, or pyrolidine
dithiocarbamate. TPA stimulated both NF-B DNA-protein binding and
COX-2 promoter activity, these effects being inhibited by genistein,
herbimycin, or pyrolidine dithiocarbamate. The TNF--induced, but not
the TPA-induced, COX-2 promoter activity was inhibited by phospholipase
C-
2 mutants, and the COX-2 promoter activity induced by either agent
was attenuated by dominant-negative mutants of PKC-,
NF-B-inducing kinase, or I-B (inhibitory protein that dissociates
from NF-B) kinase (IKK)1 or 2. IKK activity was stimulated by both
TNF- and TPA, and these effects were inhibited by staurosporine or
herbimycin. These results suggest that, in NCI-H292 epithelial cells,
TNF- might activate phospholipase C-2 via an upstream tyrosine
kinase to induce activation of PKC- and protein tyrosine kinase,
resulting in the activation of NF-B-inducing kinase and IKK1/2, and
NF-B in the COX-2 promoter, then initiation of COX-2 expression and
PGE2 release.
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