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

Abstract

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 PKCalpha/c-Src/EGFR, PDGFR/PI3K/Akt/NF-kappaB signaling in HTSMCs. CSE stimulated the phosphorylation of c-Src, EGFR, PDGFR, and Akt, which were inhibited by pretreatment with the inhibitor of PKCalpha (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 PKCalpha, Src, or Akt. Furthermore, CSE-stimulated NF-kappaB 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 NF-kappaB in HTSMCs, revealed by coimmunoprecipitation and Western blot analysis. In addition, pretreatment with the inhibitors of NF-kappaB (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 PKCalpha/c-Src/EGFR, PDGFR/PI3K/Akt leading to the recruitment of p300 with NF-kappaB complex.

Cigarette Smoke Activates the Proto-Oncogene c-Src to Promote Airway Inflammation and Lung Tissue Destruction

“…Although c-Src is typically viewed as a downstream target of EGFR, cSrc was required for the tyrosine phosphorylation and hence activation of EGFR in CSE-treated SAE cells. This study and others have shown that cigarette smoke phosphorylates c-Src via a PKCa-dependent mechanism (9,29). Once activated by cigarette smoke, c-Src phosphorylated EGFR and KRAS/Raf, and these signaling proteins further activated cSrc.…”

“…In fact, studies indicate that CSE can stimulate the phosphorylation of c-Src, EGFR, PDGFR, and Akt in a PKC-a-dependent manner (9). For this reason, we examined how PKC-a silencing affected c-Src-mediated inflammation in SAE cells.…”

“…Conversely, phosphorylation at site Tyr527 on the C-terminal end leads to binding of this region to the SH2 domain and negatively regulates kinase activity (8). Cigarette smoke is known to activate c-Src kinase through the phosphorylation of c-Src at Tyr416 in tumor cells and in tracheal smooth muscle cells (9). c-Src activates several signaling cascades in cancer that have been implicated in COPD, such as focal adhesion kinase (FAK) (10), signal transducers and activators of transcription 3 (STAT-3) (11), the epidermal growth factor receptor (EGFR) family (12), phosphatidylinositol 3-kinase (PI3K)-Akt, and mitogen-activated protein kinase (MAPK) (9).…”

Cyclooxygenase-2 and MicroRNA-155 Expression Are Elevated in Asthmatic Airway Smooth Muscle Cells

“…MiR-155-mediated repression of SHIP1 led to enhanced activation of the phosphoinositide-3-kinase (PI3K)/AKT pathway in IB3-1 cells. In hASMCs, the PI3K/AKT pathway was observed to partly mediate cigarette smoke extract induction of COX-2 in hASMCs (47). Furthermore, AKT was previously observed to enhance the stability of COX-2 mRNA in rat intestinal epithelial cells (48).…”

Si̇gara Dumaninin Neden Olduğu Akci̇ğer İnflamasyonuna Karşi Alfa-Li̇noleni̇k Asi̇ti̇n Koruyucu Rolü

“…Sigara dumanının, inflamasyon mekanizmalarında önemli rolleri olan siklooksijenaz-2 ve fosfolipaz A2 enzimini arttırdığı diğer çalışmalarda gösterilmiştir (23)(24). Bizim çalışmamızda da benzer şekilde sigara dumanı siklooksijenaz-2, Fosfolipaz A2 ve iNOS enzimlerinin miktarında artışa neden olurken antiinflamatuar özelliği gösterilen alfalinolenik asit bu artışları anlamlı olarak azalttı.…”