Neutrophil Mediated Host Responses during Influenza A Virus Infection, An In Vitro Study

Abstract

Neutrophil (Nϕ) is the most abundant cell of the immune system in humans. In an 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. Nϕs were freshly isolated from healthy volunteers and subjected to influenza H1N1 and H5N1 virus infection. The susceptibility of Nϕ to influenza A virus infection was assessed by the viral matrix gene expression and viral titration assay. Innate immune response of Nϕ was evaluated by monitoring the gene and protein expression of the naïve Nϕ with and without influenza virus infection, using qPCR assay and ELISA. The induction of the de novo neutrophil extracellular trap (NET) was evaluated by scanning electron microscopy and SYTOX green staining. Our results demonstrated that naïve Nϕs were equally susceptible to H5N1 and H1N1 virus infection with similar viral gene transcription. Productive replication was observed in H5N1 infected Nϕs only. H5N1 induced higher cytokine and chemokine gene transcription and protein secretion than H1N1 infected Nϕs, including TNFα, IFNβ, CXCL10, MCP-1, MIP-1α and IL-8. This inferred a more intense inflammatory response posed by H5N1 than H1N1 virus. Strikingly, NET formation was only observed in H1N1 infected Nϕs at 6 hpi and it was not found in H5N1 infected cells. 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.