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Rat respiratory coronavirus infection: replication in airway and alveolar epithelial cells and the innate immune response.

ABSTRACT: The rat coronavirus sialodacryoadenitis virus (SDAV) causes respiratory infection and provides a system for investigating respiratory coronaviruses in a natural host. A viral suspension in the form of a microspray aerosol was delivered by intratracheal instillation into the distal lung of 6-8-week-old Fischer 344 rats. SDAV inoculation produced a 7 % body weight loss over a 5 day period that was followed by recovery over the next 7 days. SDAV caused focal lesions in the lung, which were most severe on day 4 post-inoculation (p.i.). Immunofluorescent staining showed that four cell types supported SDAV virus replication in the lower respiratory tract, namely Clara cells, ciliated cells in the bronchial airway and alveolar type I and type II cells in the lung parenchyma. In bronchial alveolar lavage fluid (BALF) a neutrophil influx increased the population of neutrophils to 45 % compared with 6 % of the cells in control samples on day 2 after mock inoculation. Virus infection induced an increase in surfactant protein SP-D levels in BALF of infected rats on days 4 and 8 p.i. that subsided by day 12. The concentrations of chemokines MCP-1, LIX and CINC-1 in BALF increased on day 4 p.i., but returned to control levels by day 8. Intratracheal instillation of rats with SDAV coronavirus caused an acute, self-limited infection that is a useful model for studying the early events of the innate immune response to respiratory coronavirus infections in lungs of the natural virus host.

SUBMITTER: Funk CJ

PROVIDER: S-EPMC2887555 | biostudies-literature | 2009 Dec

REPOSITORIES: biostudies-literature

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Rat respiratory coronavirus infection: replication in airway and alveolar epithelial cells and the innate immune response.

Funk C Joel CJ Manzer Rizwan R Miura Tanya A TA Groshong Steve D SD Ito Yoko Y Travanty Emily A EA Leete Jennifer J Holmes Kathryn V KV Mason Robert J RJ

The Journal of general virology 20090909 Pt 12

The rat coronavirus sialodacryoadenitis virus (SDAV) causes respiratory infection and provides a system for investigating respiratory coronaviruses in a natural host. A viral suspension in the form of a microspray aerosol was delivered by intratracheal instillation into the distal lung of 6-8-week-old Fischer 344 rats. SDAV inoculation produced a 7 % body weight loss over a 5 day period that was followed by recovery over the next 7 days. SDAV caused focal lesions in the lung, which were most sev ...[more]

PMID: 19741068

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Project description:UnlabelledInfluenza is the cause of significant morbidity and mortality in pediatric populations. The contribution of pulmonary host defense mechanisms to viral respiratory infection susceptibility in very young children is poorly understood. As a surrogate to compare mucosal immune responses of infant and adult lungs, rhesus monkey primary airway epithelial cell cultures were infected with pandemic influenza A/H1N1 virus in vitro. Virus replication, cytokine secretion, cell viability, and type I interferon (IFN) pathway PCR array profiles were evaluated for both infant and adult cultures. In comparison with adult cultures, infant cultures showed significantly increased levels of H1N1 replication, reduced alpha interferon (IFN-?) protein synthesis, and no difference in cell death following infection. Age-dependent differences in expression levels of multiple genes associated with the type I IFN pathway were observed in H1N1-infected cultures. To investigate the pulmonary and systemic responses to H1N1 infection in early life, infant monkeys were inoculated with H1N1 by upper airway administration. Animals were monitored for virus and parameters of inflammation over a 14-day period. High H1N1 titers were recovered from airways at day 1, with viral RNA remaining detectable until day 9 postinfection. Despite viral clearance, bronchiolitis and alveolitis persisted at day 14 postinfection; histopathological analysis revealed alveolar septal thickening and intermittent type II pneumocyte hyperplasia. Our overall findings are consistent with the known susceptibility of pediatric populations to respiratory virus infection and suggest that intrinsic developmental differences in airway epithelial cell immune function may contribute to the limited efficacy of host defense during early childhood.ImportanceTo the best of our knowledge, this study represents the first report of intrinsic developmental differences in infant airway epithelial cells that may contribute to the increased susceptibility of the host to respiratory virus infections. Despite the global burden of influenza, there are currently no vaccine formulations approved for children <6 months of age. Given the challenges of conducting experimental studies involving pediatric patients, rhesus monkeys are an ideal laboratory animal model to investigate the maturation of pulmonary mucosal immune mechanisms during early life because they are most similar to those of humans with regard to postnatal maturation of the lung structure and the immune system. Thus, our findings are highly relevant to translational medicine, and these data may ultimately lead to novel approaches that enhance airway immunity in very young children.

Project description:ADP-ribosylation is a common posttranslational modification that may have antiviral properties and impact innate immunity. To regulate this activity, macrodomain proteins enzymatically remove covalently attached ADP-ribose from protein targets. All members of the Coronavirinae, a subfamily of positive-sense RNA viruses, contain a highly conserved macrodomain within nonstructural protein 3 (nsp3). However, its function or targets during infection remain unknown. We identified several macrodomain mutations that greatly reduced nsp3's de-ADP-ribosylation activity in vitro Next, we created recombinant severe acute respiratory syndrome coronavirus (SARS-CoV) strains with these mutations. These mutations led to virus attenuation and a modest reduction of viral loads in infected mice, despite normal replication in cell culture. Further, macrodomain mutant virus elicited an early, enhanced interferon (IFN), interferon-stimulated gene (ISG), and proinflammatory cytokine response in mice and in a human bronchial epithelial cell line. Using a coinfection assay, we found that inclusion of mutant virus in the inoculum protected mice from an otherwise lethal SARS-CoV infection without reducing virus loads, indicating that the changes in innate immune response were physiologically significant. In conclusion, we have established a novel function for the SARS-CoV macrodomain that implicates ADP-ribose in the regulation of the innate immune response and helps to demonstrate why this domain is conserved in CoVs. IMPORTANCE:The macrodomain is a ubiquitous structural domain that removes ADP-ribose from proteins, reversing the activity of ADP-ribosyltransferases. All coronaviruses contain a macrodomain, suggesting that ADP-ribosylation impacts coronavirus infection. However, its function during infection remains unknown. Here, we found that the macrodomain is an important virulence factor for a highly pathogenic human CoV, SARS-CoV. Viruses with macrodomain mutations that abrogate its ability to remove ADP-ribose from protein were unable to cause lethal disease in mice. Importantly, the SARS-CoV macrodomain suppressed the innate immune response during infection. Our data suggest that an early innate immune response can protect mice from lethal disease. Understanding the mechanism used by this enzyme to promote disease will open up novel avenues for coronavirus therapies and give further insight into the role of macrodomains in viral pathogenesis.

Dataset Information

Rat respiratory coronavirus infection: replication in airway and alveolar epithelial cells and the innate immune response.

Publications

Rat respiratory coronavirus infection: replication in airway and alveolar epithelial cells and the innate immune response.

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