Optimized DNA Vaccine Ameliorates Deficits Of Striatal Dopamine Terminals In A MPTP Model Of Parkinson’s Disease

Objective：To explore optimized DNA vaccine ameliorated deficits of striataldopamine terminals associated with MPTP and the mechanism ofimmunotherapeutic effect of optimized DNA vaccine on striatal dopamineterminals in a MPTP model of Parkinson’s disease.Methods：Firstly，we extracted and prepared sufficient highly pure emptyplasmid（pVAX1）and optimized DNA vaccine（pVAX1-IL-4/SYN-B）by endofree plasmid midi kit. All experimental mice（C57BL/6，male，22-25g） were randomly divided into four groups：normal control group（PBS+PBS，n=10），PBS PD model group（MPTP+PBS，n=10），empty plasmid PD model group（MPTP+pVAX1，n=10），optimized DNAvaccine PD model group（MPTP+pVAX1-IL-4/SYN-B，n=15）. Then，we used a chronic MPTP model of Parkinson’s disease. Mice of modelgroup were injected with MPTP，in the same way the normal control groupwere injected with the equivalent volume of PBS. After succeeded in building chronic MPTP model， the four groups were respectivelyinoculated with PBS，PBS，pVAX1，pVAX1-IL-4/SYN-B in the anteriortibial muscle of bilateral hind limb of each mouse. After the lastinoculation， Chloroquine was stereotaxically injected into the lateralventricle of5of15mice treated with pVAX1-IL-4/SYN-B，in the sameway the were injected into the lateral ventricle of other experimental micewith the equivalent volume of PBS. We observed changes of the behaviorin each group of mice. The expression level of tyrosine hydroxylase（TH），synaptophysin（SYP），alpha-synuclein（α-syn），microtubule-associatedprotein1light chain3（LC3），lysosomal-associated membrane protein1（Lamp1）and cathepsin D（CD）in striatum of mice were detected bywestern blot and immunohistochemistry.Results：1. Behavior change In the process of building chronic MPTP model，each mouse in model groups appeared the same degree of Parkinson-likesymptoms： tremor，vertical hair，warped-tail，hypokinesia，bradykinesia，staggering gait and postural instability. After the last inoculation，behavioral deficits in mice of PD model groups was increasedsignificantly，compared with normal control group（P＜0.05）. Behavioraldeficits in optimized DNA vaccine PD model group was improvedsignificantly, compared with PBS PD model group or empty plasmid PDmodel group（P＜0.05）. 2. Immunohistochemistry change Immunohistochemistry showedthe expression level of TH and SYP in striatum of mice of PD modelgroups were reduced significantly，compared with normal control group（P＜0.05）. The expression level of TH and SYP in striatum of mice ofoptimized DNA vaccine PD model groups were increased significantly，compared with PBS PD model group or empty plasmid PD model group（P＜0.05）.3. Western blot change Western blot showed the expression level ofTH and SYP in striatum of mice of PD model groups were reducedsignificantly，compared with normal control group（P＜0.05）. Theexpression level of TH and SYP in striatum of mice of optimized DNAvaccine PD model groups were increased significantly，compared with PBSPD model group or empty plasmid PD model group（P＜0.05）.Theexpression level of α-syn dimer in striatum of mice of PD model groupswere increased significantly，and the expression level of CD and Lamp1were reduced significantly，compared with normal control group（P＜0.05）.The expression level of α-syn dimer in striatum of mice of optimizedDNA vaccine PD model groups were reduced significantly，and theexpression level of CD and Lamp1were increased significantly，comparedwith PBS PD model group or empty plasmid PD model group（P＜0.05）. Conclusions：From these results we can conclude that chronic MPTP can damage tomotor function and striatal dopamine terminals in C57BL/6mice，themolecular pathological mechanisms may be lysosomal depletion by chronicMPTP，which raises the level of α-syn dimer，or interact as both cause andeffect. Optimized DNA vaccine can ameliorate deficits of behavior andstriatal dopamine terminals associated with MPTP，the immunotherapeuticmechanisms may be repairing lysosome and promoting presynaptic α-syndimer clearance via the lysosomal pathway.