新生乳鼠侧脑室注射 rAAV2 / 9 递送 SNCA 构建全脑转基因鼠
作者:
作者单位:

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

中图分类号:

R-33


Neonatal rAAV2 / 9 delivery of SNCA to generate whole-bRain transgenic mice
Author:
Affiliation:

1.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

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    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-EGFPEGFP 的重组腺相关病毒 2 / 9 ( recombinant adeno-associated virus2 / 9,rAAV2 / 9) ( 1. 5 × 1013 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。

    Abstract:

    Objective To establish a human SNCA whole-bRain transgenic mouse model, and to obtain preliminarily data on the role of α-synuclein in the central nervous system. Methods rAAV2 / 9 (1 × 1013 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 of α-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, and α-synuclein was widely expressed throughout the brain, with high expression in the olfactory bulb, cerebral cortex, hippocampus, interbrain and midbrain. Furthermore, nuclear α-synuclein was detected in the olfactory bulb, cerebral cortex, CA2 / 3 of the hippocampus and Purkinje cells of the cerebellum. Overexpression of α-synuclein caused the proliferation of astrocytes and microglia. In addition, pS129 and aggregation of α-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 of α-synuclein and enhanced gliosis and α-synuclein pathology. This model should be useful for studying the physiological function of α-synuclein and its role in Parkinson’s disease.

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李国祥,潘 玥,胡 鹏,杜廷福,马开利.新生乳鼠侧脑室注射 rAAV2 / 9 递送 SNCA 构建全脑转基因鼠[J].中国比较医学杂志,2021,31(4):91~98.

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  • 收稿日期:2020-11-24
  • 在线发布日期: 2021-05-28
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