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Rationale: Damage to airway epithelium is followed by deposition of extracellular matrix (ECM) and migration of adjacent epithelial cells. We have shown that epithelial cells from children with asthma fail to heal a wound in vitro.Objectives: To determine whether dysregulated ECM production by the epithelium plays a role in aberrant repair in asthma.Methods: Airway epithelial cells (AEC) from children with asthma (n = 36), healthy atopic control subjects (n = 23), and healthy nonatopic control subjects (n = 53) were investigated by microarray, gene expression and silencing, transcript regulation analysis, and ability to close mechanical wounds.Measurements and Main Results: Time to repair a mechanical wound in vitro by AEC from healthy and atopic children was not significantly different and both were faster than AEC from children with asthma. Microarray analysis revealed differential expression of multiple gene sets associated with repair and remodeling in asthmatic AEC. Fibronectin (FN) was the only ECM component whose expression was significantly lower in asthmatic AEC. Expression differences were verified by quantitative polymerase chain reaction and ELISA, and reduced FN expression persisted in asthmatic cells over passage. Silencing of FN expression in nonasthmatic AEC inhibited wound repair, whereas addition of FN to asthmatic AEC restored reparative capacity. Asthmatic AEC failed to synthesize FN in response to wounding or cytokine/growth factor stimulation. Exposure to 5', 2'deoxyazacytidine had no effect on FN expression and subsequent analysis of the FN promoter did not show evidence of DNA methylation.Conclusions: These data show that the reduced capacity of asthmatic epithelial cells to secrete FN is an important contributor to the dysregulated AEC repair observed in these cells. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Secreted phospholipase A 2 s (sPLA 2 s) regulate eicosanoid formation and have been implicated in asthma. Although sPLA 2 s function as enzymes, some of the sPLA 2 s bind with high affinity to a C-type lectin receptor, called PLA2R1, which has functions in both cellular signaling and clearance of sPLA 2 s. We sought to examine the expression of PLA2R1 in the airway epithelium of human subjects with asthma and the function of the murine Pla2r1 gene in a model of asthma. Expression of PLA2R1 in epithelial brushings was assessed in two distinct cohorts of children with asthma by microarray and quantitative PCR, and immunostaining for PLA2R1 was conducted on endobronchial tissue and epithelial brushings from adults with asthma. C57BL/129 mice deficient in Pla2r1 (Pla2r1 -/- ) were characterized in an ovalbumin (OVA) model of allergic asthma. PLA2R1 was differentially overexpressed in epithelial brushings of children with atopic asthma in both cohorts. Immunostaining for PLA2R1 in endobronchial tissue localized to submucosal glandular epithelium and columnar epithelial cells. After OVA sensitization and challenge, Pla2r1 -/- mice had increased airway hyperresponsiveness, as well as an increase in cellular trafficking of eosinophils to the peribronchial space and bronchoalveolar lavage fluid, and an increase in airway permeability. In addition, Pla2r1 -/- mice had more dendritic cells in the lung, higher levels of OVA-specific IgG, and increased production of both type-1 and type-2 cytokines by lung leukocytes. PLA2R1 is increased in the airway epithelium in asthma, and serves as a regulator of airway hyperresponsiveness, airway permeability, antigen sensitization, and airway inflammation.