Rhinovirus Delays Cell Repolarization in a Model of Injured/Regenerating Human Airway Epithelium

Abstract

Rhinovirus (RV), which causes exacerbation in patients with chronic airway diseases, readily infects injured airway epithelium and has been reported to delay wound closure. In this study, we examined the effects of RV on cell repolarization and differentiation in a model of injured/regenerating airway epithelium (polarized, undifferentiated cells). RV causes only a transient barrier disruption in a model of normal (mucociliary-differentiated) airway epithelium. However, in the injury/regeneration model, RV prolongs barrier dysfunction and alters the differentiation of cells. The prolonged barrier dysfunction caused by RV was not a result of excessive cell death but was instead associated with epithelial-to-mesenchymal transition (EMT)-like features, such as reduced expression of the apicolateral junction and polarity complex proteins, E-cadherin, occludin, ZO-1, claudins 1 and 4, and Crumbs3 and increased expression of vimentin, a mesenchymal cell marker. The expression of Snail, a transcriptional repressor of tight and adherence junctions, was also up-regulated in RV-infected injured/regenerating airway epithelium, and inhibition of Snail reversed RV-induced EMT-like features. In addition, compared with sham-infected cells, the RV-infected injured/regenerating airway epithelium showed more goblet cells and fewer ciliated cells. Inhibition of epithelial growth factor receptor promoted repolarization of cells by inhibiting Snail and enhancing expression of E-cadherin, occludin, and Crumbs3 proteins, reduced the number of goblet cells, and increased the number of ciliated cells. Together, these results suggest that RV not only disrupts barrier function, but also interferes with normal renewal of injured/regenerating airway epithelium by inducing EMT-like features and subsequent goblet cell hyperplasia.Keywords: barrier function; epithelial-to-mesenchymal transition; Crumbs polarity complex; epithelial growth factor receptor; goblet cell hyperplasia Clinical RelevanceTo the best of our knowledge, this is the first study to demonstrate that rhinovirus delays cell repolarization in a model of injured/regenerating, but not normal, airway epithelium by inducing epithelial-to-mesenchymal transition-like features. Furthermore, rhinovirus reduces ciliated cell differentiation and promotes goblet-cell differentiation and mucin gene expression. These changes depended partially on persistent EGFR activation induced by rhinovirus. Such changes during regeneration can lead to airway epithelium with impaired barrier and mucociliary clearance functions.Airway epithelium that lines the respiratory tract acts as a physical barrier between the external environment and the lung and plays an important role in maintaining tissue homeostasis. Paracellular permeability of airway epithelium is maintained through the co-operation of two functionally distinct structural components: tight and adherence junctions located in the apicolateral membranes. Tight junctions regulate the transport of solutes and ions across airway epithelium...

NOTCH3 contributes to rhinovirus-induced goblet cell hyperplasia in COPD airway epithelial cells

“…Since RV activates EGFR and induces acute expression of MUC5AC in normal epithelial cells,34 35 and GCH in repairing airway epithelial cell cultures,15 we reasoned that RV-induced EGFR may contribute to the observed GCH in COPD. To our surprise, RV did not increase EGFR phosphorylation in COPD cells.…”

“…Mucociliary-differentiated cell cultures were infected apically with 10 µL of phosphate buffered saline (PBS) containing RV16 at 1 multiplicity of infection (MOI) or equivalent volume of sham and incubated for 15 days as described previously 15. Medium in the basolateral chamber was changed every other day.…”

“…Number of goblet cells in cell cultures was determined as described previously 15. Briefly, after relevant treatment, apical surface of the cell cultures was washed with 0.15% sodium bicarbonate in PBS, fixed in ice cold methanol and then immunostained with antibody to human tracheobronchial mucins30 and imaged under confocal microscopy to detect goblet cells.…”

“…Chronic EGFR activation by Sendai virus induces GCH in well-differentiated airway epithelial cells by promoting survival and transdifferentiation of ciliated to goblet cells in concert with IL-13 14. Recently, we demonstrated that RV promotes goblet cell differentiation in a model of injured but not in normal airway epithelium via persistent activation of EGFR 15. Since EGFR activity is already enhanced in COPD,16 17 RV may further activate EGFR to drive GCH independently or in concert with IL-13.…”

Viral–bacterial interactions in the respiratory tract

“…Sajjan et al (2008) showed that infection of polarized airway epithelial cells with HRV can lead to redistribution of the tight junction protein zona occludens 1 (ZO-1) from the membrane to the cytoplasm. Furthermore, HRV was shown to impair the repolarization of airway epithelium regenerating after injury (Faris et al, 2016). Integrity of the epithelial barrier likewise is reduced after RSV infection (Kilani et al, 2004).…”