Research Of Avian Influenza Virus H5N1Neuraminidase Induced Th17Differentiation Of Cd4~+T Cells In Mice

Avian influenza has been plaguing the world’s poultry industry, and leading a huge economic lose. Meanwhile, AIV can transmit to human from poultry also threatens people’s health. The high mortality caused by highly pathogenic avian influenza virus can give rise to social panic. AIV neuraminidase (NA) is composed of four identical, single-chain polypeptide units, and is a glycosylated membrane protein having sialic acid hydrolysis activity. NA is one of the bases for AIV divided into different subtypes, and also related to AIV’s virulence and specificity directly. Th17cells are associated with inflammatory cell infiltration, resulting in host tissue damage and severe pathological disorders of the body. Our study found that AIV H5N1NA involved in CD4+T cell Th17differentiation in mice, which speculated that have related to excessive inflammation and cytokine storm lead by AIV. The main results obtained in this study are as follows:1. H5N1NA can infect mice and cause CD4+T cell Th17subset increase in lungIntranasal infect5week old mice with AIV H5N1NA in different dose. And then, take the mouse lung tissue T cells cultured in vitro, after48h, flow cytometry analysis Th17and Treg cell. As a result, the proportion of Th17cells increased and in a dose-dependent.2. AIV NA can promote BMDC maturationCultured BMDC in vitro, and then stimulate BMDC by lipopolysaccharide (LPS), H1N1NA, H5N1NA, H9N2NA, as well as the heat-inactivated H5N1NA and ovalbumin (OVA) in different period. Flow cytometry analysis the CD11c, MHC-II and co-stimulatory molecules CD40, CD80, CD83and CD86, which are on the surface of the BMDC. The results found that the AIV NA can promote dendritic cell maturation by the activity, and it has a dose-dependent in a certain range.3. H5N1NA induce can induce Th17and other CD4+T cells differentiationBMDC and CD4+T cells isolated from spleen of mice co-cultured in vitro. Stimulate the cultured cell with H5N1NA and other samples, and then test samples by ELISA, fluorescence quantitative PCR and flow cytometry analysis. As a result, the H5N1NA can induce CD4+T cell polarization by stimulate the BMDC in5th day and9th day. Experimental results suggest that IL-21and TGF-β induce Th17differentiation in early stage, while IL-6and TGF-β induce Th17differentiation in late stage. In this process, cells can release a series of inflammatory cytokines, such as TGF-P, IL-6, IL-21, IL-23, IL-1β,IL-17A,IL-10.