Crampton Steve P, Davis Jaeger, Hughes Christopher C W
Department of Molecular Biology and Biochemistry, University of California, Irvine, USA.
J Vis Exp. 2007(3):183. doi: 10.3791/183. Epub 2007 Apr 28.
Angiogenesis is a complex multi-step process, where in response to angiogenic stimuli, new vessels are created from the existing vasculature. These steps include: degradation of the basement membrane, proliferation and migration (sprouting) of endothelial cells (EC) into the extracellular matrix, alignment of EC into cords, lumen formation, anastomosis, and formation of a new basement membrane. Many in vitro assays have been developed to study this process, but most only mimic certain stages of angiogenesis, and morphologically the vessels often do not resemble vessels in vivo. Here we demonstrate an optimized in vitro angiogenesis assay that utilizes human umbilical vein EC and fibroblasts. This model recapitulates all of the key early stages of angiogenesis, and importantly the vessels display patent intercellular lumens surrounded by polarized EC. Vessels can be easily observed by phase-contrast and time-lapse microscopy, and recovered in pure form for downstream applications.
血管生成是一个复杂的多步骤过程，在此过程中，响应血管生成刺激，新血管从现有的脉管系统中形成。这些步骤包括：基底膜降解、内皮细胞（EC）增殖并迁移（发芽）到细胞外基质中、EC排列成索状、管腔形成、吻合以及新基底膜的形成。已经开发了许多体外试验来研究这一过程，但大多数仅模拟血管生成的某些阶段，并且在形态学上这些血管通常与体内的血管不同。在这里，我们展示了一种优化的体外血管生成试验，该试验利用人脐静脉EC和成纤维细胞。该模型概括了血管生成的所有关键早期阶段，重要的是，这些血管显示出由极化的EC包围的开放细胞间管腔。通过相差显微镜和延时显微镜可以轻松观察到血管，并且可以以纯形式回收用于下游应用。
