Host DNA released by NETosis in neutrophils exposed to seasonal H1N1 and highly pathogenic H5N1 influenza viruses

Abstract Background Neutrophil (Nϕ) is of the most abundant number in human immune system. During acute influenza virus infection, Nϕs are already active in the early phase of inflammation-a time in which clinical biopsy or autopsy material is not readily available. However, the role of Nϕ in virus infection is not well understood. Here, we studied the role of Nϕ in host defense during influenza A virus infection, specifically assessing if it contributes to the differential pathogenesis in H5N1 disease. Methods Nϕs were freshly isolated from healthy volunteers and subjected to direct influenza H1N1 and H5N1 virus infection in vitro . The ability of the naïve Nϕs to infiltrate from the basolateral to the apical phase of the influenza virus infected alveolar epithelium was assessed. The viral replication, innate immune responses and Neutrophil extracellular trap (NET) formation of Nϕs upon influenza virus infection were evaluated. Results Our results demonstrated that influenza virus infected alveolar epithelium allowed more Nϕs transmigration. Significantly more Nϕs migrated across the H5N1 influenza virus infected the epithelium than the counterpart infected by the seasonal influenza H1N1 virus infected. Nϕs were equally susceptible to H5N1 and H1N1 virus infection with similar viral gene transcription. Productive replication was observed in H5N1 infected Nϕs. Both H5N1 and H1N1 infected Nϕs induced cytokines and chemokines including TNF-α, IFN-β, CXCL10, MIP-1α and IL-8. This inferred a more intense inflammatory response posed by H5N1 than H1N1 virus. Strikingly, NADPH oxidase-independent NET formation was observed in H1N1 infected Nϕs at 6 hpi while no NET formation was observed upon H5N1 infection. Conclusion Our data is the first to demonstrate that NET formation is abrogated in H5N1 influenza virus infection. Its contribution to the differential severity of H5N1 disease requires further investigation.

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Host DNA released by NETosis in neutrophils exposed to seasonal H1N1 and highly pathogenic H5N1 influenza viruses

10.21203/rs.2.24164/v2 ◽

2020 ◽

Author(s):

Louisa L.Y. Chan ◽

John M. Nicholls ◽

J.S. Malik Peiris ◽

Yu Lung Lau ◽

Michael C.W. Chan ◽

...

Keyword(s):

Influenza Virus ◽

Virus Infection ◽

Gene Transcription ◽

H1n1 Virus ◽

Influenza Virus Infection ◽

Alveolar Epithelium ◽

H5n1 Influenza Virus ◽

H5n1 Influenza ◽

Influenza H1n1

Abstract Background: Neutrophil is of the most abundant number in human immune system. During acute influenza virus infection, neutrophils are already active in the early phase of inflammation-a time in which clinical biopsy or autopsy material is not readily available. However, the role of neutrophil in virus infection is not well understood. Here, we studied the role of neutrophil in host defense during influenza A virus infection, specifically assessing if it contributes to the differential pathogenesis in H5N1 disease. Methods: Neutrophils were freshly isolated from healthy volunteers and subjected to direct influenza H1N1 and H5N1 virus infection in vitro. The ability of the naïve neutrophils to infiltrate from the basolateral to the apical phase of the influenza virus infected alveolar epithelium was assessed. The viral replication, innate immune responses and Neutrophil extracellular trap (NET) formation of neutrophils upon influenza virus infection were evaluated. Results: Our results demonstrated that influenza virus infected alveolar epithelium allowed more neutrophils transmigration. Significantly more neutrophils migrated across the H5N1 influenza virus infected the epithelium than the counterpart infected by the seasonal influenza H1N1 virus infected. Neutrophils were equally susceptible to H5N1 and H1N1 virus infection with similar viral gene transcription. Productive replication was observed in H5N1 infected neutrophils. H5N1 induced higher cytokine and chemokine gene transcription than H1N1 infected neutrophils, including TNF-α, IFN-β, CXCL10, MIP-1α and IL-8. This inferred a more intense inflammatory response posed by H5N1 than H1N1 virus. Strikingly, NADPH oxidase-independent NET formation was only observed in H1N1 infected neutrophils at 6 hpi while no NET formation was observed upon H5N1 infection.Conclusion: Our data is the first to demonstrate that NET formation is abrogated in H5N1 influenza virus infection and might contribute to the severity of H5N1 disease.

