Tmem119-EGFP and Tmem119-CreERT2 transgenic mice for labeling and manipulating microglia
Tmem119-EGFP and Tmem119-CreERT2 transgenic mice for labeling and manipulating microglia
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DOI：
10.1523/eneuro.0448-18.2019
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发表时间：
2019-07-01
期刊：
影响因子：
3.4
通讯作者：
Feng, Guoping
中科院分区：
文献类型：
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作者：
Kaiser, Tobias;Feng, Guoping
小胶质细胞是专门的脑内巨噬细胞，在健康和疾病中具有重要功能。为了提高我们对这些细胞的理解，研究界需要遗传工具来识别和控制它们，使它们与密切相关的细胞类型区分开来。我们已经靶向最近发现的小胶质细胞特异性Tmem 119基因，以产生表达EGFP（JAX#031823）或CreERT 2（JAX#031820）的敲入小鼠，分别用于小胶质细胞的鉴定和操作。基因座的遗传表征和基于qPCR的分析证明了转基因的正确定位和内源性Tmem 119在敲入小鼠模型中的完整表达。免疫荧光分析进一步表明，实质小胶质细胞，而不是其他脑巨噬细胞，是完全和忠实地标记在EGFP-线在不同的时间点的发展。流式细胞术显示EGFP在CD 11b + CD 4510小胶质细胞中高度选择性表达。类似地，使用Cre依赖性报告小鼠系的免疫荧光和流式细胞术分析证明了在施用他莫昔芬后，CreERT 2主要在小胶质细胞中的活性，同时警告在软脑膜细胞中的活性。最后，流式细胞术分析分别揭示Tmem 119-EGFP和Tmem 119-CreERT 2系的血液单核细胞中不存在EGFP表达和CreERT 2的最小活性。这些新的转基因细胞系通过提供目前最特异的遗传标记和对小鼠骨髓腔室中这些细胞的控制来扩展小胶质细胞工具箱。
Microglia are specialized brain-resident macrophages with important functions in health and disease. To improve our understanding of these cells, the research community needs genetic tools to identify and control them in a manner that distinguishes them from closely related cell- types. We have targeted the recently discovered microglia-specific Tmem119 gene to generate knock-in mice expressing EGFP (JAX#031823) or CreERT2 (JAX#031820) for the identification and manipulation of microglia, respectively. Genetic characterization of the locus and qPCR-based analysis demonstrate correct positioning of the transgenes and intact expression of endogenous Tmem119 in the knock-in mouse models. Inununofluorescence analysis further shows that parenchymal microglia, but not other brain macrophages, are completely and faithfully labeled in the EGFP-line at different time points of development. Flow cytometiy indicates highly selective expression of EGFP in CD11b + CD4510 microglia. Similarly, immunofluorescence and flow cytomeliy analyses using a Cre-dependent reporter mouse line demonstrate activity of CreERT2 primarily in microglia upon tamoxifen administration with the caveat of activity in leptomeningeal cells. Finally, flow cytometric analyses reveal absence of EGFP expression and minimal activity of CreERT2 in blood monocytes of the Tmem119-EGFP and Tmem119-CreERT2 lines, respectively. These new transgenic lines extend the microglia toolbox by providing the currently most specific genetic labeling and control over these cells in the myeloid compartment of mice.