Lipoteichoic acid-induced nitric oxide synthase expression in RAW 264.7 macrophages is mediated by cyclooxygenase-2, prostaglandin E2, protein kinase A, p38 MAPK, and nuclear factor-kappaB pathways.
We recently reported that lipoteichoic acid (LTA), a cell wall component of the gram-positive bacterium Staphylococcus aureus, stimulated inducible nitric oxide synthase (iNOS) expression, nitric oxide (NO) release, and cyclooxygenase-2 (COX-2) expression in RAW 264.7 macrophages. This study was carried out to further investigate the roles of COX-2 and prostaglandin E2 (PGE2) in LTA-induced iNOS expression and NO release in RAW 264.7 macrophages. Treatment of RAW 264.7 macrophages with LTA caused a time-dependent increase in PGE2 release. LTA-induced iNOS expression and NO release were inhibited by a non-selective COX inhibitor (indomethacin), a selective COX-2 inhibitor (NS-398), an adenylyl cyclase (AC) inhibitor (dideoxyadenosine, DDA), and a protein kinase A (PKA) inhibitor (KT-5720). Furthermore, both PGE2 and the direct PKA activator, dibutyryl-cAMP, also induced iNOS expression in a concentration-dependent manner. Stimulation of RAW 264.7 macrophages with LTA, PGE2, and dibutyryl-cAMP all caused p38 MAPK activation in a time-dependent manner. LTA-mediated p38 MAPK activation was inhibited by indomethacin, NS-398, and SB 203580, but not by PD 98059. The PGE2-mediated p38 MAPK activation was inhibited by DDA, KT-5720, and SB 203580, but not by PD 98059. LTA caused time-dependent activation of the nuclear factor-kappaB (NF-kappaB)-specific DNA-protein complex formation. The LTA-induced increase in kappaB-luciferase activity was inhibited by indomethacin, NS-398, KT-5720, and a dominant negative mutant of p38 alphaMAPK (p38 alphaMAPK DN). These results suggest that LTA-induced iNOS expression and NO release involve COX-2-generated PGE2 production, and AC, PKA, p38 MAPK, and NF-kappaB activation in RAW 264.7 macrophages.
Chang
Yau-Chong
YC
Department of Cardiac Surgery, Taipei Medical University, Wang Fang Hospital, Taipei, Taiwan; Graduate Institute of Injury Prevention, College of Public Health and Nutrition, Taipei Medical University, Taipei, Taiwan.
Li
Pei-Chih
PC
Chen
Bing-Chang
BC
Chang
Ming-Shyan
MS
Wang
Jui-Ling
JL
Chiu
Wen-Ta
WT
Lin
Chien-Huang
CH
D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000494:pharmacology; D054746:Cyclic AMP-Dependent Protein Kinase Type II; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D051546:Cyclooxygenase 2 / Q000378:metabolism*; D015232:Dinoprostone / Q000096:biosynthesis / Q000378:metabolism / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D007213:Indomethacin / Q000494:pharmacology; D007700:Kinetics; D008070:Lipopolysaccharides / Q000494:pharmacology*; D020935:MAP Kinase Signaling System / Q000187:drug effects; D008264:Macrophages / Q000187:drug effects* / Q000201:enzymology; D051379:Mice; D008954:Models, Biological; D016328:NF-kappa B / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II / Q000378:metabolism*; D009573:Nitrites / Q000378:metabolism; D013682:Teichoic Acids / Q000494:pharmacology*; D013997:Time Factors; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism*
D008070:Lipopolysaccharides; D016328:NF-kappa B; D009573:Nitrites; D013682:Teichoic Acids; D009569:Nitric Oxide; C009900:lipoteichoic acid; D000242:Cyclic AMP; D052247:Nitric Oxide Synthase Type II; D051546:Cyclooxygenase 2; D054746:Cyclic AMP-Dependent Protein Kinase Type II; D017868:Cyclic AMP-Dependent Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D000262:Adenylyl Cyclases; D015232:Dinoprostone; D007213:Indomethacin