YC-1-induced cyclooxygenase-2 expression is mediated by cGMP-dependent activations of Ras, phosphoinositide-3-OH-kinase, Akt, and nuclear factor-kappaB in human pulmonary epithelial cells.
We demonstrated previously that 3-(5'-hydroxymethyl-2'-furyl)-1-benzylindazole (YC-1), an activator of soluble guanylate cyclase (sGC), induces cyclooxygenase-2 (COX-2) expression via cGMP- and p44/42 mitogen-activated protein kinase-dependent pathways in human pulmonary epithelial A549 cells. In this study, we explore the role of Ras, phosphoinositide-3-OH-kinase (PI3K), Akt, and transcription factor nuclear factor-kappaB (NF-kappaB) in YC-1-induced COX-2 expression in A549 cells. A Ras inhibitor (manumycin A), a PI3K inhibitor (wortmannin), an Akt inhibitor (1l-6-Hydroxymethyl-chiro-inositol2-[(R)-2-O-methyl-3-O-octadecylcarbonate]), and an NF-kappaB inhibitor [pyrrolidine dithiocarbamate (PDTC)] all reduced YC-1-induced COX-2 expression. The YC-1-induced increase in COX activity was also blocked by manumycin A, wortmannin, PDTC, and the dominant-negative mutants for Ras (RasN17), Akt (Akt DN), and IkappaBalpha (IkappaBalphaM). The YC-1-induced increase in Ras activity was inhibited by an sGC inhibitor [1H-(1,2,4)oxadiazolo[4,3-a]quinozalin-1-one (ODQ)], a protein kinase G (PKG) inhibitor [1-oxo-9.12-epoxy-1H-diindolo[1,2,3-fg:3',2',1'-kl]pyrrolo[3,4-I][1,6]benzodiazocine-10-carboxylic acid methyl ester (KT-5823)], and manumycin A. YC-1-induced Akt activation was also inhibited by ODQ, KT-5823, manumycin A, and wortmannin. YC-1 caused the formation of an NF-kappaB-specific DNA-protein complex and an increase in kappaB-luciferase activity. YC-1-induced kappaB-luciferase activity was inhibited by ODQ, KT-5823, manumycin A, wortmannin, an Akt inhibitor, PDTC, RasN17, Akt DN, and IkappaBalphaM. Likewise, YC-1-induced IKKalpha/beta activation was inhibited by ODQ, KT-5823, manumycin A, wortmannin, and an Akt inhibitor. Furthermore, YC-1-induced COX-2 promoter activity was inhibited by manumycin A, RasN17, Akt DN, PDTC, and IkappaBalphaM. Taken together, these results indicate that YC-1 might activate the sGC/cGMP/PKG pathway to induce Ras and PI3K/Akt activation, which in turn initiates IKKalpha/beta and NF-kappaB activation and finally induces COX-2 expression in A549 cells.
Chang
Ming-Shyan
MS
Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, 250 Wu-Hsing Street, Taipei 110, Taiwan.
Lee
Wen-Sen
WS
Chen
Bing-Chang
BC
Sheu
Joen-Rong
JR
Lin
Chien-Huang
CH
D006152:Cyclic GMP / Q000031:analogs & derivatives / Q000378:metabolism* / Q000494:pharmacology; D017869:Cyclic GMP-Dependent Protein Kinases / Q000378:metabolism; D051546:Cyclooxygenase 2; D015232:Dinoprostone / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004847:Epithelial Cells / Q000187:drug effects / Q000201:enzymology; D015870:Gene Expression; D006801:Humans; D051550:I-kappa B Kinase; D020672:I-kappa B Proteins / Q000378:metabolism; D007191:Indazoles / Q000737:chemistry / Q000494:pharmacology*; D007375:Interleukin-1 / Q000378:metabolism; D007527:Isoenzymes / Q000378:metabolism*; D008168:Lung / Q000166:cytology; D008565:Membrane Proteins; D009154:Mutation; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D011090:Polyenes / Q000494:pharmacology; D053284:Polyunsaturated Alkamides; D011392:Proline / Q000031:analogs & derivatives* / Q000494:pharmacology; D011451:Prostaglandin-Endoperoxide Synthases / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt; D013859:Thiocarbamates / Q000494:pharmacology; D014157:Transcription Factors / Q000378:metabolism; D018631:ras Proteins / Q000378:metabolism*
D006152:Cyclic GMP; D017869:Cyclic GMP-Dependent Protein Kinases; D051546:Cyclooxygenase 2; D015232:Dinoprostone; D004791:Enzyme Inhibitors; D051550:I-kappa B Kinase; D020672:I-kappa B Proteins; D007191:Indazoles; D007375:Interleukin-1; D007527:Isoenzymes; D008565:Membrane Proteins; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B; D019869:Phosphatidylinositol 3-Kinases; D011090:Polyenes; D053284:Polyunsaturated Alkamides; D011392:Proline; D011451:Prostaglandin-Endoperoxide Synthases; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D013859:Thiocarbamates; D014157:Transcription Factors; D018631:ras Proteins; C000606204:NFKBIA protein, human; C066229:prolinedithiocarbamate; C090937:3-(5'-hydroxymethyl-2'-furyl)-1-benzylindazole; C016276:8-bromocyclic GMP; C496540:PTGS2 protein, human; C494918:AKT1 protein, human; C496559:CHUK protein, human; C496560:IKBKB protein, human; C496565:IKBKE protein, human; C054474:manumycin