Karolinska Institutet, Department of Medical Biochemistry and Biophysics, Division of Vascular Biology, Solnavägen 9, SE171 65, Stockholm, Sweden.
Uppsala University, Dept. Immunology, Genetics and Pathology, Rudbeck Laboratory, Dag Hammarskjölds, väg 20, SE751 85, Uppsala, Sweden.
Sci Rep. 2018 Jul 13;8(1):10672. doi: 10.1038/s41598-018-28770-7.
Establishment of the functional blood vasculature involves extensive cellular rearrangement controlled by growth factors, chemokines and flow-mediated shear forces. To record these highly dynamic processes in mammalians has been technically demanding. Here we apply confocal and wide field time-lapse in vivo microscopy to characterize the remodelling vasculature of the wounded mouse cornea. Using mouse lines with constitutive or inducible endogenous fluorescent reporters, in combination with tracer injections and mosaic genetic recombination, we follow processes of sprouting angiogenesis, sprout fusion, vessel expansion and pruning in vivo, at subcellular resolution. We describe the migratory behaviour of endothelial cells of perfused vessels, in relation to blood flow directionality and vessel identity. Live-imaging following intravascular injection of fluorescent tracers, allowed for recording of VEGFA-induced permeability. Altogether, live-imaging of the remodelling vasculature of inflamed corneas of mice carrying endogenous fluorescent reporters and conditional alleles, constitutes a powerful platform for investigation of cellular behaviour and vessel function.
功能性血管系统的建立涉及受生长因子、趋化因子和流动介导的剪切力控制的广泛细胞重排。在哺乳动物中记录这些高度动态的过程在技术上具有挑战性。在这里，我们应用共聚焦和宽场延时活体显微镜来描述受伤小鼠角膜的血管重塑。使用具有组成型或诱导型内源性荧光报告基因的小鼠系，结合示踪剂注射和马赛克基因重组，我们以亚细胞分辨率在体内跟踪发芽血管生成、芽融合、血管扩张和修剪过程。我们描述了灌注血管内皮细胞的迁移行为，与血流方向性和血管身份有关。荧光示踪剂静脉内注射后的活体成像，允许记录 VEGFA 诱导的通透性。总之，携带内源性荧光报告基因和条件等位基因的炎症性角膜的血管重塑的活体成像，为研究细胞行为和血管功能提供了一个强大的平台。
