Contributed equally to this work and, therefore, share first authorship.
Kinetics of plasma membrane and mitochondrial alterations in cells undergoing apoptosis
Article first published online: 8 MAR 2005
Copyright © 1995 Wiley-Liss, Inc.
Volume 21, Issue 3, pages 275–283, 1 November 1995
How to Cite
Lizard, G., Fournel, S., Genestier, L., Dhedin, N., Chaput, C., Flacher, M., Mutin, M., Panaye, G. and Revillard, J.-P. (1995), Kinetics of plasma membrane and mitochondrial alterations in cells undergoing apoptosis. Cytometry, 21: 275–283. doi: 10.1002/cyto.990210308
- Issue published online: 23 JUN 2005
- Article first published online: 8 MAR 2005
- Manuscript Accepted: 26 APR 1995
- Manuscript Received: 11 AUG 1994
- etoposide (VP-16);
- sodium azide (NaN3);
- rhodamine 123;
- fluorescein diacetate
Programmed cell death or apoptosis is characterized by typical morphological alterations. By transmission electron microscopy, apoptotic cells are identified by condensation of the chromatin in tight apposition to the nuclear envelope, alteration of the nuclear envelope and fragmentation of the nucleus, whereas integrity of the plasma membrane and organelles is preserved. Conversely cells undergoing necrosis display an early desintegration of cytoplasmic membrane and swelling of mitochondria. In this study we assessed by flow cytometry the sequential alterations of forward angle light scatter, 90° light scatter, and fluorescence associated with fluorescein diacetate, rhodamine 123, and propidium iodide in two human B cell lines undergoing apoptosis induced by the topoisomerase II inhibitor VP-16. The kinetics of these modifications were compared to those of cells undergoing necrosis induced by sodium azide. At the same time intervals, cells were examined by transmission electron microscopy and by UV microscopy after staining with Hoechst 33342. We report that sequential changes in light scatters and fluorescein diacetate are similar in cells undergoing apoptosis or necrosis, whereas apoptosis is characterized by a slightly delayed decrease of mitochondrial activity as assessed by rhodamine 123 staining. Surprisingly a part of cells undergoing apoptosis displayed an early uptake of propidium iodide followed by a condensation and then a fragmentation of their nuclei. It is concluded that uptake of propidium iodide is a very early marker of cell death which does not discriminate between necrosis and apoptosis. Along with biochemical criteria, nuclear morphology revealed by staining with Hoechst 33342 would seem to be of the most simple and most discriminative assay of apoptosis. © 1995 Wiley-Liss, Inc.