Integrin-linked kinase regulates inducible nitric oxide synthase and cyclooxygenase-2 expression in an NF-kappa B-dependent manner.
Nitric oxide (NO) and prostaglandins are produced as a result of the stimulation of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2, respectively, in response to cytokines or lipopolysaccharide (LPS). We demonstrate that the activity of integrin-linked kinase (ILK) is stimulated by LPS activation in J774 macrophages. Inhibition of ILK activity by dominant-negative ILK or a highly selective small molecule ILK inhibitor, in epithelial cells or LPS-stimulated J774 cells and murine macrophages, resulted in inhibition of iNOS expression and NO synthesis. LPS stimulates the phosphorylation of IkappaB on Ser-32 and promotes its degradation. Inhibition of ILK suppressed this LPS-stimulated IkappaB phosphorylation and degradation. Similarly, ILK inhibition suppressed the LPS-stimulated iNOS promoter activity. Mutation of the NF-kappaB sites in the iNOS promoter abolished LPS- and ILK-mediated regulation of iNOS promoter activity. Overexpression of ILK-stimulated NF-kappaB activity and inhibition of ILK or protein kinase B (PKB/Akt) suppressed this activation. We conclude that ILK can regulate NO production in macrophages by regulating iNOS expression through a pathway involving PKB/Akt and NF-kappaB. Furthermore, we also demonstrate that ILK activity is required for LPS stimulated cyclooxygenase-2 expression in murine and human macrophages. These findings implicate ILK as a potential target for anti-inflammatory applications.
Tan
Clara
C
Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, British Columbia V6H 3Z6, Canada.
Mui
Alice
A
Dedhar
Shoukat
S
D000818:Animals; D002460:Cell Line; D051545:Cyclooxygenase 1; D051546:Cyclooxygenase 2; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D007527:Isoenzymes / Q000235:genetics*; D008070:Lipopolysaccharides / Q000494:pharmacology; D008264:Macrophages; D008565:Membrane Proteins; D051379:Mice; D016328:NF-kappa B / Q000378:metabolism*; D019001:Nitric Oxide Synthase / Q000235:genetics*; D052247:Nitric Oxide Synthase Type II; D010766:Phosphorylation; D011401:Promoter Regions, Genetic / Q000187:drug effects; D011451:Prostaglandin-Endoperoxide Synthases / Q000235:genetics*; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D000099200:Scaffold Protein ILK
D051545:Cyclooxygenase 1; D051546:Cyclooxygenase 2; D007527:Isoenzymes; D008070:Lipopolysaccharides; D008565:Membrane Proteins; D016328:NF-kappa B; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; D011451:Prostaglandin-Endoperoxide Synthases; D017346:Protein Serine-Threonine Kinases; D011993:Recombinant Fusion Proteins; D000099200:Scaffold Protein ILK; C496319:NOS2 protein, human; C496317:Nos2 protein, mouse; C496511:PTGS1 protein, human; C496540:PTGS2 protein, human; C496512:Ptgs1 protein, mouse