Respiratory syncytial virus increases IL-8 gene expression and protein release in A549 cells

1995 ◽  
Vol 269 (6) ◽  
pp. L865-L872 ◽  
Author(s):  
M. A. Fiedler ◽  
K. Wernke-Dollries ◽  
J. M. Stark
Keyword(s):  
Gene Expression ◽  
Respiratory Syncytial Virus ◽  
Viral Replication ◽  
Mrna Expression ◽  
A549 Cells ◽  
Protein Release ◽  
Tnf Alpha ◽  
Epithelial Cell Line ◽  
Airway Epithelial ◽  
Syncytial Virus

The mechanism of respiratory syncytial virus (RSV)-induced inflammation in the airways of infants and children is not fully understood. We hypothesized that RSV directly induces interleukin (IL)-8 gene expression in airway epithelial cells, independent of IL-1 beta and tumor necrosis factor-alpha (TNF-alpha) production. Exposure of A549 cells (an airway epithelial cell line) to RSV resulted in increased IL-8 mRNA expression and IL-8 protein release from the cells as early as 2 h after treatment. Neither IL-1 beta nor TNF-alpha (mRNA or protein) were detected. Viral replication was not necessary for the effects of RSV on IL-8 mRNA expression and protein release early in the infectious process. However, sustained levels of increased IL-8 production required RSV replication. A dose-response relationship was observed between the multiplicity of infection and IL-8 production with both active and nonreplicative RSV at the 2-h time point. Both active RSV and nonreplicative RSV increased the transcriptional activity of the 1.6-kb 5' flanking region of the IL-8 gene. Neither active RSV nor nonreplicative RSV increased the stability of the IL-8 mRNA in A549 cells. We conclude that RSV increases IL-8 gene expression in A549 cells in a biphasic pattern independent of viral replication early (2 h) but dependent on viral replication late (24 h).

Mechanism of RSV-induced IL-8 gene expression in A549 cells before viral replication

1996 ◽  
Vol 271 (6) ◽  
pp. L963-L971 ◽  
Author(s):  
M. A. Fiedler ◽  
K. Wernke-Dollries ◽  
J. M. Stark
Keyword(s):  
Gene Expression ◽  
Transcription Factor ◽  
Viral Replication ◽  
Transcriptional Activation ◽  
Mutational Analysis ◽  
A549 Cells ◽  
Nf Kappa B ◽  
Mobility Shift ◽  
Syncytial Virus ◽  
Time Point

Previous studies demonstrated that respiratory syncytial virus (RSV) infection of A549 cells induced interleukin (IL)-8 gene expression and protein release from the cells as early as 2 h after treatment [M. A. Fiedler, K. Wernke-Dollries, and J. M. Stark. Am. J. Physiol. 269 (Lung Cell. Mol. Physiol. 13): L865-L872, 1995; J. G. Mastronarde, M. M. Monick, and G. W. Hunninghake. Am. J. Respir. Cell Mol. Biol. 13: 237-244, 1995]. Furthermore, the effects of RSV at the 2-h time point were not dependent on viral replication. The studies reported here were designed to test the hypothesis that active and inactive RSV induce IL-8 gene expression in A549 cells at the 2-h time point by a mechanism dependent on the activation of the nuclear transcription factor NF-kappa B Northern blot analysis indicated that IL-8 gene expression occurred independent of protein synthesis 2 h after A549 cells were treated with RSV. Analysis of nuclear extracts from RSV-treated A549 cells by electrophoretic mobility shift assays demonstrated that NF-kappa B was activated as early as 15 min after RSV was added to the cells and remained activated for at least 90 min. In contrast, baseline levels of NF-IL-6 and activator protein-1 (AP-1) did not change over this period of time. Deoxyribonuclease footprint analysis of a portion of the 5'-flanking region of the IL-8 gene demonstrated two potential regions for transcription factor binding, which corresponded to the potential AP-1 binding site, and potential NF-IL-6 and NF-kappa B binding sites. Mutational analysis of the 200-bp 5'-untranslated region of the IL-8 gene demonstrated that activation of NF-kappa B and NF-IL-6 were required for RSV-induced transcriptional activation of the IL-8 gene.