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Host DNA released by NETosis in neutrophils exposed to seasonal H1N1 and highly pathogenic H5N1 influenza viruses

Respiratory Research ◽

10.1186/s12931-020-01425-w ◽

2020 ◽

Vol 21 (1) ◽

Cited By ~ 1

Author(s):

Louisa L. Y. Chan ◽

John M. Nicholls ◽

J. S. Malik Peiris ◽

Yu Lung Lau ◽

Michael C. W. Chan ◽

...

Keyword(s):

Influenza Virus ◽

Virus Infection ◽

Gene Transcription ◽

H1n1 Virus ◽

Influenza Virus Infection ◽

Alveolar Epithelium ◽

H5n1 Influenza Virus ◽

H5n1 Influenza ◽

Influenza H1n1

Abstract Background Neutrophil is of the most abundant number in human immune system. During acute influenza virus infection, neutrophils are already active in the early phase of inflammation - a time in which clinical biopsy or autopsy material is not readily available. However, the role of neutrophil in virus infection is not well understood. Here, we studied the role of neutrophil in host defense during influenza A virus infection, specifically assessing if it contributes to the differential pathogenesis in H5N1 disease. Methods Neutrophils were freshly isolated from healthy volunteers and subjected to direct influenza H1N1 and H5N1 virus infection in vitro. The ability of the naïve neutrophils to infiltrate from the basolateral to the apical phase of the influenza virus infected alveolar epithelium was assessed. The viral replication, innate immune responses and Neutrophil extracellular trap (NET) formation of neutrophils upon influenza virus infection were evaluated. Results Our results demonstrated that influenza virus infected alveolar epithelium allowed neutrophil transmigration. Significantly more neutrophils migrated across the H5N1 influenza virus infected the epithelium than the counterpart infected by the seasonal influenza H1N1 virus infected. Neutrophils were equally susceptible to H5N1 and H1N1 virus infection with similar viral gene transcription. Productive replication was observed in H5N1 infected neutrophils. H5N1 induced higher cytokine and chemokine gene transcription than H1N1 infected neutrophils, including TNF-α, IFN-β, CXCL10, MIP-1α and IL-8. This inferred a more intense inflammatory response posed by H5N1 than H1N1 virus. Strikingly, NADPH oxidase-independent NET formation was only observed in H1N1 infected neutrophils at 6 hpi while no NET formation was observed upon H5N1 infection. Conclusion Our data is the first to demonstrate that NET formation is abrogated in H5N1 influenza virus infection and might contribute to the severity of H5N1 disease.

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Cross-Reactive, Cell-Mediated Immunity and Protection of Chickens from Lethal H5N1 Influenza Virus Infection in Hong Kong Poultry Markets

Journal of Virology ◽

10.1128/jvi.75.6.2516-2525.2001 ◽

2001 ◽

Vol 75 (6) ◽

pp. 2516-2525 ◽

Cited By ~ 161

Author(s):

Sang Heui Seo ◽

Robert G. Webster

Keyword(s):

T Cells ◽

Influenza Virus ◽

Hong Kong ◽

Virus Infection ◽

Cellular Immunity ◽

Influenza Virus Infection ◽

Influenza Viruses ◽

H5n1 Influenza Virus ◽

H9n2 Influenza Virus ◽

H5n1 Influenza

ABSTRACT In 1997, avian H5N1 influenza virus transmitted from chickens to humans resulted in 18 confirmed infections. Despite harboring lethal H5N1 influenza viruses, most chickens in the Hong Kong poultry markets showed no disease signs. At this time, H9N2 influenza viruses were cocirculating in the markets. We investigated the role of H9N2 influenza viruses in protecting chickens from lethal H5N1 influenza virus infections. Sera from chickens infected with an H9N2 influenza virus did not cross-react with an H5N1 influenza virus in neutralization or hemagglutination inhibition assays. Most chickens primed with an H9N2 influenza virus 3 to 70 days earlier survived the lethal challenge of an H5N1 influenza virus, but infected birds shed H5N1 influenza virus in their feces. Adoptive transfer of T lymphocytes or CD8+ T cells from inbred chickens (B2/B2) infected with an H9N2 influenza virus to naive inbred chickens (B2/B2) protected them from lethal H5N1 influenza virus. In vitro cytotoxicity assays showed that T lymphocytes or CD8+ T cells from chickens infected with an H9N2 influenza virus recognized target cells infected with either an H5N1 or H9N2 influenza virus in a dose-dependent manner. Our findings indicate that cross-reactive cellular immunity induced by H9N2 influenza viruses protected chickens from lethal infection with H5N1 influenza viruses in the Hong Kong markets in 1997 but permitted virus shedding in the feces. Our findings are the first to suggest that cross-reactive cellular immunity can change the outcome of avian influenza virus infection in birds in live markets and create a situation for the perpetuation of H5N1 influenza viruses.

