Induction of cytosolic phospholipase A2 by lipopolysaccharide in canine tracheal smooth muscle cells: involvement of MAPKs and NF-kappaB pathways.
Cytosolic phospholipase A2 (cPLA2) plays a pivotal role in mediating agonist-induced arachidonic acid (AA) release for prostaglandins (PG) synthesis induced by bacterial lipopolysaccharide (LPS) and cytokines. However, the intracellular signaling pathways mediating LPS-induced cPLA2 expression and PGE2 synthesis in canine tracheal smooth muscle cells (TSMCs) remains unknown. LPS-induced expression of cPLA2 and release of PGE2 was attenuated by inhibitors of tyrosine kinase (genistein), phosphatidylcholine-phospholipase C (D609), phosphatidylinositol-phospholipase C (U73122), PKC (GF109203X and staurosporine), removal of Ca2+ by BAPTA/AM plus EDTA, MEK1/2 (PD98059), p38 (SB202190), JNK (SP600125), and phosphatidylinositol 3-kinase (PI3-K; LY294002 and wortmannin). The involvement of MPAKs in LPS-induced responses was further confirmed by transfection of TSMCs with dominant negative mutants of ERK2 and p38. LPS-induced cPLA2 expression and PGE2 synthesis was inhibited by a selective NF-kappaB inhibitor (helenalin) and transfection with dominant negative mutants of NF-kappaB inducing kinase (NIK), IkappaB kinase (IKK)-alpha, and IKK-beta, consistent with that LPS-stimulated both IkappaB-alpha degradation and NF-kappaB translocation into nucleus in these cells. LPS-stimulated cPLA2 phosphorylation was inhibited by PD98059, GF109203X, and staurosporine, indicating the regulation by p42/p44 MAPK and PKC. Moreover, LPS-induced up-regulation of cPLA2 and COX-2 linked to PGE2 synthesis was inhibited by AACOCF3 (a selective cPLA2 inhibitor), implying the involvement of cPLA2 in these responses. These findings suggest that phosphorylation and expression of cPLA2 correlates with the release of PGE2 from LPS-challenged TSMCs, at least in part, mediated through MAPKs and NF-kappaB signaling pathways. LPS-mediated responses were modulated by PLC, Ca2+, PKC, tyrosine kinase, and PI3-K in TSMCs.
Luo
Shue-Fen
SF
Department of Internal Medicine, College of Medicine, Chang Gung University, Division of Allergy, Immunology, and Rheumatology, Chang Gung Memorial Hospital, 5 Fu-Shin Street, Kwei-San, Tao-Yuan, Taiwan. lsf@00076@adm.cgmh.org.tw
Lin
Wei-Ning
WN
Yang
Chuen-Mao
CM
Lee
Chiang-Wen
CW
Liao
Chang-Hui
CH
Leu
Yann-Lii
YL
Hsiao
Li-Der
LD
D000818:Animals; D001952:Bridged-Ring Compounds / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D003600:Cytosol / Q000201:enzymology*; D015232:Dinoprostone / Q000096:biosynthesis; D004285:Dogs; D004790:Enzyme Induction / Q000187:drug effects; D004963:Estrenes / Q000494:pharmacology; D019833:Genistein / Q000494:pharmacology; D008070:Lipopolysaccharides / Q000494:pharmacology*; D020935:MAP Kinase Signaling System / Q000187:drug effects*; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D032389:Myocytes, Smooth Muscle / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D016328:NF-kappa B / Q000037:antagonists & inhibitors / Q000378:metabolism*; D009636:Norbornanes; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010741:Phospholipases A / Q000037:antagonists & inhibitors / Q000096:biosynthesis*; D054467:Phospholipases A2; D010766:Phosphorylation / Q000187:drug effects; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D011760:Pyrrolidinones / Q000494:pharmacology; D013859:Thiocarbamates; D013871:Thiones / Q000494:pharmacology; D014132:Trachea / Q000166:cytology*
D001952:Bridged-Ring Compounds; D004963:Estrenes; D008070:Lipopolysaccharides; D016328:NF-kappa B; D009636:Norbornanes; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011760:Pyrrolidinones; D013859:Thiocarbamates; D013871:Thiones; C060229:1-(6-((3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione; C046498:tricyclodecane-9-yl-xanthogenate; D019833:Genistein; D011493:Protein Kinase C; D020928:Mitogen-Activated Protein Kinases; D010741:Phospholipases A; D054467:Phospholipases A2; D015232:Dinoprostone; D002118:Calcium