Abstract: Objective　To construct AD tree shrew model by double transgenic injection of APPswe/ MAPTp30s in the hippocampus and to evaluate the pathological characteristics of AD. Methods　Experimental animals were randomly divided into a control group and an experimental group, with 6 animals in each group (half male and half female). AAV9-EGFP were injected into the left and right hippocampus of control group tree shrews with 2 μL per side (titer:1×1013 GC/mL) by brain stereotaxic technique.AAV9 vectors expressing APPswe genes were injected into the left hippocampus with 2 μL (titer: 1×1013 GC/mL) and AAV9 vectors expressing MAPTp301s genes were injected into the right hippocampus with 2 μL (titer: 1×1013 GC/mL) of experimental group tree shrews (APPswe/ MAPTp301s) by brain stereotaxic technique. The AD model pathological features were detected using immunofluorescence, immunohistochemistry, HE staining, Bieloschsky silver staining, Tunel staining, and Nissl staining. Serum SOD, MDA, and AchE activities were measured, and relevant protein expression was measured by Western blot. Results　Immunofluorescence confirmed the APPswe/ MAPTp301s was expressed in the experimental group tree shrews (APPswe/MAPTp301s) hippocampus. Compared to the control group (AAV9-EGFP), the experimental group (APPswe/MAPTp301s) showed significant Aβ plaque deposition, reduced neuron count, partial neuronal degeneration and necrosis, disappearance of Nissl body in the hippocampus (P<0.05) and cerebral cortex, activation of glial cells, significant neurofibrillary tangle formation (P<0.01), a significant decrease in serum SOD activity (P< 0.01), a significant increase in AchE enzyme activity (P<0.01), and significantly elevated expression levels of phosphorylated Tau protein (P-Tau181 and P-Tau217) in the olfactory bulb(P<0.01,P< 0.05).Conclusion　The APPswe/MAPTp301s double transgenic AD tree shrew model was successfully constructed, which provided a potential animal model for the pathological mechanism study, drug development and treatment of AD.

Key words: familial Alzheimer’s disease, APP Swedish mutation, MAPTp301s mutation, tree shrew, animal model

摘要： 目的　建立APPswe/MAPTp301s双转基因的阿尔茨海默病(AD)树鼩模型,评价AD病理特征。方法　将实 验动物随机分为对照组和实验组,每组各6只,雌雄各半,通过脑立体定位技术在对照组树鼩(AAV9-EGFP)左右脑 海马中均注射AAV9-EGFP,注射体积为左右脑各2 μL(滴度:1×1013 GC/mL),在实验组树鼩(APPswe/MAPTp301s) 左脑海马注射2 μL AAV9-APPswe(滴度:1×1013 GC/mL),右脑海马注射2 μL AAV9-MAPTp301s(滴度:1×1013 GC/ mL)。利用免疫荧光、免疫组化、HE染色、Bieloschsky镀银染色和尼氏体染色等方法检测AD病理特征,同时检测 血清SOD、MDA、AchE活性,采用Western blot检测相关蛋白表达。结果　免疫荧光显示APPswe/MAPTp301s在实 验组(APPswe/MAPTp301s)海马体内表达。与对照组相比(AAV9-EGFP),实验组(APPswe/MAPTp301s)Aβ斑块沉 积明显、神经元数量减少,部分神经元退化和坏死、海马和大脑皮层中的Nissl体消失,胶质细胞活化、产生明显神 经原纤维缠结(P<0.05,P<0.01)、血清SOD活性显著下降(P<0.01), AchE酶活性显著上升(P<0.01)、嗅球中磷 酸化的Tau蛋白(P-Tau181和P-Tau217)表达水平显著升高(P<0.01,P<0.05)。结论　成功构建APPswe/ MAPTp301s双转基因AD树鼩模型,为AD的病理机制研究、药物开发与治疗提供了一个潜在的动物模型。

关键词: 家族性阿尔茨海默病, APPswe突变, MAPTp301s突变, 树鼩, 动物模型