Yu Qing Cissy, Song Wenqian, Lai Dengwen, Zeng Yi Arial
State Key Laboratory of Cell Biology, CAS Center for Excellence in Molecular Cell Science, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences;
State Key Laboratory of Cell Biology, CAS Center for Excellence in Molecular Cell Science, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences.
J Vis Exp. 2017 Aug 3(126):55795. doi: 10.3791/55795.
Endothelial cells (ECs) are the fundamental building blocks of the vascular architecture and mediate vascular growth and remodeling to ensure proper vessel development and homeostasis. However, studies on endothelial lineage hierarchy remain elusive due to the lack of tools to gain access as well as to directly evaluate their behavior in vivo. To address this shortcoming, a new tissue model to study angiogenesis using the mammary fat pad has been developed. The mammary gland develops mostly in the postnatal stages, including puberty and pregnancy, during which robust epithelium proliferation is accompanied by extensive vascular remodeling. Mammary fat pads provide space, matrix, and rich angiogenic stimuli from the growing mammary epithelium. Furthermore, mammary fat pads are located outside the peritoneal cavity, making them an easily accessible grafting site for assessing the angiogenic potential of exogenous cells. This work also describes an efficient tracing approach using fluorescent reporter mice to specifically label the targeted population of vascular endothelial stem cells (VESCs) in vivo. This lineage tracing method, coupled with subsequent tissue whole-mount microscopy, enable the direct visualization of targeted cells and their descendants, through which the proliferation capability can be quantified and the differentiation commitment can be fate-mapped. Using these methods, a population of bipotent protein C receptor (Procr) expressing VESCs has recently been identified in multiple vascular systems. Procr VESCs, giving rise to both new ECs and pericytes, actively contribute to angiogenesis during development, homeostasis, and injury repair. Overall, this manuscript describes a new mammary fat pad transplantation and in vivo lineage tracing techniques that can be used to evaluate the stem cell properties of VESCs.
内皮细胞（ECs）是血管结构的基本组成部分，介导血管生长和重塑，以确保血管正常发育和内环境稳定。然而，由于缺乏在体内获取并直接评估其行为的工具，关于内皮细胞谱系层次的研究仍然难以捉摸。为了解决这一缺点，已开发出一种利用乳腺脂肪垫研究血管生成的新组织模型。乳腺主要在出生后阶段发育，包括青春期和妊娠期，在此期间，旺盛的上皮细胞增殖伴随着广泛的血管重塑。乳腺脂肪垫为生长中的乳腺上皮提供空间、基质和丰富的血管生成刺激。此外，乳腺脂肪垫位于腹腔外，使其成为评估外源性细胞血管生成潜力的易于获取的移植部位。这项工作还描述了一种高效的追踪方法，使用荧光报告基因小鼠在体内特异性标记血管内皮干细胞（VESCs）的目标群体。这种谱系追踪方法，结合随后的组织整装显微镜检查，能够直接观察目标细胞及其后代，通过这种方法可以量化增殖能力并绘制分化命运图谱。使用这些方法，最近在多个血管系统中鉴定出一群表达双能蛋白C受体（Procr）的VESCs。Procr VESCs既能产生新的内皮细胞，也能产生周细胞，在发育、内环境稳定和损伤修复过程中积极促进血管生成。总体而言，本文描述了一种新的乳腺脂肪垫移植和体内谱系追踪技术，可用于评估VESCs的干细胞特性。
