Cigarette smoke extract induces COX-2 expression via a PKC /c-Src/EGFR, PDGFR/PI3K/Akt/NF- B pathway and p300 in tracheal smooth muscle cells

Yang C-M, Lee I-T, Lin C-C, Yang Y-L, Luo S-F, Kou YR, Hsiao L-D. Cigarette smoke extract induces COX-2 expression via a PKC /c-Src/EGFR, PDGFR/PI3K/Akt/NFB pathway and p300 in tracheal smooth muscle cells. Am J Physiol Lung Cell Mol Physiol 297: L892–L902, 2009. First published August 28, 2009; doi:10.1152/ajplung.00151.2009.—Exposure to cigarette smoke extract (CSE) leads to airway or lung inflammation, which may be mediated through cyclooxygenase-2 (COX-2) expression and its product prostaglandin E2 (PGE2) synthesis. The aim of this study was to investigate the molecular mechanisms underlying CSE-induced COX-2 expression in human tracheal smooth muscle cells (HTSMCs). Here, we describe that COX-2 induction is dependent on PKC /cSrc/EGFR, PDGFR/PI3K/Akt/NFB signaling in HTSMCs. CSE stimulated the phosphorylation of c-Src, EGFR, PDGFR, and Akt, which were inhibited by pretreatment with the inhibitor of PKC (Gö6976 or Gö6983), c-Src (PP1), EGFR (AG1478), PDGFR (AG1296), or PI3K (LY294002). Moreover, CSE induced a significant increase in COX-2 expression, which was reduced by pretreatment with these inhibitors or transfection with siRNA of PKC , Src, or Akt. Furthermore, CSE-stimulated NFB p65 phosphorylation and translocation were also attenuated by pretreatment with Gö6976, PP1, AG1478, AG1296, or LY294002. CSE-induced COX-2 expression was also mediated through the recruitment of p300 associated with NFB in HTSMCs, revealed by coimmunoprecipitation and Western blot analysis. In addition, pretreatment with the inhibitors of NFB (helenalin) and p300 (garcinol) or transfection with p65 siRNA and p300 siRNA markedly inhibited CSE-regulated COX-2 expression. However, CSE-induced PGE2 generation was reduced by pretreatment with the inhibitor of COX-2 (NS-398). These results demonstrated that in HTSMCs, CSE-induced COX-2-dependent PGE2 generation was mediated through PKC /c-Src/EGFR, PDGFR/PI3K/Akt leading to the recruitment of p300 with NFB complex.