Roflumilast inhibits lipopolysaccharide-induced inflammatory mediators via suppression of nuclear factor-kappaB, p38 mitogen-activated protein kinase, and c-Jun NH2-terminal kinase activation.
Roflumilast, a potent and selective phosphodiesterase 4 (PDE4) inhibitor, has been demonstrated to be an effective anti-inflammatory agent in airway inflammatory diseases. In the present study, we investigated the mechanism of anti-inflammatory effects of roflumilast in murine macrophage cell line RAW264.7 cells. Roflumilast inhibited NO, tumor necrosis factor (TNF)-alpha, and interleukin (IL)-1beta production via suppression of their gene expressions in lipopolysaccharide (LPS)-stimulated macrophages. To elucidate the mechanism by which roflumilast inhibits the production of inflammatory mediators, we examined the effect of roflumilast on the activation of nuclear factor-kappaB (NF-kappaB) in these cells. Roflumilast inhibited the DNA binding activity of NF-kappaB by preventing inhibitor kappaBalpha phosphorylation and degradation. The phosphorylation of mitogen-activated protein (MAP) kinases, including stress-activated protein kinase/c-Jun NH2-terminal kinase (JNK) and p38 MAP kinase, was also markedly inhibited by roflumilast. Similar to the effects of roflumilast, treatment of either SB203580 [4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)imidazole] or SP600125 [anthra(1,9-cd)pyrazol-6(2H)-one 1,9-pyrazoloanthrone], specific inhibitors of p38 MAP kinase and JNK, respectively, suppressed NO, TNF-alpha, and IL-1beta production. Consistent with in vitro results, administration of roflumilast recovered the survival rate of LPS-treated mice, with concurrent suppression of plasma levels of nitrite/nitrate, TNF-alpha, and IL-1beta. These results suggest that the inhibitory activity of roflumilast on the production of inflammatory mediators seems to be mediated via inhibition of NF-kappaB, p38 MAP kinase, and JNK activation in macrophages.
Kwak
Hyun Jeong
HJ
Laboratory of Molecular Pharmacology and Physiology, Medicinal Science Division, Korea Research Institute of Chemical Technology, P.O. Box 107, Yusung-Gu, Taejon 305-343, Korea.
Song
Jin Sook
JS
Heo
Jun Young
JY
Yang
Sung Don
SD
Nam
Ji-Youn
JY
Cheon
Hyae Gyeong
HG
D000631:Aminopyridines / Q000633:toxicity*; D000818:Animals; D001549:Benzamides / Q000633:toxicity*; D002478:Cells, Cultured; D003521:Cyclopropanes / Q000633:toxicity; D024202:Electrophoretic Mobility Shift Assay; D004789:Enzyme Activation / Q000187:drug effects; D007375:Interleukin-1 / Q000032:analysis / Q000037:antagonists & inhibitors; D048031:JNK Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors*; D008070:Lipopolysaccharides / Q000494:pharmacology; D017737:Macrophages, Peritoneal / Q000187:drug effects; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D016328:NF-kappa B / Q000037:antagonists & inhibitors*; D009573:Nitrites / Q000032:analysis; D010726:Phosphodiesterase Inhibitors / Q000633:toxicity*; D018675:Toxicity Tests; D014409:Tumor Necrosis Factor-alpha / Q000032:analysis / Q000037:antagonists & inhibitors; D048051:p38 Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors*
D000631:Aminopyridines; D001549:Benzamides; D003521:Cyclopropanes; D007375:Interleukin-1; D008070:Lipopolysaccharides; D016328:NF-kappa B; D009573:Nitrites; D010726:Phosphodiesterase Inhibitors; D014409:Tumor Necrosis Factor-alpha; C424423:Roflumilast; D048031:JNK Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases