表达Aβ3-10基因疫苗免疫阻止β-淀粉样蛋白沉积对Alzheimer病（AD）神经变性的影响

Firstly, the APPswe/Tauvlw double transgenic mice will be immunized prior to Aβ deposits with our newly constructed vaccine which can clear the Aβ deposits without inducing inflammatory reaction within the brain, in order to keep there is no Aβ deposit in the brain from the beginning to the end. Secondly, the same kind of mice will be immunized at different time point from the very beginning of Aβ deposit to the time when Aβ plaques obviously deposit with severe cognitive impairment occurring, in order to clear Aβ from different phase. Then based on the results from changes of structure and function of synapse and brain cells, Aβdeposit, tau protein phosphorylation, changes of neurofibrillary tangles and the behavioural changes of the transgenic mice, it will be aimed to discuss whether Aβis the primary pathogenic event, whether the Amyloid Cascade Hypothesis is true or not, more importantly to illustrate whether immunotherapy is effective or not, then to make sure whether neurodegeneration is initiated by Aβ and when the starting point is, to make clear that at what time immunization can prevent or obviously delay neurodegeneration, providing further evidence for immunotherapy of AD; at the same time through the detection of Aβ specific antigen in the serum of the transgenic mice immunized with our newly constructed vaccine at different stages, to discuss the existing duration of the anti- Aβ specific antigen, then to provide important evidence for further research on prevention and treatment of AD with our newly constructed vaccine.

首先，在Aβ开始沉积之前以我们构建的能够清除Aβ沉积而不引起炎症反应的疫苗来免疫APPswe/Tauvlw双转基因鼠，使脑内始终不发生Aβ沉积；另一方面，从Aβ刚刚开始沉积到明显沉积并出现认知功能损害之间的不同时间点来免疫上述转基因鼠, 目的在于从病程的不同阶段清除已沉积的Aβ等两方面入手，进而通过神经突触结构及功能变化、脑细胞的结构与数量变化、Aβ沉积、tau蛋白磷酸化、神经元纤维缠结及转基因鼠行为学变化等研究，来探讨Aβ是否为AD发病的核心因素，更重要的是阐明免疫治疗AD是否有效，在此基础上确定Aβ是否启动神经变性及其启动点，明确何时免疫能够阻止或者延缓神经变性，为免疫治疗AD提供进一步的理论依据；与此同时通过不同阶段接受我们新构建的疫苗免疫后的转基因鼠血浆中Aβ特异性抗体的检测，来探讨Aβ特异性抗体的存在时间，为我们新构建的疫苗对AD的进一步防治研究提供重要依据。

选用Aβ1-42 的N 末端片段Aβ3-10 为抗原，为提高其免疫原性再偶联KLH制成Aβ3-10 -KLH抗原，以该抗原免疫APP/PS1转基因鼠后获得高浓度抗体，且避开了与炎症反应相关的T-细胞反应，表现为TH2反应；免疫后明显减少APP/PS1转基因鼠脑内老年斑的形成，海马和皮层分别减少80.77% 和 81.86%；改善鼠的学习记忆功能，经突触素、突触膜NR2B免疫组织化学及突触的电子显微镜观察研究显示对突触有明显保护作用，且突触的保护作用具持续性，均明显优于Aβ1-42免疫，该优势可能与可溶性Aβ有关。以该疫苗再免疫三转基因AD小鼠（3×Tg-AD）后经免疫组织化学、ELISA等检测表明tau蛋白的磷酸化明显减少，与可溶性Aβ1-X减少相一致，进一步揭示可溶性Aβ而非老年斑在AD发病中的重要性。