Influenza A Virus Production in Mouse Lung Is Inhibited by Inhalation of Aerosol Polyethylenimine/Short Hairpin RNA Plasmid Complexes

Abstract

Influenza pandemics are a global threat to human health, with existing vaccines and antiviral drugs providing limited protection. There is an urgent need for new prophylactic and treatment strategies. In this study, 12 short hairpin (sh)RNAs targeting conserved regions of influenza A virus (IAV) matrix protein (M)2, nucleocapsid protein (NP), nonstructural protein (NS), and polymerase acidic (PA) were synthesized, and their effects on IAV replication in cells were investigated using Madin–Darby canine kidney (MDCK) cells transfected with the shRNA plasmids. Additionally, mice were administered a polyethyleneimine PEI/pLKD-NP-391 complex in aerosol form and then infected with AIV, and viral particles in the mouse lung were detected. IAV production was markedly lower in MDCK cells transfected with pLKD-M-121, pLKD-M-935, pLKD-NP-391, pLKD-NP-1291, pLKD-PA-1365, and pLKD-PA-1645 plasmids than in control cells ( $p<0.01$ ). The viral load in MDCK cells was decreased by transfection of plasmids pLKD-M-121 ( $p<0.05$ ) and pLKD-M-935, pLKD-NP-391, pLKD-NP-1291, pLKD-PA-1365, and pLKD PA-1645 ( $p<0.01$ ) compared to an empty plasmid. The viral load was significantly lower in the lungs of mice transfected with pLKD-NP-391 than in the control plasmid and mock transfection groups ( $p<0.01$ and $p<0.05$ , respectively). Thus, IAV production was inhibited by shRNAs targeting matrix IAV components; moreover, inhalation of a PEI/pLKD-NP-391 complex in aerosol form suppressed IAV production in infected mice. Thus, these shRNAs can be effective for the prevention and treatment of influenza virus infection.