| Dendritic cells(DCs)are the most important antigen-presenting cells.Vaccine preparations promote subsequent adaptive immune responses by inducing activation and maturation of DCs.Although existing vaccine preparations can mobilize DCs,they do not provide DCs with more efficient immune responses.To solve this problem,we constructed PLGA nanospheres and conjugated DEC205 antibody to achieve better DCs function and further improve immune response.The specific research content includes the following four parts:(1)The multifunctional nano-microsphere vaccine adjuvant targeting DCwas fabricated.Using PLGA as material and OVA as model antigen,PLGA nanospheres were prepared by double emulsion-solvent extraction combined with rapid membrane emulsification.The biomimetic material dopamine was employed to coat the surface of PLGA nanospheres,and further coupled with DEC205 antibody,and the multi-functional nanospheres targeting DC were successfully constructed.Nanospheres showed pH-sensitive release profile of antigens in an environment that mimics intracellular endosomes,lysosomes and cytoplasm in vitro.The cytotoxicity test and biosafety evaluation suggested that the nanospheres had good biosafety.Nanospheres exhbited little effect on the viability of dendritic cells at a concentration of 250 μg/mL.(2)The immune efficacy of DC-targeted multifunctional nanospheres as vaccine adjuvants was evaluated at the animal level.Nanoparticles that target DCs could effectively induce the activation of lymphocytes(including B cells,CD4+T cells,and CD8+T cells),produce powerful immune effects in the effect phase,and could promote the secretion of cytokines from splenocytes,resulting in higher Levels of antigen-specific antibodies standing for increase of humoral immunity and enhance immune memory.When there were secondary incursions of pathogens,they could rapidly mobilize immune responses and produce immune cells that exert immune functions.(3)The immune mechanism of multi-functional nanospheres targeting DC as vaccine adjuvants was expored.From the cellular level,DC-targeting multi-functional nanospheres could promote DCs to take up more antigens,promote antigens to escape from lysosomes,present antigens via MHC I pathways,and significantly increase the level of cross-presentation of antigens.From the animal level,DC-targeted multi-functional nanospheres could recruit more immune cells at the injection site,especially DCs,which could effectively promote the secretion of cytokines and chemokines at the injection site and increase lymph node DCs and Thf.By blocking the surface DEC205 ligand and DTR mouse experiments on DCs,it was confirmed that the nanospheres exerted immune function mainly through DCs,and compared with DC microspheres without targeting DCs,exerted more effective immune function of DCs(4)Tumor immune effects of DC-targeted multifunctional nanospheres as vaccine adjuvants were evaluated.By constructing two kinds of B16F10 melanoma models for prevention and treatment,the anti-tumor immunity prevention and tumor immunotherapy effects of nano microspheres were examined.Nanoparticles targeting DC could significantly inhibit the growth of tumors,prolong the survival time of mice,enhance the ability of spleen lymphocytes of mice to specifically kill tumor cells after immunization,and increase the number of CD4+T cells and CD8+T cells in spleen cells.The ratio of IFN-y and Grammzy B in the killer function promoted the secretion of TGF-β and lowered the level of the negative regulatory cytokine TGF-β. |