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Role of Hypercytokinemia in NF-κB p50-Deficient Mice after H5N1 Influenza A Virus Infection

Journal of Virology ◽

10.1128/jvi.01071-08 ◽

2008 ◽

Vol 82 (22) ◽

pp. 11461-11466 ◽

Cited By ~ 36

Author(s):

Karoline Droebner ◽

Sarah Julia Reiling ◽

Oliver Planz

Keyword(s):

Influenza Virus ◽

Virus Infection ◽

Transgenic Mouse ◽

Proinflammatory Cytokines ◽

Influenza A ◽

Influenza Virus Infection ◽

H5n1 Influenza Virus ◽

Cytokines And Chemokines ◽

H5n1 Influenza

ABSTRACT During H5N1 influenza virus infection, proinflammatory cytokines are markedly elevated in the lungs of infected hosts. The significance of this dysregulated cytokine response in H5N1-mediated pathogenesis remains to be determined. To investigate the influence of hypercytokinemia, or “cytokine storm,” a transgenic mouse technology was used. The classical NF-κB pathway regulates the induction of most proinflammatory cytokines. Deletion of the p50 subunit leads to a markedly reduced expression of the NF-κB-regulated cytokines and chemokines. Here we show that H5N1 influenza virus infection of this transgenic mouse model resulted in a lack of hypercytokinemia but not in altered pathogenesis.

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A critical role of IL-17 in modulating the B-cell response during H5N1 influenza virus infection

Cellular and Molecular Immunology ◽

10.1038/cmi.2011.38 ◽

2011 ◽

Vol 8 (6) ◽

pp. 462-468 ◽

Cited By ~ 43

Author(s):

Xiaohui Wang ◽

Chris CS Chan ◽

Min Yang ◽

Jun Deng ◽

Vincent KM Poon ◽

...

Keyword(s):

Influenza Virus ◽

Virus Infection ◽

B Cell ◽

Cell Response ◽

Critical Role ◽

Influenza Virus Infection ◽

H5n1 Influenza Virus ◽

H5n1 Influenza ◽

B Cell Response

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Next Generation of Computationally Optimized Broadly Reactive HA Vaccines Elicited Cross-Reactive Immune Responses and Provided Protection against H1N1 Virus Infection

Vaccines ◽

10.3390/vaccines9070793 ◽

2021 ◽

Vol 9 (7) ◽

pp. 793

Author(s):

Ying Huang ◽

Monique S. França ◽

James D. Allen ◽

Hua Shi ◽

Ted M. Ross

Keyword(s):

Influenza Virus ◽

Virus Infection ◽

Immune Responses ◽

H1n1 Virus ◽

Influenza Virus Infection ◽

H1n1 Influenza ◽

Influenza Viruses ◽

Next Generation ◽

Wild Type ◽

Influenza A H1n1

Vaccination is the best way to prevent influenza virus infections, but the diversity of antigenically distinct isolates is a persistent challenge for vaccine development. In order to conquer the antigenic variability and improve influenza virus vaccine efficacy, our research group has developed computationally optimized broadly reactive antigens (COBRAs) in the form of recombinant hemagglutinins (rHAs) to elicit broader immune responses. However, previous COBRA H1N1 vaccines do not elicit immune responses that neutralize H1N1 virus strains in circulation during the recent years. In order to update our COBRA vaccine, two new candidate COBRA HA vaccines, Y2 and Y4, were generated using a new seasonal-based COBRA methodology derived from H1N1 isolates that circulated during 2013–2019. In this study, the effectiveness of COBRA Y2 and Y4 vaccines were evaluated in mice, and the elicited immune responses were compared to those generated by historical H1 COBRA HA and wild-type H1N1 HA vaccines. Mice vaccinated with the next generation COBRA HA vaccines effectively protected against morbidity and mortality after infection with H1N1 influenza viruses. The antibodies elicited by the COBRA HA vaccines were highly cross-reactive with influenza A (H1N1) pdm09-like viruses isolated from 2009 to 2021, especially with the most recent circulating viruses from 2019 to 2021. Furthermore, viral loads in lungs of mice vaccinated with Y2 and Y4 were dramatically reduced to low or undetectable levels, resulting in minimal lung injury compared to wild-type HA vaccines following H1N1 influenza virus infection.

