Frémont S, Echard A
Institut Pasteur, Paris, France; Centre National de la Recherche Scientifique, Paris, France.
Methods Cell Biol. 2017;137:239-251. doi: 10.1016/bs.mcb.2016.03.028. Epub 2016 May 6.
Cytokinesis is an essential step of cell proliferation leading to the physical separation of the dividing cells. Cytokinesis relies on both large scale and local scale cell shape changes, and terminates with the final abscission cut that requires close apposition of the plasma membrane. While furrow ingression is a prominent feature of the early phase of cytokinesis and is easy to visualize in all models, from dividing eggs to culture cells, the later steps of cytokinesis until abscission can be much more difficult to visualize. One key issue is to combine live-cell imaging over several hours and detailed, structural analysis of the cell shape changes in 3D, in particular at the time of cytokinetic abscission. Here, we describe the methodologies that we recently developed for studying cytokinetic abscission in human culture cells using live-cell phase-contrast microscopy, combined with correlative scanning electron microscopy. This allows us to determine the membrane surface and underlying cytoskeleton of the intercellular bridge with unprecedented precision and to determine the fate of the midbody remnant after abscission.
胞质分裂是细胞增殖的关键步骤，可导致分裂细胞的物理分离。胞质分裂依赖于大规模和局部的细胞形状变化，并以最终的脱离切割结束，这需要质膜紧密贴合。虽然沟的内陷是胞质分裂早期的一个显著特征，并且在所有模型中都很容易观察到，从分裂的卵子到培养细胞，但胞质分裂直到脱离的后期步骤可能更难观察到。一个关键问题是要将数小时的活细胞成像与对细胞形状在三维空间中的变化进行详细的结构分析相结合，特别是在胞质分裂脱离时。在这里，我们描述了我们最近开发的用于研究人类培养细胞中胞质分裂脱离的方法，该方法使用活细胞相差显微镜，并结合相关扫描电子显微镜。这使我们能够以前所未有的精度确定细胞间桥的膜表面和潜在的细胞骨架，并确定脱离后中间体残余物的命运。
