Journal
FREE RADICAL BIOLOGY AND MEDICINE
Volume 48, Issue 2, Pages 240-254Publisher
ELSEVIER SCIENCE INC
DOI: 10.1016/j.freeradbiomed.2009.10.047
Keywords
Cigarette smoke; Oxidative stress; Free radicals; Cyclooxygenase-2; Prostaglandin E-2; Human tracheal smooth muscle cells
Funding
- [NSC95-2320-B182-010]
- [NSC95-2314-B182-040]
- [CMRPD170331]
- [CMRPG350652]
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Exposure to cigarette smoke extract (CSE) leads to airway and lung inflammation through an oxidant-antioxidant imbalance. Cyclooxygenase-2 (COX-2) and prostaglandin E-2 (PGE(2)) have been shown to play critical roles in respiratory inflammation. Here, we show that COX-2/PGE(2)/IL-6 induction is dependent on Toll-like receptor 4 (TLR4)/NADPH oxidase signaling in human tracheal smooth muscle cells (HTSMCs). CSE induced COX-2 expression in vitro in HTSMCs and in vivo in the airways of mice. CSE also directly caused an increase in TLR4. Moreover, CSE-regulated COX-2, PGE(2), and IL-6 generation was inhibited by pretreatment with TLR4 Ab: inhibitors of c-Src (PP1), NADPH oxidase (diphenylene iodonium chloride and apocynin), p38 MAPK (SB202190), MEK1/2 (U0126), JNK1/2 (SP600125), and NF-kappa B (helenalin); a ROS scavenger (N-acetyl-L-cysteine); and transfection with siRNA of TLR4, MyD88, TRAF6, Src, p47(phox), p38, p42, JNK2, or p65. CSE-induced leukocyte numbers in BAL fluid were also reduced by pretreatment with these inhibitors. Furthermore, CSE induced p47(phox) translocation and TLR4/MyD88/TRAF6 and c-Src/p47(phox), complex formation. We found that PGE(2) enhanced IL-6 production in HTSMCs and leukocyte count in BAL fluid. In addition, treatment with nicotine could induce COX-2, PGE(2), and IL-6 generation in in vivo and in Vitro Studies. These results demonstrate that CSE-induced ROS generation was mediated through the TLR4/MyD88/TRAF6/c-Src/NADPH oxidase pathway, in turn initiated the activation of MAPKs and NF-kappa B, and ultimately induced COX-2/PGE(2)/IL-6-dependent airway inflammation. (C) 2009 Elsevier Inc. All rights reserved.
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