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Oseltamivir-Ribavirin Combination Therapy for Highly Pathogenic H5N1 Influenza Virus Infection in Mice

Antiviral Research ◽

10.1016/j.antiviral.2007.01.020 ◽

2007 ◽

Vol 74 (3) ◽

pp. A31-A32 ◽

Cited By ~ 2

Author(s):

N ILYUSHINA ◽

R WEBSTER ◽

E GOVORKOVA

Keyword(s):

Influenza Virus ◽

Virus Infection ◽

Combination Therapy ◽

Influenza Virus Infection ◽

H5n1 Influenza Virus ◽

Highly Pathogenic ◽

H5n1 Influenza ◽

Pathogenic H5n1

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Tropism and Infectivity of a Seasonal A(H1N1) and a Highly Pathogenic Avian A(H5N1) Influenza Virus in Primary Differentiated Ferret Nasal Epithelial Cell Cultures

Journal of Virology ◽

10.1128/jvi.00080-19 ◽

2019 ◽

Vol 93 (10) ◽

Cited By ~ 6

Author(s):

Hui Zeng ◽

Cynthia S. Goldsmith ◽

Amrita Kumar ◽

Jessica A. Belser ◽

Xiangjie Sun ◽

...

Keyword(s):

Influenza Virus ◽

Virus Infection ◽

Avian Influenza ◽

H5n1 Virus ◽

H1n1 Virus ◽

Influenza Virus Infection ◽

Influenza Viruses ◽

Host Interactions ◽

Culture Model

ABSTRACTFerrets represent an invaluable animal model to study influenza virus pathogenesis and transmission. To further characterize this model, we developed a differentiated primary ferret nasal epithelial cell (FNEC) culture model for investigation of influenza A virus infection and virus-host interactions. This well-differentiated culture consists of various cell types, a mucociliary clearance system, and tight junctions, representing the nasal ciliated pseudostratified respiratory epithelium. Both α2,6-linked and α2,3-linked sialic acid (SA) receptors, which preferentially bind the hemagglutinin (HA) of human and avian influenza viruses, respectively, were detected on the apical surface of the culture with different cellular tropisms. In accordance with the distribution of SA receptors, we observed that a pre-2009 seasonal A(H1N1) virus infected both ciliated and nonciliated cells, whereas a highly pathogenic avian influenza (HPAI) A(H5N1) virus primarily infected nonciliated cells. Transmission electron microscopy revealed that virions were released from or associated with the apical membranes of ciliated, nonciliated, and mucin-secretory goblet cells. Upon infection, the HPAI A(H5N1) virus replicated to titers higher than those of the human A(H1N1) virus at 37°C; however, replication of the A(H5N1) virus was significantly attenuated at 33°C. Furthermore, we found that infection with the A(H5N1) virus induced higher expression levels of immune mediator genes and resulted in more cell damage/loss than with the human A(H1N1) virus. This primary differentiated FNEC culture model, recapitulating the structure of the nasal epithelium, provides a useful model to bridgein vivoandin vitrostudies of cellular tropism, infectivity, and pathogenesis of influenza viruses during the initial stages of infection.IMPORTANCEAlthough ferrets serve as an important model of influenza virus infection, much remains unknown about virus-host interactions in this species at the cellular level. The development of differentiated primary cultures of ferret nasal epithelial cells is an important step toward understanding cellular tropism and the mechanisms of influenza virus infection and replication in the airway milieu of this model. Using lectin staining and microscopy techniques, we characterized the sialic acid receptor distribution and the cellular composition of the culture model. We then evaluated the replication of and immune response to human and avian influenza viruses at relevant physiological temperatures. Our findings offer significant insight into this first line of defense against influenza virus infection and provide a model for the evaluation of emerging influenza viruses in a well-controlledin vitroenvironmental setting.

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