Transfection of rAAV9-Galectin-9 into PBMCs inh ibits Th17 immune responses in mice with RSV infec-tion

Abstract

Objective To evaluate the transfection efficiency of a recombinant adeno -associated virus serotype9 （ rAAV9 ） vector carrying Galectin-9 gene （ rAAV9-Galectin-9 ） into peripheral blood mononuclear cells （ PBMCs） and to analyze the effects of Galectin-9 on Th17 cell differentiation in a mouse model of respira -tory syncytial virus （RSV） infection.Meth ods A mouse model of RSV infection was established.Samples of lung sections were stained with HE or PAS to observe inflammatory infiltration.PBMCs were isolated from mice and transfected with rAAV9-Galectin-9.The transfection efficiency was evaluated by using fluorescence micro-scope.The expression of RORγt at mRNA level was detected by quantitative PCR.Flow cytometry analysis was performed to measure the levels of Th17 and Treg cells.ELISA was performed to measure the levels of IL-10, IL-17 and TGF-β1 in the supernatants of cell culture.Results Compared with control group, the mice with RSV infection showed an increased expression of ROR γt at mRNA level and elevated levels of T1h7 cells and IL-17 （all P〈0.05）, but decreased levels of Treg cells and IL-10 （all P〈0.05）.No significant difference with TGF-β1 level was observed.The expression of RORγt at mRNA level and the levels of Th17 cells and IL-17 were decreased in mice with RSV infection upon the transfection of Galectin -9 gene（all P 〈0.05）.Moreover, the levels of Treg cells and IL-10 were increased in mice from RSV＋Galectin-9 treated group （all P〈0.05）.No significant differences were observed between RSV infection group and RSV＋ZsGreen treated group.Conclusion rAAV9-Galectin-9 was successful transfected into PBMCs.rAAV9-Galectin9-could regulate Th17 immune re-sponses through Galectin-9 pathway during RSV infection. Key words: Respiratory syncytial virus; Galectin-9; Thl7 cell; Recombinant adeno-associated virusserotype 9