LTD4 INDUCES HB-EGF-DEPENDENT CXCL8 RELEASE THROUGH EGFR ACTIVATION IN HUMAN BRONCHIAL EPITHELIAL CELLS.

Airway epithelial cells release proinflammatory mediators that may contribute to airway remodeling and leukocyte recruitment. We explored the hypothesis that leukotriene D4 (LTD4) may trigger the release of proremodeling factors through activation of the EGF receptor (EGFR). We particularly focused on the effects of LTD4 on release of heparin-binding EGF-like factor (HB-EGF) and IL-8 (CXCL8), a potent neutrophil chemoattractant that may be released downstream of EGFR activation. To address this hypothesis, both primary (NHBE) and transformed bronchial human epithelial cells (BEAS-2B) were grown on an air-liquid interface and stimulated with LTD4. HB-EGF and CXCL8 were evaluated by ELISA in cell culture supernatants. To explore the EGFR signaling pathway, we used a broad-spectrum matrix metalloproteinase (MMP) inhibitor, GM-6001, two selective EGFR tyrosine kinase inhibitors, AG-1478 and PD-153035, an HBEGF neutralizing antibody, and a specific small interfering RNA (siRNA) against the EGFR. Expression of the CysLT1 cysteinyl leukotriene receptor was demonstrated by RT-PCR and immunocytochemistry in both BEAS-2B and NHBE cells. Four hours after stimulation with LTD4, HB-EGF and CXCL8 were significantly increased in cell culture supernatant. GM-6001 and montelukast, a specific CysLT1 receptor antagonist, blocked the LTD4-induced increase in HB-EGF. All inhibitors/antagonists decreased LTD4-induced CXCL8 release. siRNA against EGFR abrogated CXCL8 release following stimulation with LTD4 and exogenous HB-EGF. These findings suggest LTD4 induced EGFR transactivation through the release of HB-EGF in human bronchial epithelial cells with downstream release of CXCL8. These effects may contribute to epithelial-mediated airway remodeling in asthma and other conditions associated with cysteinyl leukotriene release.

AIR-100 , Airway remodeling, Amphiregulin, BEAS-2B, comparison, CysLT1 Cysteinyl leukotriene receptor, Cysteinyl leukotriene receptor, Cysteinyl leukotrienes, EGF receptor, Heparin binding EGF-like factor (HB-EGF), Histology, IL-8 (CXCL8), siRNA, TGF-α, Tight junctions

EGFR tyrosine kinase inhibitors, AG-1478, Leukotriene D4(LTD4), Matrix metalloproteinase (MMP) inhibitor, GM-6001, Montelukast, PD-153035