Caspase-dependent and independent cell death in rat hepatoma 5123tc cells.
The objective of this study was to establish whether apoptosis in 5123tc rat hepatoma cells required the caspase-3 dependent pathway. Apoptosis was induced by either growth factor deprivation or treatment with a topoisomerase II inhibitor, VM26, in the absence or presence of caspase inhibitors (DEVD-fmk, z-VAD-fmk and BAF). The results indicated that, although these inhibitors at 10 microM concentration completely blocked caspase-3 activity, they had no effect on either the rate of cell death or on any other apoptotic features, e.g., chromatin condensation, DNA fragmentation, protein cleavage, suggesting that caspase-3 was not required to mediate nuclear destruction in these hepatoma cells. At higher concentrations, up to 100 microM, z-VAD-fmk and BAF, but not DEVD-fmk, did block apoptosis, however, they also caused cell swelling and membrane permeabilization, which are the hallmarks of necrotic cell death. Clearly, high concentrations of these inhibitors must have interfered non-specifically with other metabolic pathways, e.g., z-VAD-fmk at a high concentration blocked protein phosphorylation, and caused cell death by a different mechanism.
Apoptosis Research Group, Institute for Biological Sciences, National Research Council of Canada, Ottawa, Ontario. pandey@nrc.ca
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D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D015153:Blotting, Western; D006528:Carcinoma, Hepatocellular; D053148:Caspase 3; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism*; D002467:Cell Nucleus / Q000378:metabolism; D016895:Culture Media, Serum-Free; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology*; D053938:DNA Fragmentation; D016521:Electrophoresis, Gel, Pulsed-Field; D015180:Electrophoresis, Gel, Two-Dimensional; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D034882:Lamins; D008856:Microscopy, Fluorescence; D009687:Nuclear Proteins / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases; D011506:Proteins / Q000378:metabolism; D051381:Rats; D013713:Teniposide / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D000590:Amino Acid Chloromethyl Ketones; D061945:Caspase Inhibitors; D016895:Culture Media, Serum-Free; D015853:Cysteine Proteinase Inhibitors; D004791:Enzyme Inhibitors; D034882:Lamins; D009687:Nuclear Proteins; D011506:Proteins; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; D013713:Teniposide; C499322:Parp1 protein, rat; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases; C505407:CASP3 protein, human; C505410:Casp3 protein, rat; D053148:Caspase 3; D020169:Caspases