| Influenza virus, belongs to the family of Orthomyxoviridae, could generate severe contagious respiratory disease, which poses rigorous threat to public health worldwide, not only because of the individual cases of infection and death, but also because of the spread of highly pathogenic influenza virus could frequently transmitted to human and mammals worldwide, leading to a potential novel influenza pandemic. It is therefore urgently necessary to develop a novel and broad-spectrum medication of preventing influenza dissemination.Silver nanoparticles (Silver-nps) have shown efficient antiviral activities against human immunodeficiency virus (HIV) and hepatitis B virus (HBV). However, the inhibitory effects and mechanisms of Silver-nps on influenza A virus (IFV) have not been evaluated. In this study, we will explore the inhibitory effects of Silver-nps against influenza A/Human/Hubei/3/2005 virus (H3N2-IFV) and their possible mechanismsInitially, following the interaction of Silver-nps with H3N2-IFV, we observed that Silver-nps weakened the viral neuraminidase activities and inhibited the ability of H3N2-IFV to agglutinate the erythrocytes. Compared with the titers 1:1024 of both the virus control and solvent group, the viral HA titers in silver-np groups were 1:16, 1:8, 1:4, and below 1:2 at 30 min, 60 min, 90 min and 120 min, respectively. Furthermore, in the presence of Silver-nps, the results of viral HA titers in embryonating eggs among diverse samples were statistically significant. Compared with the titers 1:1024 of both the virus control and solvent group, the viral HA titers in Silver-nps groups were below 1:2 (P < 0.001). These results suggest that Silver-nps play a significant role by obstructing the binding of H3N2-IFV to the RBCs. Silver-nps added to canine kidney cells (MDCK) before and after the cells was adsorpted by H3N2-IFV influenza virus, using cytopathic effect(CPE)assay, MTT assay and the immunofluorescence test to analysis the preventive effect, directly off deactivation and the inhibit formation of progeny virions of Silver-nps on H3N2-IFV virus. MDCK cells infected with H3N2-IFV influenza virus appeared much specific immunofluorescence when compared to the cells treated with Silver-nps in the three different ways. The results showed Silver-nps could inactivate H3N2-IFV virus, remarkedly prevented them invasing to the cells and reproducting when H3N2-IFV entered to cells. Under Transmission Electron Microscope(TEM) negative staining assay, the viral morphological structures were changed obviously when H3N2-IFV influenza viruses were interactived with Silver-nps.The results demonstrated that Silver-nps maight destroy viral morphological structures so that they could disturb viral functions. Moreover, TEM analysis and a flow cytometry(FCM) assay were used to detect whether Silver-nps could reduce H3N2-IFV-induced apoptosis in MDCK cells. Normal cells and cells infected with the silver-np-treated virus for 2 h first revealed normal silhouettes while virus control displayed characteristics of apoptosis and apoptotic bodies in some cells, suggesting that Silver-nps can inhibit H3N2-IFV infectivity in vitro. To gain a further insight into the mechanism of the inhibitory effects of Silver-nps on H3N2-IFV, the reverse transcription-polymerase chain reaction (RT-PCR) was used for monitoring changes in gene expression patterns occurred by the nanoparticles. Both the virus control and the solvent control groups appeared as significant bright bands in the 1700bp area, whereas the Silver-nps group did not. Here, we present the initial findings that Silver-nps have efficacious anti-H3N2-IFV activities and the underlying mechanisms of which may be that Silver-nps interferes with H3N2-IFV both at viral entry and viral release during the process of viral replication. The efficient inhibitory activities of Silver-nps against H3N2-IFV suggest that Silver-nps may play an important role for treatment and prevention of IFV infection.
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