scholarly journals The Interferon Type I/III Response to Respiratory Syncytial Virus Infection in Airway Epithelial Cells Can Be Attenuated or Amplified by Antiviral Treatment

2015 ◽  
Vol 90 (4) ◽  
pp. 1705-1717 ◽  
Author(s):  
K. M. McCutcheon ◽  
R. Jordan ◽  
M. E. Mawhorter ◽  
S. L. Noton ◽  
J. G. Powers ◽  
...  
Keyword(s):  
Life Cycle ◽  
Respiratory Syncytial Virus ◽  
Epithelial Cells ◽  
Viral Replication ◽  
Airway Epithelial Cells ◽  
Therapeutic Interventions ◽  
Airway Epithelial ◽  
Antiviral Activities ◽  
Syncytial Virus ◽  
Interferon Responses

ABSTRACTHuman respiratory syncytial virus (RSV) is a single-stranded RNA virus that causes acute, and occasionally fatal, lower respiratory illness in young infants, the elderly, and immunocompromised patients. Therapeutic interventions able to cut short viral replication and quickly return the airways to normal function are needed. An understanding of antiviral activities and their effects on host defense mechanisms is important for the design of safe and effective therapy. We targeted functionally and temporally distinct steps within the viral life cycle using small-molecule RSV inhibitors and studied their antiviral activities and their effects on innate interferon responses of airway epithelial cellsin vitro. Antivirals acting upstream of RSV polymerase activity (i.e., compounds targeting the fusion protein or the nucleoprotein) reduced viral load immediately postinfection and partially attenuated interferon responses. In contrast, antivirals directed to the RSV polymerase demonstrated activity throughout the viral replication cycle and specifically modulated the RIG-I/mitochondrial antiviral signaling protein (MAVS)/TBK1/IRF3/interferon-stimulated gene (ISG) axis, causing either an upregulation or a downregulation of interferon responses, depending on the mechanism of polymerase inhibition. Notably, polymerase inhibition leading to the accumulation of abortive RNA products correlated with the amplification of interferon-stimulated genes to up to 10 times above normal infection levels. Understanding how antiviral activities and their modulation of innate immunity may affect recovery from RSV infection will help guide the development of safe and effective therapies.IMPORTANCERSV circulates seasonally, causing acute lower respiratory disease. Therapeutic interventions with efficacy throughout the viral replication cycle, rapid viral clearance, and prevention of potentially harmful inflammatory responses are desirable. Compounds targeting the RSV polymerase inhibited virus replication late in the viral life cycle and, depending on the functional domain targeted, either attenuated or amplified RIG-I and downstream interferon pathways in infected cells. These data will help guide the development of safe and effective therapies by providing new molecular evidence that the mechanism of inhibition by an antiviral compound can directly impact innate antiviral immune responses in the airway epithelium.

scholarly journals Viral N6-methyladenosine upregulates replication and pathogenesis of human respiratory syncytial virus

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Miaoge Xue ◽  
Boxuan Simen Zhao ◽  
Zijie Zhang ◽  
Mijia Lu ◽  
Olivia Harder ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Rational Design ◽  
A549 Cells ◽  
Human Respiratory Syncytial Virus ◽  
Gene Transcript ◽  
Airway Epithelial ◽  
Live Attenuated Vaccine ◽  
Epithelial Cultures ◽  
Cotton Rats ◽  
Syncytial Virus

Abstract N6-methyladenosine (m6A) is the most prevalent internal modification of mRNAs in most eukaryotes. Here we show that RNAs of human respiratory syncytial virus (RSV) are modified by m6A within discreet regions and that these modifications enhance viral replication and pathogenesis. Knockdown of m6A methyltransferases decreases RSV replication and gene expression whereas knockdown of m6A demethylases has the opposite effect. The G gene transcript contains the most m6A modifications. Recombinant RSV variants expressing G transcripts that lack particular clusters of m6A display reduced replication in A549 cells, primary well differentiated human airway epithelial cultures, and respiratory tracts of cotton rats. One of the m6A-deficient variants is highly attenuated yet retains high immunogenicity in cotton rats. Collectively, our results demonstrate that viral m6A methylation upregulates RSV replication and pathogenesis and identify viral m6A methylation as a target for rational design of live attenuated vaccine candidates for RSV and perhaps other pneumoviruses.

