新生乳鼠侧脑室注射rAAV2/9递送SNCA构建全脑转基因鼠  

Neonatal rAAV2/9 delivery of SNCA to generate whole-brain transgenic mice

作　　者：李国祥 潘玥[1] 胡鹏 杜廷福 马开利[1,2] LI Guoxiang;PAN Yue;HU Peng;DU Tingfu;MA Kaili(Institute of Medical Biology,Chinese Academy of Medical Sciences and Peking Union Medical College,Kunming 650118;China.2.Medical Primate Research Center&Neuroscience Center,Chinese Academy of Medical Sciences and Peking Union Medical College,Beijing 100005)

机构地区：[1]中国医学科学院&北京协和医学院医学生物学研究所药物安全性评价研究中心,昆明650118 [2]中国医学科学院&北京协和医学院医学灵长类研究中心&神经科学中心,北京100005

出　　处：《中国比较医学杂志》2021年第4期91-98,共8页Chinese Journal of Comparative Medicine

基　　金：中国医学科学院医学与健康科技创新工程(2016-I2M-2-001;2016-I2M-1-004)。

摘　　要：目的快速高效地构建一种人源SNCA(hSNCA)的全脑转基因鼠,并初步探究α-突触核蛋白过表达对小鼠中枢神经系统的影响。方法对新生乳鼠进行双侧侧脑室(intracerebroventricular,ICV)注射携带人源SNCA-EGFP或EGFP的重组腺相关病毒2/9(recombinant adeno-associated virus2/9,rAAV2/9)(1.5×10^(13) genome copies(GC)/mL),在2周和3个月龄用免疫荧光及Western blot检测α-突触核蛋白的表达模式及亚细胞定位,并用免疫荧光及免疫组化探究星形胶质细胞、小胶质细胞及病理性α-突触核蛋白的变化。结果用rAAV2/9侧脑室注射的方式成功构建了hSNCA全脑转基因鼠,α-突触核蛋白在整个脑中广泛表达,且趋向于在嗅球、皮层、海马、间脑及中脑中高表达。进一步研究发现,在嗅球、皮层、海马的CA2/3区及小脑的浦肯野细胞中观测到α-突触核蛋白在神经元胞核中表达的现象,α-突触核蛋白过表达引起了胶质增生。此外,在嗅球及大脑皮层检测到α-突触核蛋白Ser129磷酸化(pS129)及聚集。结论快速成功地构建了一种hSNCA全脑转基因小鼠模型,其α-突触核蛋白持久地高表达,并出现胶质增生和病理性α-突触核蛋白表达的现象,为研究α-突触核蛋白的生理功能及其在帕金森病(Parkinson’s disease,PD)中的作用奠定一定的基础。Objective To establish a human SNCA whole-brain transgenic mouse model,and to obtain preliminarily data on the role ofα-synuclein in the central nervous system.Methods rAAV2/9(1×10^(13)genome copies(GC)/mL)carrying either human SNCA-EGFP or EGFP was bilateral injected intracerebroventricularly in mice at postnatal day 0.The expression pattern and subcellular localization ofα-synuclein was examined at 2 weeks and 3 months of age by immunofluorescence and Western blot.Glial profile and pathological changes were analyzed by immunofluorescence and immunohistochemical staining.Results hSNCA transgenic mice were successfully constructed,andα-synuclein was widely expressed throughout the brain,with high expression in the olfactory bulb,cerebral cortex,hippocampus,interbrain and midbrain.Furthermore,nuclearα-synuclein was detected in the olfactory bulb,cerebral cortex,CA2/3 of the hippocampus and Purkinje cells of the cerebellum.Overexpression ofα-synuclein caused the proliferation of astrocytes and microglia.In addition,pS129 and aggregation ofα-synuclein were observed in the olfactory bulb and cerebral cortex.Conclusions hSNCA whole-brain transgenic mouse model was established successfully,with high long-term expression ofα-synuclein and enhanced gliosis andα-synuclein pathology.This model should be useful for studying the physiological function ofα-synuclein and its role in Parkinson’s disease.

关 键 词：α-突触核蛋白 转基因 乳鼠注射 胶质增生 

分 类 号：R-33[医药卫生]