Als3 T Cell Epitope And Adjuvant Synergize To Enhance Cellular Immune Responses Induced By Staphylococcus Aureus TRAP In Mice

Staphylococcus aureus(S.aureus)not only threats human health,but also causes serious economic loss.Study shows that Target of RNAIII Activating Protein(TRAP),a membrane-related protein of S.aureus,responds to anti-oxidative stress and induces better immunoprotection against S.aureus infections.Als3,an adhensin of Candida albicans,can induce robust cellular immune responses and cross-protection against S.aureus.In order to enhance TRAP induced-cellular immune responses,we constructed a recombinant protein ATT by fusing Als3 T cell epitope to TRAP and formulated ATT with different adjuvants,and detected the antibody levels,specific cytokines,and immunoprotection induced by ATT in the immunized mice.In this study,a linker was used to connect the Als3 T cell epitope and the N-terminal of TRAP to form the recombinant protein,which was named ATT.While TRAP,e Als(the Als3 T cell epitope repeated six times linked to p ET-28 a vector),e Als+TRAP and PBS were used as the controls.All the expressed proteins and PBS were emulsified in isovolumetric complete Freund's adjuvant before immunizing the mice via the tibialis anterior.Three weeks after the immunization,antigens were emulsified in isovolumetric incomplete Freund's adjuvant to give the mice a booster immunization.After the booster immunization,the mice were challenged with the S.aureus strains Newman and Wood46.Then the concentrations of cytokines(IFN-?,IL-4,IL-10,and IL-17A),the levels of antibodies,and the subclasses of IgG were analyzed.The results showed that the ATT group displayed significant IFN-?,IL-4,IL-10,and IL-17 A responses,and the levels of total antibodies and the production of of IgG1 were significantly elevated,in comparison with the other groups(except the e Als+TRAP group).The concentrations of cytokines(IFN-?,IL-4,IL-10,IL-17A),the levels of antibodies and subclasses of IgG induced by ATT were significantly higher than those induced by TRAP.Furthermore,the survival rates of the ATT group were higher than the other groups after challenge.These results suggested that the recombinant protein ATT had a better preventive effect than TRAP,and could enhance the immunogenicity of TRAP protein.Further,ATT protein was combined with corresponding adjuvants,including FIA,CpG,MDP,MDP+CpG,CpG+FIA,MDP+FIA,and CpG+MDP+FIA.And the C57BL/6 mice were immunized and challenged according to the above methods.The results showed that the IgG antibody titers and the production of IgG1 in the mouse sera had a great difference between the groups,and the concentrations of IFN-? and IL-17 in the splenic cells of the ATT+CpG+MDP+FIA group increased significantly(p<0.01),compared with other groups.Moreover,the survival rate of the mice immunized with ATT+CpG+MDP+FIA was the highest after the challenge with two S.aureus strains.These results suggested that the vaccine formulation,ATT+CpG+MDP+FIA,was more potent than other vaccine formulation in enhancing cellular immune responses and immune protection.This study provided a reference for developing the novel and high-efficiency vaccines against S.aureus infections.