scholarly journals Oxidant Tone Regulates RANTES Gene Expression in Airway Epithelial Cells Infected with Respiratory Syncytial Virus

2001 ◽  
Vol 276 (23) ◽  
pp. 19715-19722 ◽  
Author(s):  
Antonella Casola ◽  
Nathalie Burger ◽  
Tianshuang Liu ◽  
Mohammad Jamaluddin ◽  
Allan R. Brasier ◽  
...  
Keyword(s):  
Gene Expression ◽  
Respiratory Syncytial Virus ◽  
Epithelial Cells ◽  
Airway Epithelial Cells ◽  
Airway Epithelial ◽  
Syncytial Virus

Amiloride inhibits cytokine production in epithelium infected with respiratory syncytial virus

1996 ◽  
Vol 271 (2) ◽  
pp. L201-L207 ◽  
Author(s):  
J. G. Mastronarde ◽  
M. M. Monick ◽  
T. J. Gross ◽  
G. W. Hunninghake
Keyword(s):  
Respiratory Syncytial Virus ◽  
Airway Epithelium ◽  
Cytokine Production ◽  
A549 Cells ◽  
Protein Release ◽  
Respiratory Pathogen ◽  
Mrna Induction ◽  
Limited Effectiveness ◽  
Syncytial Virus ◽  
Few Data

Respiratory syncytial virus (RSV) is an important respiratory pathogen in infants and children. RSV preferentially infects airway epithelium and causes local production of inflammatory cytokines. Ribavirin, the only specific agent available for treatment of RSV infection, has limited effectiveness. There are few data regarding the ability of drugs to modulate the inflammatory response of epithelium infected with RSV. This study evaluated the effect of amiloride and ribavirin on cytokine production by RSV-infected epithelium. We observed a dose-dependent reduction in interleukin (IL)-8 protein release with both amiloride and ribavirin in RSV-infected A549 epithelial cells. Peak effects were observed at concentrations of 200 microM amiloride and 60 microM ribavirin. Both amiloride and ribavirin inhibited IL-8 mRNA induction. Pretreatment with either agent was not required to inhibit IL-8 release. Both drugs also inhibited IL-6 release. However, unlike ribavirin, amiloride did not inhibit viral replication or infection. Amiloride also inhibited IL-8 release from A549 cells stimulated with IL-1 or tumor necrosis factor. Amiloride similarly inhibited IL-8 protein release from primary human airway epithelium infected with RSV. These data demonstrate that both amiloride and ribavirin inhibit cytokine production in RSV-infected airway epithelium. These results suggest amiloride, as well as ribavirin, may be useful as a therapeutic agent in RSV infections.

scholarly journals Incomplete Regulation of NF-κB by IκBα during Respiratory Syncytial Virus Infection in A549 Cells

1999 ◽  
Vol 73 (5) ◽  
pp. 4502-4507 ◽  
Author(s):  
Michael A. Fiedler ◽  
Kara Wernke-Dollries
Keyword(s):  
Respiratory Syncytial Virus ◽  
Epithelial Cells ◽  
A549 Cells ◽  
Inhibitory Effect ◽  
Airway Epithelial Cells ◽  
Airway Epithelial ◽  
Necrosis Factor Alpha ◽  
Tnf Α ◽  
Rsv Infection ◽  
Syncytial Virus

ABSTRACT Respiratory syncytial virus (RSV) infection of airway epithelial cells results in persistent NF-κB activation and NF-κB-mediated interleukin-8 production. Previous studies in airway epithelial cells demonstrated that tumor necrosis factor alpha (TNF-α)-induced NF-κB activation is transient due to regulation by IκBα. However, during RSV infection, IκBα has only a partial inhibitory effect on NF-κB activation. Studies presented here demonstrate that neither increased IκBα production which occurs as a result of RSV-induced NF-κB activation nor inhibition of proteasome-mediated IκBα degradation results in a reversal of RSV-induced NF-κB activation. Thus, while manipulation of IκBα results in reversal of TNF-α-induced NF-κB activation, manipulation of IκBα does not result in a reversal of RSV-induced NF-κB activation.

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