title: Stage-Specific Changes in the Water, Na+, Cl- and K+ Contents of Organelles during Apoptosis, Demonstrated by a Targeted Cryo Correlative Analytical Approach.
abstract: Many studies have demonstrated changes in the levels of several ions during apoptosis, but a few recent studies have reported conflicting results concerning the changes in water content in apoptotic cells. We used a correlative light and cryo-scanning transmission electron microscopy method to quantify water and ion/element contents simultaneously at a nanoscale resolution in the various compartments of cells, from the onset to the end of apoptosis. We used stably transfected HeLa cells producing H2B-GFP to identify the stages of apoptosis in cells and for a targeted elemental analysis within condensed chromatin, nucleoplasm, mitochondria and the cytosol. We found that the compartments of apoptotic cells contained, on average, 10% more water than control cells. During mitochondrial outer membrane permeabilization, we observed a strong increase in the Na+ and Cl- contents of the mitochondria and a strong decrease in mitochondrial K+ content. During the first step in apoptotic volume decrease (AVD), Na+ and Cl- levels decreased in all cell compartments, but remained higher than those in control cells. Conversely, during the second step of AVD, Na+ and Cl- levels increased considerably in the nucleus and mitochondria. During these two steps of AVD, K+ content decreased steadily in all cell compartments. We also determined in vivo ion status during caspase-3 activity and chromatin condensation. Finally, we found that actinomycin D-tolerant cells had water and K+ contents similar to those of cells entering apoptosis but lower Na+ and Cl- contents than both cells entering apoptosis and control cells.
authors: Laboratoire de Recherche en Nanosciences, EA 4682, Université de Reims Champagne Ardenne, Reims, France.
authors: Frédérique
authors: F
authors: Nolin
authors: Laboratoire de Recherche en Nanosciences, EA 4682, Université de Reims Champagne Ardenne, Reims, France.
authors: Jean
authors: J
authors: Michel
authors: Laboratoire de Recherche en Nanosciences, EA 4682, Université de Reims Champagne Ardenne, Reims, France.
authors: Laurence
authors: L
authors: Wortham
authors: CNRS UMR 7369, Université de Reims Champagne Ardenne, Reims, France.
authors: Pavel
authors: P
authors: Tchelidze
authors: Laboratoire de Recherche en Nanosciences, EA 4682, Université de Reims Champagne Ardenne, Reims, France.
authors: Vincent
authors: V
authors: Banchet
authors: CNRS UMR 7369, Université de Reims Champagne Ardenne, Reims, France.
authors: Nathalie
authors: N
authors: Lalun
authors: Platform of Cellular and Tissular Imaging (PICT), Université de Reims Champagne Ardenne, Reims, France.
authors: Christine
authors: C
authors: Terryn
authors: CNRS UMR 7369, Université de Reims Champagne Ardenne, Reims, France.
authors: Dominique
authors: D
authors: Ploton
mesh_terms: D000838:Anions / Q000032:analysis; D017209:Apoptosis / Q000187:drug effects; D053148:Caspase 3 / Q000032:analysis; D002412:Cations / Q000032:analysis; D002463:Cell Membrane Permeability; D048429:Cell Size; D002712:Chlorides / Q000032:analysis*; D020285:Cryoelectron Microscopy / Q000379:methods*; D045304:Cytochromes c / Q000032:analysis; D003609:Dactinomycin / Q000494:pharmacology; D006367:HeLa Cells; D006801:Humans; D007091:Image Processing, Computer-Assisted; D008855:Microscopy, Electron, Scanning / Q000379:methods; D008928:Mitochondria / Q000737:chemistry; D051336:Mitochondrial Membranes; D036103:Nanotechnology / Q000379:methods*; D015388:Organelles / Q000737:chemistry*; D011064:Poly Adenosine Diphosphate Ribose / Q000032:analysis; D011188:Potassium / Q000032:analysis*; D012964:Sodium / Q000032:analysis*; D013052:Spectrometry, X-Ray Emission / Q000379:methods*; D059008:Time-Lapse Imaging / Q000379:methods*; D014867:Water / Q000032:analysis*
chemical_list: D000838:Anions; D002412:Cations; D002712:Chlorides; D014867:Water; D003609:Dactinomycin; D011064:Poly Adenosine Diphosphate Ribose; D045304:Cytochromes c; D012964:Sodium; C505407:CASP3 protein, human; D053148:Caspase 3; D011188:Potassium