| Objective：1.The experimental group early build the optimized Parkinson nucleicacid vaccine pVAX1-IL-4/SYN-B and full-length DNA vaccinepVAX1-hαS1-140, for the prevention of vaccinated mice to furtherinvestigate the DNA vaccine-induced immune response results, in order tofilter out the more effective vaccines, and to lay the foundation for futureresearch.2. To evaluate the security of optimization the nucleic the vaccinepVAX1-IL-4/SYN-B and full-length DNA vaccine pVAX1-hαS1-140byprevention of inoculated mice, in order to filter out the safer vaccine, to laythe foundation for future research.MethodsIn this study, first to optimize DNA vaccine、 empty vector、full-length DNA vaccine using an endotoxin-free plasmid with highpumping kit for extraction and preparation.The next choice of32C57BL/6Jmice were randomly divided into control group, empty vector group, optimized DNA vaccine group, full-length DNA vaccine group, four groupsof animals with bilateral hind tibia bone anterior muscle of the same partsof the same depth of inoculation PBS, pVAX1, pVAX1-IL-4/SYN-B,pVAX1-hαS1-140, each of the injection of50μl inoculated five times atintervals of three weeks. After2weeks after immunization and eachimmunization vaccination tail blood, separation of serum stored at-80℃.In addition to re-selection of eight C57BL/6J mice, subcutaneous injectionof rotenone (Sigma, USA)1mg/kg (Preparation method: DMSO: rotenone:saline=3.5:1g:0.5L),1time/d, continuous injection of40d. For thepreparation of chronic rotenone Parkinson mouse brain slices. In miceimmune and model preparation process, to observe mouse behavior change.Indirect ELISA determination of serum antibody titer of anti-α-syn andIL-4and IFN-r levels; immunohistochemistry and α-syn peptide antigens inthe assay antiserum chronic rotenone Parkinson mouse brain slices.thespecific binding of the α-syn-positive inclusion bodies; by RT-PCRdetection of the mice in the substantia nigra of COX-2expression.Results1.In the process of immunization, the mice behavior science noabnormal changes. The average body weight of each group of mice beforeinoculation and after inoculation, no significant difference in change.Chronic rotenone Parkinson’s mouse model preparation process, the micetail arched uplift, stiffness, tremors, lack of exercise, huddled, limb movement asymmetry and other types of Parkinson-like symptoms.2. Compared with the control group, pVAX1-IL-4/SYN-B vaccine2weeks after the first immunization, α-syn antibodies, with the increase inthe number of strengthening the immune antibody titers increasedsignificantly (p <0.01).3.Immune factors in serum: IL-4of optimizing nucleic acid vaccinewas significantly higher, IFN-r is less than the control group. The otherthree groups no significant difference.4.Immunohistochemical staining showed: optimized DNA vaccine andthe full-length DNA vaccine in mice serum and α-syn-positive inclusionbodies immune chronic fish rotenone Parkinson mouse brain slices andalpha-syn peptides and the control group andempty plasmid group, noreaction.5.RT-PCR showed: COX-2expression of optimization DNA vaccinegroup was significantly less than the full-length nucleic acid vaccine group(p <0.01), and the control group, empty vector group no significantdifference (p>0.05).ConclusionOptimization DNA vaccine pVAX1-IL-4/SYN-B can in mice producehigh titers of α-syn antibody, mainly caused by the Th2immune response,and safer than full-length DNA vaccine pVAX1-hαS1-140and found novaccination causing side effects. Therefore, our results suggest that optimized DNA vaccine more than the full-length vaccine in accordancewith immunotherapy requirements of Parkinson’s disease, provide theconditions for the Next immune Parkinson animal studies, laid thefoundation for further screening for optimized DNA vaccine... |
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