Induction of epithelial-mesenchymal transition in primary airway epithelial cells from patients with asthma by transforming growth factor-beta1.

TitleInduction of epithelial-mesenchymal transition in primary airway epithelial cells from patients with asthma by transforming growth factor-beta1.
Publication TypeJournal Article
Year of Publication2009
AuthorsHackett, T-L, Warner, SMary, Stefanowicz, D, Shaheen, F, Pechkovsky, DV, Murray, LA, Argentieri, R, Kicic, A, Stick, SM, Bai, TR, Knight, DA
JournalAm J Respir Crit Care Med
Volume180
Issue2
Pagination122-33
Date Published2009 Jul 15
ISSN1535-4970
KeywordsAdolescent, Asthma, Case-Control Studies, Cell Culture Techniques, Cell Dedifferentiation, Child, Child, Preschool, Epithelial Cells, Extracellular Matrix Proteins, Female, Humans, Intercellular Signaling Peptides and Proteins, Male, Mesenchymal Stromal Cells, Recombinant Proteins, Signal Transduction, Transforming Growth Factor beta1, Young Adult
Abstract

RATIONALE: Airway remodeling in asthma is associated with the accumulation of fibroblasts, the primary cell responsible for synthesis and secretion of extracellular matrix proteins. The process by which the number of fibroblasts increases in asthma is poorly understood, but epithelial-mesenchymal transition (EMT) may play a significant role.OBJECTIVES: To evaluate whether EMT occurs in primary airway epithelial cells (AECs), the mechanisms involved, and if this process is altered in asthmatic AECs.METHODS: AECs were obtained from subjects with asthma (n = 8) and normal subjects without asthma (n = 10). Monolayer and air-liquid interface-AEC (ALI-AEC) cultures were treated with transforming growth factor (TGF)-beta1 (10 ng/ml) for 72 hours and assayed for mesenchymal and epithelial markers using quantitative polymerase chain reaction, confocal microscopy, and immunoblot. The involvement of BMP-7, Smad3, and MAPK-mediated signaling were also evaluated.MEASUREMENTS AND MAIN RESULTS: TGF-beta1-induced EMT in AEC monolayers derived from subjects with asthma and normal donors. EMT was characterized by changes in cell morphology, increased expression of mesenchymal markers EDA-fibronectin, vimentin, alpha-smooth muscle actin, and collagen-1, and loss of epithelial markers E-cadherin and zonular occludin-1. Inhibition of TGF-beta1-induced signaling with Smad3-inhibiting siRNA or TGF-beta1-neutralizing antibodies prevented and reversed EMT, respectively, whereas BMP-7 had no effect. In ALI-AEC cultures derived from normal subjects, EMT was confined to basally situated cells, whereas in asthmatic ALI-AEC cultures EMT was widespread throughout the epithelium.CONCLUSIONS: TGF-beta1 induces EMT in a Smad3-dependent manner in primary AECs. However, in asthmatic-derived ALI-AEC cultures, the number of cells undergoing EMT is greater. These findings support the hypothesis that epithelial repair in asthmatic airways is dysregulated.

DOI10.1164/rccm.200811-1730OC
Alternate JournalAm. J. Respir. Crit. Care Med.
PubMed ID19406982