核定位人源α-突触核蛋白转基因小鼠的建立  

Establishment of the human α-synuclein nuclear localizationsignal transgenic mice

作　　者：韦梦晨 范胜涛[1] 吴海婷 张艺薇 王子鸥 黄璋琼[1] WEI Mengchen;FAN Shengtao;WU Haiting;ZHANG Yiwei;WANG Ziou;HUANG Zhangqiong(Institute of Medical Biology,Chinese Academy of Medical Sciences and Peking Union Medical College,Kunming 650118,China)

机构地区：[1]北京协和医学院中国医学科学院医学生物学研究所,昆明650118

出　　处：《中国实验动物学报》2024年第3期307-316,共10页Acta Laboratorium Animalis Scientia Sinica

基　　金：云南省技术创新人才培养对象项目(202105AD160018,202005AD160006);中国医学科学院医学与健康科技创新工程项目(2021-I2M-1-043);国家自然科学基金联合基金项目(U2202214);科技创新2030-重大专项(2023ZD0406306)。

摘　　要：目的 建立一种核定位人源α-突触核蛋白(α-synuclein, α-syn)转基因小鼠,并探究人源α-syn核输入对小鼠行为的影响。方法 构建hSNCA-NLS和EGFP的慢病毒载体,利用显微注射法建立转基因小鼠模型,通过PCR和Western Blot方法鉴定转基因首建鼠及其子代基因型和蛋白表达情况。采用免疫荧光鉴定人源α-syn在小鼠脑组织中的共定位情况,同时采用旷场、转棒、O迷宫实验评价转基因小鼠的行为改变情况。结果 hSNCA-NLS基因成功插入小鼠基因组中,人源α-syn成功表达,并且有明显的核定位现象。进一步研究发现,核定位人源α-syn转基因小鼠在2月龄时开始表现出了运动功能障碍,发生了星形胶质细胞增生和炎症反应,于9月龄时表现出了明显的焦虑样症状,γ-氨基丁酸(GABA)基因表达减少,这些表现一直持续到小鼠12月龄。结论 成功构建了核定位人源α-syn转基因小鼠,小鼠表现出明显的运动功能障碍和焦虑样症状。该模型的成功建立可以为研究α-syn核定位现象在帕金森病中的作用提供一定的研究基础。Objective To establish a humanα-synuclein nuclear localization signal transgenic mouse model and investigate the effects ofα-synuclein nuclear localization on the behavior of mice.Methods Humanα-synuclein nuclear localization signal and EGFP lentiviral vectors were constructed.Transgenic mice were created with the microinjection method.Using PCR and Western Blot method to identify the genotypes and protein expression of the transgenic founder mice and their offsprings.The immunofluorescence was used to examine the localization of humanα-synuclein in the mouse brain tissue.The behavioral changes of the transgenic mice were evaluated by the open field test,rotarod test,and O maze test.Results The h SNCA-NLS gene was successfully inserted into the mouse genome,the humanα-syn was successfully expressed,and the humanα-syn has localized with the nuclear.Further studies found that humanα-synuclein nuclear localization signal transgenic mice had significant motor dysfunction,astrocyte proliferation and inflammatory response at 2 months of age and exhibited significant anxiety-like symptoms and reduced expression of theγ-aminobutyric acid(GABA)gene at 9 months of age,which persisted until 12 months of age.Conclusions A humanα-synuclein nuclear localization signal transgenic mouse model has been successfully established.The mice exhibit significant motor dysfunction and anxiety-like symptoms.The successful establishment of this model provides a foundation for studying the role ofα-syn nuclear localization in Parkinson’s disease.

关 键 词：人源α-突触核蛋白 小鼠 核定位 转基因 运动功能障碍 

分 类 号：Q95-33[生物学—动物学]