Relationship between intracellular pH, metabolic co-factors and caspase-3 activation in cancer cells during apoptosis

• Published in: Biochimica et biophysica acta. Molecular cell research Vol. 1864; no. 3; pp. 604 - 611
• Main Authors: Sergeeva, Tatiana F., Shirmanova, Marina V., Zlobovskaya, Olga A., Gavrina, Alena I., Dudenkova, Varvara V., Lukina, Maria M., Lukyanov, Konstantin A., Zagaynova, Elena V.
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.03.2017
• Subjects: Animals; Antineoplastic Agents - pharmacology; Apoptosis; Apoptosis - drug effects; Apoptosis - genetics; Caspase 3 - genetics; Caspase 3 - metabolism; Caspase-3; Cell Line, Tumor; Coumarins - chemistry; Enzyme Activation - drug effects; Epithelial Cells - cytology; Epithelial Cells - drug effects; Epithelial Cells - metabolism; FAD; Flavin-Adenine Dinucleotide - metabolism; Fluorescence Resonance Energy Transfer; Gene Expression Regulation; Genes, Reporter; Glycolysis - drug effects; Hydrogen-Ion Concentration; Intracellular pH; Mice; Microscopy, Fluorescence, Multiphoton; Molecular Probes - chemistry; NAD(P)H; NADP - metabolism; Oxidative Phosphorylation - drug effects; Signal Transduction; Staurosporine - pharmacology; pHi; Intracellular pH; NHE1; STS; ROS; FAD; OXPHOS; FRET; NAD(P)H; FLIM; Caspase-3; Apoptosis
• ISSN: 0167-4889; 1879-2596
• DOI: 10.1016/j.bbamcr.2016.12.022

A complex cascade of molecular events occurs in apoptotic cells but cell-to-cell variability significantly complicates determination of the order and interconnections between different processes. For better understanding of the mechanisms of programmed cell death, dynamic simultaneous registration of several parameters is required. In this paper we used multiparameter fluorescence microscopy to analyze energy metabolism, intracellular pH and caspase-3 activation in living cancer cells in vitro during staurosporine-induced apoptosis. We performed metabolic imaging of two co-factors, NAD(P)H and FAD, and used the genetically encoded pH-indicator SypHer1 and the FRET-based sensor for caspase-3 activity, mKate2-DEVD-iRFP, to visualize these parameters by confocal fluorescence microscopy and two-photon fluorescence lifetime imaging microscopy. The correlation between energy metabolism, intracellular pH and caspase-3 activation and their dynamic changes were studied in CT26 cancer cells during apoptosis. Induction of apoptosis was accompanied by a switch to oxidative phosphorylation, cytosol acidification and caspase-3 activation. We showed that alterations in cytosolic pH and the activation of oxidative phosphorylation are relatively early events associated with the induction of apoptosis. •Multiparameter fluorescence microscopy is applied for apoptosis study in cancer cells.•Genetically encoded sensors are used for pHi and caspase-3 activation imaging.•Metabolic co-factors, pHi and caspase-3 activity are visualized in apoptotic cells.•Dynamic changes in metabolism, pHi and caspase-3 activity are shown in apoptosis.