TNF-α–mediated bronchial barrier disruption and regulation by src-family kinase activation

• Published in: Journal of allergy and clinical immunology Vol. 132; no. 3; pp. 665 - 675.e8
• Main Authors: Hardyman, Michelle A., PhD, Wilkinson, Emily, BSc, Martin, Emma, BMedSci, Jayasekera, Nivenka P., MD, Blume, Cornelia, PhD, Swindle, Emily J., PhD, Gozzard, Neil, PhD, Holgate, Stephen T., MD, DSc, Howarth, Peter H., MD, DM, Davies, Donna E., PhD, Collins, Jane E., PhD
• Format: Journal Article
• Language: English
• Published: New York, NY Elsevier Inc 01.09.2013; Elsevier
• Subjects: Adherens Junctions - drug effects; Adherens Junctions - metabolism; Airway; Allergy and Immunology; Asthma - metabolism; barrier; Biological and medical sciences; Bronchi - cytology; Bronchi - metabolism; bronchial; Cadherins - metabolism; Catenins - metabolism; Cells, Cultured; cytokines; Cytokines - metabolism; Delta Catenin; epithelial; Epithelial Cells - drug effects; Epithelial Cells - metabolism; Fundamental and applied biological sciences. Psychology; Fundamental immunology; Humans; Immunopathology; Indoles - pharmacology; Medical sciences; proMMP-9; Protein Kinase Inhibitors - pharmacology; Sarcoidosis. Granulomatous diseases of unproved etiology. Connective tissue diseases. Elastic tissue diseases. Vasculitis; src kinase; src-Family Kinases - antagonists & inhibitors; src-Family Kinases - metabolism; SU6656; Sulfonamides - pharmacology; Tight Junctions - drug effects; Tight Junctions - metabolism; TNF-α; Tumor Necrosis Factor-alpha - antagonists & inhibitors; Tumor Necrosis Factor-alpha - genetics; Tumor Necrosis Factor-alpha - metabolism; Fluorescein isothiocyanate; TER; Thymic stromal lymphoprotein; HRP; Src family kinase; Horseradish peroxidase; Zonula occluden; Madin-Darby canine kidney epithelial; Monoclonal antibody; Epithelial volt ohm meter; LDH; FITC; barrier; Human bronchial epithelial cell; Tissue inhibitor of metalloproteinase; SFK; cytokines; proMMP-9; MDCK; Transepithelial electrical resistance; MSD; TSLP; bronchial; epithelial; AJ; Lactate dehydrogenase; SU6656; Adherens junctions; Air-liquid interface; Electrochemiluminescence; Airway; ECL; EVOM; mAb; MMP; TNF-α; ZO; TJ; HBEC; Matrix metalloprotease; Tight junction; src kinase; Meso Scale Discovery; ALI; TIMP; Activation; Respiratory system; Respiratory tract; Regulation(control); Immunology; Family environment; Protein-tyrosine kinase; Tumor necrosis factor α; Protooncogene; Immunopathology; Enzyme; Family study; Transferases; Cytokine; Bronchus; Kinase; C-Onc gene
• ISSN: 0091-6749; 1097-6825
• DOI: 10.1016/j.jaci.2013.03.005

Background Because TNF-α is increased in severe asthma, we hypothesized that TNF-α contributes to barrier dysfunction and cell activation in bronchial epithelial cells. We further hypothesized that src-family kinase inhibition would improve barrier function in healthy cells in the presence of TNF-α and directly in cultures of severe asthmatic cells where the barrier is disrupted. Objectives We assessed the effect of TNF-α, with or without src-family kinase inhibitor SU6656, on barrier properties and cytokine release in differentiated human bronchial epithelial cultures. Further, we tested the effect of SU6656 on differentiated primary cultures from severe asthma. Methods Barrier properties of differentiated human bronchial epithelial air-liquid interface cultures from healthy subjects and subjects with severe asthma were assessed with transepithelial electrical resistance and fluorescent dextran passage. Proteins were detected by immunostaining or Western blot analysis and cytokines by immunoassay. Mechanisms were investigated with src kinase and other inhibitors. Results TNF-α lowered transepithelial electrical resistance and increased fluorescent dextran permeability, caused loss of occludin and claudins from tight junctions with redistribution of p120 catenin and E-cadherin from adherens junctions, and also increased endogenous TNF-α, IL-6, IL-1β, IL-8, thymic stromal lymphoprotein, and pro–matrix metalloprotease 9 release. SU6656 reduced TNF-α–mediated paracellular permeability changes, restored occludin, p120, and E-cadherin and lowered autocrine TNF-α release. Importantly, SU6656 improved the barrier properties of severe asthmatic air-liquid interface cultures. Redistribution of E-cadherin and p120 was observed in bronchial biopsies from severe asthmatic airways. Conclusions Inhibiting TNF-α or src kinases may be a therapeutic option to normalize barrier integrity and cytokine release in airway diseases associated with barrier dysfunction.