Abstract
BackgroundLung cancer often exhibits molecular changes, such as the overexpression of the ErbB1 gene. ErbB1 encodes epidermal growth factor receptor (EGFR), a tyrosine kinase receptor, involved mainly in cell proliferation and survival. EGFR overexpression has been associated with more aggressive disease, poor prognosis, low survival rate and low response to therapy. ErbB1 amplification and mutation are associated with tumor development and are implicated in ineffective treatment. The aim of the present study was to investigate whether the ErbB1 copy number affects EGFR expression, cell proliferation or cell migration by comparing two different cell lines.MethodsThe copies of ErbB1 gene was evaluated by FISH. Immunofluorescence and Western blotting were performed to determine location and expression of proteins mentioned in the present study. Proliferation was studied by flow cytometry and cell migration by wound healing assay and time lapse.ResultsWe investigated the activation and function of EGFR in the A549 and HK2 lung cancer cell lines, which contain 3 and 6 copies of ErbB1, respectively. The expression of EGFR was lower in the HK2 cell line. EGFR was activated after stimulation with EGF in both cell lines, but this activation did not promote differences in cellular proliferation when compared to control cells. Inhibiting EGFR with AG1478 did not modify cellular proliferation, confirming previous data. However, we observed morphological alterations, changes in microfilament organization and increased cell migration upon EGF stimulation. However, these effects did not seem to be consequence of an epithelial-mesenchymal transition.ConclusionEGFR expression did not appear to be associated to the ErbB1 gene copy number, and neither of these aspects appeared to affect cell proliferation. However, EGFR activation by EGF resulted in cell migration stimulation in both cell lines.
Highlights
Lung cancer is the most common type of cancer in men and women worldwide, and it is the main cause of cancerrelated death in the United States, Europe, Japan and China [1,2,3,4,5]
Cell culture The human non-small cell lung cancer A549 cell line was obtained from ATCC, and the HK2 cell line was established in our laboratory [25]
epidermal growth factor receptor (EGFR) activation by EGF did not induce cell proliferation Once we demonstrated that EGFR is phosphorylated by EGF exposure in A549 and HK2 cells, the step was to determine if the EGFR activation contribute to cell proliferation
Summary
Lung cancer is the most common type of cancer in men and women worldwide, and it is the main cause of cancerrelated death in the United States, Europe, Japan and China [1,2,3,4,5]. Lung cancer cells frequently manifest alterations in their molecular genetics, including changes in epidermal growth factor receptor (EGFR) expression. EGFR is a receptor encoded by the ErbB1 gene, which is located in the 7p12 region of chromosome 7. EGFR is activated by its ligands (EGF, TGF-α, amphiregulin, betacellulin, heparin or epiregulin) that bind to the extracellular domain, causing a conformational change and promoting homo- or hetero-dimerization. This results in the autophosphorylation of the tyrosine kinase domain, thereby initiating an intracellular signaling cascade [9]. ErbB1 encodes epidermal growth factor receptor (EGFR), a tyrosine kinase receptor, involved mainly in cell proliferation and survival. The aim of the present study was to investigate whether the ErbB1 copy number affects EGFR expression, cell proliferation or cell migration by comparing two different cell lines
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