A fluorescent Tie1 reporter allows monitoring of vascular development and endothelial cell isolation from transgenic mouse embryos
A fluorescent Tie1 reporter allows monitoring of vascular development and endothelial cell isolation from transgenic mouse embryos
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DOI：
10.1096/fj.01-1043com
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发表时间：
2002-11-01
期刊：
影响因子：
4.8
通讯作者：
Alitalo, K
中科院分区：
文献类型：
--
作者：
Iljin, K;Petrova, TV;Alitalo, K
Tie1 是一种内皮受体酪氨酸激酶，对于血管系统的发育和维护至关重要。在这里，我们报告了表达增强型绿色荧光蛋白（EGFP）或由 Zeosin 抗性标记和 EGFP 在小鼠 Tie1 启动子控制下组成的嵌合蛋白的转基因小鼠的产生。活体荧光监测表明，EGFP 报告基因重现了发育中脉管系统中的 Tie1 表达模式，并且使用 EGFP 的流式细胞术可以从转基因小鼠胚胎中分离出基本上纯的表达 Tie1 的内皮细胞。然而，EGFP 和 LacZ 转基因标记在成人脉管系统中强烈下调；与 Tie1-LacZ 敲入基因座不同，启动子在肿瘤新生血管形成过程中不会重新激活，表明 Tie1 基因座中存在额外的调控元件。从妊娠中期开始，Tie1 启动子活性在动脉内皮中比在静脉内皮中更强；在成年小鼠中，在小动脉、毛细血管和淋巴管中观察到启动子活性，表明不同类型的内皮细胞具有显着程度的特异性。我们的结果将 Tie1-Z/EGFP 转基因小鼠确立为研究胚胎血管发育的有用模型和分离原代内皮细胞的便捷来源。
Tie1 is an endothelial receptor tyrosine kinase essential for development and maintenance of the vascular system. Here we report generation of transgenic mice expressing enhanced green fluorescent protein (EGFP) or a chimeric protein consisting of a Zeosin resistance marker and EGFP under the control of mouse Tie1 promoter. Intravital monitoring of fluorescence showed that the EGFP reporter recapitulates the Tie1 expression pattern in the developing vasculature, and flow cytometry using EGFP allowed the isolation of essentially pure Tie1-expressing endothelial cells from transgenic mouse embryos. However, EGFP and LacZ transgenic markers were strongly down-regulated in the adult vasculature; unlike the Tie1-LacZ knock-in locus, the promoter was not reactivated during tumor neovascularization, indicating the presence of additional regulatory elements in the Tie1 locus. Starting at midgestation, Tie1 promoter activity became stronger in the arterial than in the venous endothelium; in adult mice, promoter activity was observed in arterioles, capillaries, and lymphatic vessels, indicating a significant degree of specificity in different types of endothelial cells. Our results establish Tie1-Z/EGFP transgenic mice as a useful model to study embryonic vascular development and a convenient source for the isolation of primary endothelial cells.