Roflumilast Inhibits Respiratory Syncytial Virus Infection in Human Differentiated Bronchial Epithelial Cells

Manuel Mata,Herman Tenor,Isidoro Martinez,Jose A Melero,Julio Cortijo,Steven M Varga

doi:10.1371/journal.pone.0069670

Abstract

Respiratory syncytial virus (RSV) causes acute exacerbations in COPD and asthma. RSV infects bronchial epithelial cells (HBE) that trigger RSV associated lung pathology. This study explores whether the phosphodiesterase 4 (PDE4) inhibitor Roflumilast N-oxide (RNO), alters RSV infection of well-differentiated HBE (WD-HBE) in vitro. WD-HBE were RSV infected in the presence or absence of RNO (0.1-100 nM). Viral infection (staining of F and G proteins, nucleoprotein RNA level), mRNA of ICAM-1, ciliated cell markers (digital high speed videomicroscopy, β-tubulin immunofluorescence, Foxj1 and Dnai2 mRNA), Goblet cells (PAS), mRNA of MUC5AC and CLCA1, mRNA and protein level of IL-13, IL-6, IL-8, TNFα, formation of H2O2 and the anti-oxidative armamentarium (mRNA of Nrf2, HO-1, GPx; total antioxidant capacity (TAC) were measured at day 10 or 15 post infection. RNO inhibited RSV infection of WD-HBE, prevented the loss of ciliated cells and markers, reduced the increase of MUC5AC and CLCA1 and inhibited the increase of IL-13, IL-6, IL-8, TNFα and ICAM-1. Additionally RNO reversed the reduction of Nrf2, HO-1 and GPx mRNA levels and consequently restored the TAC and reduced the H2O2 formation. RNO inhibits RSV infection of WD-HBE cultures and mitigates the cytopathological changes associated to this virus.

Highlights

Respiratory syncytial virus (RSV) is an enveloped negativesense single-stranded RNA virus of the Paramyxoviridae family
After RSV infection, we have found an increase of mRNA and protein release of IL-8, IL-6 and TNF-α in well-differentiated normal human bronchial epithelial cells (WD-HBE)
A key observation from the current study has been that RNO inhibits RSV infection in WD-HBE cultures which reproduce many aspects of normal human epithelium

Summary

Introduction

Respiratory syncytial virus (RSV) is an enveloped negativesense single-stranded RNA virus of the Paramyxoviridae family It is the main cause of severe lower respiratory tract infections (bronchiolitis and pneumonia) in infants and young children and it is a pathogen of considerable importance in the elderly and in immunocompromised adults [1,2]. RSV infection causes the release of numerous pro-inflammatory chemokines and cytokines from bronchial epithelial cells and entails an imbalance between an enhanced reactive oxygen species (ROS) production and a compromised antioxidant enzymatic armamentarium [10,11]. Given these numerous effects, bronchial epithelial cells are considered key players of RSV-induced lung diseases

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