Effect of specific cellular immune response induced by anti-Aedes albopictus salivary protein rAlb-34k2-1 on DENV-2 replication in BMDM cells
Objective To investigate the effect of Aedes albopictus salivary protein rAlb-34k2-1-induced mouse splenocytes immune response on dengue virus type 2(DENV-2)-infected bone marrow-derived macrophages(BMDM)in vitro.Methods The specific cell proliferation response of splenic cells from immunized/non-immunized mice to Alb34k2-1 protein was analyzed using the CCK-8 assay.The antigen-presenting cells(APCs)loaded with mosquito saliva proteins rAlb-34k2-1 protein/rAalb_CTL1 protein were prepared and co-cultured with lymphocytes from spleens of immunized/non-immunized mice.On the one hand,the levels of DENV-2 nucleic acids in BMDM treated with cell supernatants from mixed lymphocyte co-cultures were analyzed by qRT-PCR,and the effect of specific T cell response induced by rAlb-34k2-1 on DENV-2 replication in BMDM was analyzed.On the other hand,the expression of the DENV-2 E gene in BMDM pretreated with rAlb-34k2-1 protein was also evaluated to determine the direct effect of immunized mouse spleen lymphocytes co-cultured at 48 h and 72 h and the effect of spleen lymphocytes from immunized or non-immunized mice on DENV-2 replication in pretreated BMDMs was analyzed.Results Compared to the non-immunized group,the rAlb-34k2-1 protein in vitro caused a specific proliferation of immunized-splenocytes,with a proliferation index(SI)>2(P<0.000 1).Mixed lymphocyte culture supernatants loaded with rAlb-34k2-1 protein inhibited DENV-2 replication in BMDM,while the mixed lymphocyte culture supernatants loaded with rAalb_CTL1 protein had no such effect(2-ΔΔCT<0.5,P<0.05).There was no difference in DENV-2 intracellular replication in rAlb-34k2-1 pretreated BMDM after co-culture with either immunized-lymphocytes or non-immunized-lymphocytes.Conclusions The supernatant induced by the mouse T cell response to the rAlb-34k2-1 protein can inhibit DENV-2 replication in BMDM cells,providing preliminary experimental evidence to explore the role of mosquito salivary protein-induced immune responses in the pathogenesis of mosquito-borne viruses.
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