Respiratory Syncytial Virus Infection Disrupts Monolayer Integrity and Function in Cystic Fibrosis Airway Cells

Michele Kong,Rhonda Szczesniak,John Clancy,Wayne Sullender,Patrick Maeng,Jeong Hong,Eric Sorscher

doi:10.3390/v5092260

Abstract

Background: Respiratory Syncytial Virus (RSV) infection is a common contributor to pulmonary symptoms in children with cystic fibrosis (CF). Here we examined RSV infection in immortalized bronchial epithelial cells (CFBE41o-) expressing wild-type (wt) or F508del cystic fibrosis transmembrane conductance regulator (CFTR), for monolayer integrity and RSV replication. Methods: CFBE41o- monolayers expressing wt or F508del CFTR were grown on permeable supports and inoculated with RSV A2 strain. Control experiments utilized UV-inactivated RSV and heat-killed RSV. Monolayer resistance and RSV production was monitored for up to six days post-infection. Results: Within 24 h, a progressive decrease in monolayer resistance was observed in RSV infected F508del CFBE41o- cells, while the monolayer integrity of RSV infected wt CFTR CFBE41o- cells remained stable. RSV replication was necessary to disrupt F508del CFBE41o- monolayers as UV-irradiated and heat killed RSV had no effect on monolayer integrity, with an earlier and much more pronounced peak in RSV titer noted in F508del relative to wt CFTR-expressing cells. RSV infection of wt CFBE41o- monolayers also resulted in blunting of CFTR response. Conclusions: These findings identify an enhanced sensitivity of CFBE41o- cells expressing F508del CFTR to RSV infection, replication and monolayer disruption independent of the cellular immune response, and provide a novel mechanism by which cystic fibrosis airway epithelia are susceptible to RSV-dependent injury.

Highlights

Cystic fibrosis (CF) is an autosomal recessive disorder, caused by mutations in the gene that encodes the cystic fibrosis transmembrane conductance regulator (CFTR) protein [1]
Recent studies demonstrate the importance of CFTR function in CF airway host defense, wound repair and airway remodeling [4,5], with CFTR dysfunction implicated in excessive lung inflammation, bacterial infection and colonization [6]
Despite the declining resistance from days 4–6 in the Respiratory Syncytial Virus (RSV) infected wt CFTR cells, monolayer resistance remains above the pre-RSV infection values at all time points

Summary

Introduction

Cystic fibrosis (CF) is an autosomal recessive disorder, caused by mutations in the gene that encodes the cystic fibrosis transmembrane conductance regulator (CFTR) protein [1]. Deletion of phenylalanine at position 508 (F508del) is the most common disease causing mutation, resulting in protein misfolding and endoplasmic reticulum degradation of the CFTR protein [2]. Epidemiologic studies have shown that pulmonary exacerbations caused by RSV disease, in children with CF results in frequent acute outpatient visits and hospitalizations. In CF children, RSV can cause significant clinical manifestations [7,8,9], and may predispose to early respiratory colonization with CF-specific bacterial pathogens. Respiratory Syncytial Virus (RSV) infection is a common contributor to pulmonary symptoms in children with cystic fibrosis (CF). We examined RSV infection in immortalized bronchial epithelial cells (CFBE41o-) expressing wild-type (wt) or F508del cystic fibrosis transmembrane conductance regulator (CFTR), for monolayer integrity and RSV replication. Results: Within 24 h, a progressive decrease in monolayer resistance was observed in RSV infected

Methods

Results

Conclusion