Department of Physiological Chemistry and Metabolism, Graduate School of Medicine, The University of Tokyo, 7-3-1, Hongo, Bunkyo-ku, Tokyo, 113-0033, Japan.
Development. 2011 Nov;138(21):4763-76. doi: 10.1242/dev.068023. Epub 2011 Sep 28.
Angiogenesis is a complex process, which is accomplished by reiteration of modules such as sprouting, elongation and bifurcation, that configures branching vascular networks. However, details of the individual and collective behaviors of vascular endothelial cells (ECs) during angiogenic morphogenesis remain largely unknown. Herein, we established a time-lapse imaging and computer-assisted analysis system that quantitatively characterizes behaviors in sprouting angiogenesis. Surprisingly, ECs moved backwards and forwards, overtaking each other even at the tip, showing an unknown mode of collective cell movement with dynamic 'cell-mixing'. Mosaic analysis, which enabled us to monitor the behavior of individual cells in a multicellular structure, confirmed the 'cell-mixing' phenomenon of ECs that occurs at the whole-cell level. Furthermore, an in vivo EC-tracking analysis revealed evidence of cell-mixing and overtaking at the tip in developing murine retinal vessels. In parametrical analysis, VEGF enhanced tip cell behavior and directed EC migration at the stalk during branch elongation. These movements were counter-regulated by EC-EC interplay via γ-secretase-dependent Dll4-Notch signaling, and might be promoted by EC-mural cell interplay. Finally, multiple regression analysis showed that these molecule-mediated tip cell behaviors and directed EC migration contributed to effective branch elongation. Taken together, our findings provide new insights into the individual and collective EC movements driving angiogenic morphogenesis. The methodology used for this analysis might serve to bridge the gap in our understanding between individual cell behavior and branching morphogenesis.
血管生成是一个复杂的过程，通过不断重复发芽、延伸和分支等模块来完成，从而构建出分支的血管网络。然而，血管内皮细胞（ECs）在血管生成形态发生过程中的个体和集体行为的细节在很大程度上仍然未知。在此，我们建立了一个延时成像和计算机辅助分析系统，该系统可以定量表征发芽血管生成中的行为。令人惊讶的是，EC 细胞会前后移动，甚至在尖端也会超越彼此，表现出一种未知的集体细胞运动模式，具有动态的“细胞混合”。马赛克分析使我们能够监测多细胞结构中单个细胞的行为，证实了 EC 细胞在整个细胞水平上发生的“细胞混合”现象。此外，体内 EC 追踪分析在发育中的鼠视网膜血管中发现了尖端处细胞混合和超越的证据。在参数分析中，VEGF 在分支延伸过程中增强了尖端细胞的行为并指导 EC 迁移到茎部。这些运动通过 EC-EC 相互作用通过 γ-分泌酶依赖性 Dll4-Notch 信号进行反向调节，并且可能受到 EC-壁细胞相互作用的促进。最后，多元回归分析表明，这些分子介导的尖端细胞行为和定向 EC 迁移有助于有效的分支延伸。总的来说，我们的发现为推动血管生成形态发生的个体和集体 EC 运动提供了新的见解。用于这种分析的方法可能有助于弥合我们对单个细胞行为和分支形态发生之间理解的差距。
