Experimental Study Of Construction Of Dendritic Cell(DC)-targeting DNA Vaccine PRSC-NLDC145·gD-IL-21 And Its Prophylactic Function Of Herpes Simplex Keratitis

Objective:To construct DC-targeting DNA vaccine pRSC-NLDC145 · gD-IL-21 and to study its ability to induce inmmune response and to resist the HSV-1 infection of cornea in mice.Methods:The recombinant plasmid of DC-targeting DNA vaccine pRSC-NLDC145 · gD-IL-21 was constructed through gen cloning method.First through the restriction enzyme,we removed the original PRSC-gD-IL-21(existing plasmid)in gD gene,then connected gD to NLDC145 which cloning to gD original site by the enzyme digestion and connection,while keeping the original cloning site of another immune adjuvant IL-21.After being identified by molecular methods,we constructed chitosan nanoparticles,then the vaccine pRSC-NLDC145·gD-IL-21+chitosan,pRSC-gD-IL-21+chitosan,pRSC+chitosan and the chitosan were respectively inoculated into BALB/c mice for 3 times with 2weeks interval.The levels of immune response and the eye infection of HSV-1 immune protective reaction were detected 2 weeks after the last immunization.Result:The data presented showed that the targeted genes were successfully expressed in the transfected 293T cells after the DNA vaccine was identified by DNA sequencing,the restricted endonuclease analysis.The agarose gel electrophoresis analysis confirmed that the nanoparticles could be combined with plasmids through electrostatic interactions.Compared with the control mice,the DC-targeting DNA vaccine pRSC-NLDC145·gD-IL-21+chitosan induced mice to generate higher levels of antibody,specificity sIgA secretion levels in mice tears,and enhanced the activites of CTL and NK.Meanwhile,the vaccine could elicit a stronger immunoprotective effect against the infection of HSV-1 in mice.Conclusion:These findings indicate that the DC-targeting DNA vaccine pRSC-NLDC 145·gD-IL-21 +chitosan may induce a stronger immune response in immunized mice,also indirectly prove that the purpose of antigen can eventually targeted to DC.More importantly,This study will fundamentally help with the vaccine for the future application in the treatment of recurrent HSK.