| BackgroundAlzheimer's disease (AD) is the most common form of dementia and afflicts more than 20 million people world-wide. Currently, its etiology is not fully understood so there is no effective way to prevent or cure this devastating disease,Aβhas become a therapeutic target for the prevention and treatment of AD because of its presence in neuritic plaques, its neurotoxicity in vitro and in vivo, and its in-creased levels in humans with familial AD mutations in APP or the presenilin (PS1, PS2) genes. Unfortunately, the phaseⅡa trial of AN1792 was halted when 6% of the participants in the clinical trial developed aseptic meningoencephalitis. Further study found that some epitopes on the C-terminal of Aβ1-42 is associated with T-cell responses,while the Aβ4-10 on the N-terminal was definiteed as B-cell epitopes is associated with B-cell responses and the affinity between anti-Aβand Aβmaybe markedly enhanced if sequence coupled with the third amino acid on the N-terminal of Aβ1-42.ObjectiveTo detect the type of immune responses to DNA Aβ3-10 immunization for BALB/c miceMethodsWith the endonucleases EcoRI, NotI digested pcDNA3.1+-(Aβ3-10) 10-CpG, then the product was combined with T4 ligase enzyme to construct pcDNA3.1+-(Aβ3-10)10;then extract plasmid pcDNA3.1+-(Aβ3-10) 10-CpG,pcDNA3.1+-(Ap3-10)10 by alkaline lysis method;BALB/c mice were immunized with the plasmids by musle injection five times; meanwhile settingthe Aβ1-42 as positive control group and setting PBS as negative control group;splenocytes from experimental and control mices were divided on 96 well plates in 1640 supplimented with 10% FCS(fetal calf serum) and restimulated respectively with antigen[(Aβ3-10)4] or ConA and incubated at 37℃with 5%CO2 for 48 h;Then detected the splenocyte proliferation by MTT method;Detected the concentration of IFN-y and IL-4 contained in the supernatant of splenocytes by ELISA method. Analyze statistical data by t test with SPSS softwareResultsThe result of splenocyte proliferation that every group restimulated with corresp antigen demonstrate that the SI of experimental groupl are significantly higher than the SI of experimental group2((.P<0.05);the SI of groupl significantly lower than the Aβ1-42 group(positive control) (P<0.05);the SI of group2 significantly higher than the PBS group(P<0.05) (negative control);groups restimulated with ConA demonstrated that SI of groupl are significantly higher than the SI of group2(P<0.05);the SI of group1 significantly lower than the Aβ1-42 group(P<0.05);The result of Cytokine assay of groups restimulated with corresp antigen is that the ration (IL-4/IFN-γ)of groupl is higher than the Aβ1-42 group(P<0.05);groups restimulated with ConA demonstrate that the concentration of IFN-y:groupl or group2 were significantly lower than Aβ1-42 group(P<0.05).ConclusionThe antigenicity of vaccine pcDNA3.1+-(Aβ3-10) 10-CpG in our research is valid but that lower than Aβ1-42; the immune response induced by the vaccine is maintaly Th2 reaction. Vaccine pcDNA3.1+-(AP3-10) 10-CpG antigen solid and corroborating, but less than Aβ1-42; vaccine-induced responses to Th2 type of response to the main; CpG sequence to enhance the Th2 response mainly, but also a certain degree of strengthening into a Thl response. Experimental results with the experimental expectations, can be as Alzheimer's disease vaccine candidates for trials of genetically modified animals provide the basis for further study of active immune clearance mechanism of senile plaques laid the foundation for the Alzheimer's disease Gene vaccine provides a new means... |
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