The Effects Of Human Avian Influenza H5N1 Virus And Its NS1 Protein On Chemokine Expression In Host Cells

Human avian influenza is a newly emerging infectious disease which is characterized by acute respiratory tract infection in human and is directly caused by avian influenza A viruses including H5N1, H7N7 and H9N2. Severe clinical features and high mortality were observed in human H5N1 infection. But there isn't a consistent answer as to its pathogenesis mechanisms until now. Recent years, Some studies show that abundant immunological dysfunctions were detected in H5N1 patients, including high production of pro-inflammatory cytokines and chemokines, such as IL-1, IL-6 , TNF-α, IL-8, MCP-1, IP-10, MIG and so on, In the body's normal immune response, these cytokines can protect the body from the defence of foreign pathogens, but in a specific environment, the immune cells will attack normal tissue due to excessive activation, so it's probably that the elevated cytokines may contribute to the high pathogenicity of H5N1 viruses. Respiratory epithelial cells and monocyte/macrophages are the primary target cells of influenza virus. In virus-infected cells, there was abundant viral nonstructural NS1 protein. NS1 protein can participate in the antiviral immune response, regulate cell apoptosis, viral replication and inhibit the protein expression in host. Here, we compared the chemokine response after infect host cells with human influenza viruses, recombinant viruses and transfect cells with recombinant NS1 plasmids. There are the main findings in our study.1. In contrast to human influenza virus H1N1, stronger IP-10 response was detected in human H5N1 virus-infected cells, which is virus and cell type-specific. There is more than 10-fold in both mRNA and protein level between A/Anhui/1/2005(H5N1)virus and human influenza virus A/Puerto Rico/8/1934(H1N1). The strongest IP-10 response was also detected in human epithelial cell line BEAS-2B infected with A/Anhui/1/2005(H5N1)than other H5N1 viruses. Our finding suggested that the hyper-induction of IP-10 may be associated with the pathogenesis in human H5N1 disease.2. The single NS1 gene from virus A/Anhui/1/2005(H5N1), NS1 gene inserted with 80-84 amino acids from virus A/Anhui/1/2005(H5N1) and NS1 gene from virus A/Puerto Rico/8/1934(H1N1) can down regulate IP-10 expression level in BEAS-2B cells (p < 0.01). But no significant difference was detected among them. The finding indicated that NS1 protein maybe involved in the downregulation of IP-10 expression. 3 Three recombinant viruses of A/Puerto Rico/8/1934(H1N1)virus backbone with different NS gene (PR8-RG, PR8-NS-RG and PR8-mNS-RG virus) were used to infect BEAS-2B cells and monocyte-derived macrophages. Differential chemokine expression level was detected in both cell types with the similar infectivity and similar expression level and kinetics of NS protein of these three viruses. The chemokine profile is different in virus-infected BEAS-2B cells and macrophages. In BEAS-2B cells, PR8-RG virus induced higher level of IP-10 and RANTES than PR8-NS-RG, PR8-mNS-RG and mock (p<0.05). While in monocyte-derived macrophages, PR8-RG induced higher level of RANTES and MIG than others. The effect of PR8-mNS-RG on chemokine response is the lowest. These results suggested that the NS gene of H5N1 was more potential in down-regulating chemokine expression than NS gene of H1N1 virus and the deletion of 80-84 amino acid may play a role in antagonism to cytokine/chemokine response in host cell.Higher chemokine response such as IP-10 could be induced by human H5N1 virus. However, the downregulation effect of H5N1 NS on chemokine response was detected in our study by using the transfection and recombinant virus infection model. It suggested that other factors than viral NS protein are involved in the interaction of virus and host cells which may contribute to the hyperinflamatory response in human H5N1 disease.