Oxidative stress as a necessary factor in room temperature-induced apoptosis of HL-60 cells.
HL-60 cells undergo apoptosis when placed at room temperature (RT) [Shimura et al. (1997) FEBS Lett. 417, 379-384]. We report that superoxide anion radical, one of the reactive oxygen species (ROS), was produced after RT treatment. Affinity blot analysis with a biotinylated YVAD-CHO detected the generation of processed peptides with molecular masses of 15-25 kDa. Activation of such an ICE-like protease was completely abolished by N-acetylcysteine and exogenously expressed Bcl-2, known as antioxidants. We concluded that oxidative stress was a critical factor in the signal cascade of the apoptosis. Western blot analysis and experiments using tetrapeptide inhibitors suggested that caspases-1, -3, -4, -6, and -9 did not have an essential role in the apoptotic cascade. It is interesting that cyclosporin A (CsA) blocked RT-induced apoptosis with an inhibition of cytochrome c release from mitochondria. CsA, however, generated a significant amount of ROS with considerable reduction of mitochondrial membrane potential, implying that oxidative stress was one necessary factor for RT-induced apoptosis. It is also likely that mitochondrial membrane potential and the release of apoptotic factors from cytoplasm are differently regulated. Taken together with the reports that some Burkitt lymphoma cells showed apoptosis when exposed at low temperature followed by rewarming, and that hepatocytes or liver endothelial cells are susceptible to cold-induced apoptosis through the ROS function, we propose that studying the mechanism of RT-induced apoptosis of HL-60 cells may provide a therapeutic strategy for pathological conditions involving ROS, such as neurodegenerative diseases and ischemia.
Department of Intractable Diseases, International Medical Center of Japan, Tokyo.
M
M
Shimura
Y
Y
Osawa
A
A
Yuo
K
K
Hatake
F
F
Takaku
Y
Y
Ishizaka
D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D017209:Apoptosis / Q000502:physiology*; D001710:Biotin / Q000031:analogs & derivatives / Q000494:pharmacology; D061945:Caspase Inhibitors; D020169:Caspases / Q000502:physiology; D016572:Cyclosporine / Q000494:pharmacology; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D003574:Cytochrome c Group / Q000378:metabolism; D010450:Endopeptidases / Q000502:physiology; D019254:Genes, bcl-2; D018922:HL-60 Cells / Q000166:cytology* / Q000187:drug effects; D006801:Humans; D007425:Intracellular Membranes / Q000187:drug effects / Q000378:metabolism; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D008970:Molecular Weight; D009363:Neoplasm Proteins / Q000037:antagonists & inhibitors / Q000502:physiology; D009842:Oligopeptides / Q000494:pharmacology; D018384:Oxidative Stress; D010539:Permeability / Q000187:drug effects; D011480:Protease Inhibitors / Q000494:pharmacology; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000502:physiology; D017382:Reactive Oxygen Species / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000502:physiology; D013482:Superoxide Dismutase / Q000494:pharmacology; D013481:Superoxides / Q000378:metabolism; D013696:Temperature; D051020:bcl-X Protein
D000590:Amino Acid Chloromethyl Ketones; C494811:BCL2L1 protein, human; D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors; D003574:Cytochrome c Group; C098738:N-acetyl-tyrosyl-valyl-alanyl-aspartyl chloromethyl ketone; D009363:Neoplasm Proteins; D009842:Oligopeptides; D011480:Protease Inhibitors; D019253:Proto-Oncogene Proteins c-bcl-2; D017382:Reactive Oxygen Species; D011993:Recombinant Fusion Proteins; C110923:acetyl-valyl-isoleucyl-aspartyl-aldehyde; C108637:aspartyl-glutamyl-valyl-aspartal; D051020:bcl-X Protein; C412038:biotin-tyrosyl-valyl-alanyl-aspartic acid aldehyde; D013481:Superoxides; D001710:Biotin; D016572:Cyclosporine; D013482:Superoxide Dismutase; D010450:Endopeptidases; D020169:Caspases