AbstractsBiology & Animal Science

Does the epidermal growth factor receptor mediate airway epithelial repair following a single chlorine exposure in mice?

by Neil Verma

Institution: McGill University
Department: Department of Physiology
Degree: MS
Year: 2015
Keywords: Biology - Physiology
Record ID: 2057894
Full text PDF: http://digitool.library.mcgill.ca/thesisfile130492.pdf


Chlorine gas exposure can occur through inadvertent mixing of household cleaning products, as a result of industrial accidents and intentionally in chemical warfare. Chlorine dissolves in the epithelial lung lining fluid to produce hypochlorous and hydrochloric acid, which cause oxidative stress. Oxidative stress has been shown to activate matrix metalloproteinases and to stimulate the synthesis of cysteinyl leukotrienes, which may, in turn, release ligands for the epidermal growth factor receptor (EGFR). The release of EGFR ligands and an increase in the IL-8 ortholog CXCL1 post chlorine in the mouse suggest the possible involvement of the EGFR in chlorine-induced airway injury. The hypothesis addressed in this thesis is that the EGFR mediates airway epithelial repair following chlorine exposure in mice. We studied the time-course of chlorine injury in the BALB/C mouse at 2, 6, 24 and 48 hours post nose-only exposure to 100ppm of Cl2 for 5 minutes. The time course of changes involving the EGFR-EGFR ligand axis was studied at the transcriptional and protein levels using qPCR, Western blots and ELISAs to detect EGFR, heparin-binding EGF-like growth factor (HB-EGF) and amphiregulin (AREG) expression levels. The latter two proteins are ligands of the EGFR. Airway morphometry was used to quantify apoptotic cell numbers and proliferation visualized with TUNEL and PCNA immunostaining respectively. Airway inflammation was quantified with a total cell count and a differential cell count. Gefitinib, an EGFR inhibitor, was administered via oral gavage (50mg/kg) to assess the effect of inhibiting the EGFR pathway on chlorine injury, epithelial repair and airway mechanics (respiratory resistance and elastance) in response to methacholine challenge. Total EGFR protein was decreased following chlorine exposure starting at 2 hours onwards with a gradual recovery by 24 hours while the transcription of EGFR was upregulated at 6h and 24h post exposure. Transcriptional levels of HB-EGF and AREG were not upregulated following chlorine exposure. Increased transcription of EGFR was prevented by the antioxidant DMTU (100mg/kg) given i.p. 1 hour prior to chlorine. HB-EGF ligand concentrations in the BALF increased with chlorine exposure and were also inhibited by DMTU, suggesting oxidative stress-dependent ligand release. Gefitinib inhibited airway hyperresponsiveness to methacholine relative to controls and diminished neutrophilia in the BALF at 6 hours post-chlorine exposure. The EGFR-EGFR ligand axis seems to be involved in the development of chlorine injury with a rapid increase in the receptor's ligand in BALF. Inhibitors of EGFR may reduce AHR as a consequence of inhibiting the neutrophilic inflammation, which may exacerbate the original injury caused by chlorine gas. L'inhalation de vapeurs de chlore peut arriver à la suite d'un mélange accidentel de produits industriels, à la suite d'accidents du travail et intentionnellement pendant les attaques terroristes. Le chlore se dissout dans le liquide sur la surface des voies…