Time sequence analysis of caspase-3-independent programmed cell death and apoptosis in X-irradiated human leukemic MOLT-4 cells.
It has been demonstrated that caspase-3 is responsible for determining the mode of cell death, i.e., apoptosis or necrosis. To characterize the mode of cell death induced by the inhibition of caspase-3, we have studied the effects of Ac-DEVD-CHO, Ac-YVAD-CHO, and Ac-IETD-CHO, inhibitors of caspases, on structural changes in X-irradiated human leukemic MOLT-4 cells. When cells were irradiated with X-rays and incubated in the presence of Ac-DEVD-CHO, the expression of cell death, as measured by the dye exclusion test, was inhibited, whereas no such change was observed in colony-forming ability. The hallmarks of apoptosis, i.e., nuclear condensation and DNA ladder formation, were depressed. However, a new type of nuclear morphology appeared. The sum of the frequencies of apoptosis and this new type of nuclear structure corresponded to the frequency of X-ray-induced apoptosis for cells incubated in the absence of Ac-DEVD-CHO. Removal of Ac-DEVD-CHO during the course of post-irradiation incubation increased apoptotic nuclear condensation accompanied by a slight decrease in the frequency of the new type of nuclear structure. When Ac-IETD-CHO was used in place of Ac-DEVD-CHO, inhibition of cell death (apoptosis) was also observed, but not in the case of Ac-YVAD-CHO. These results suggest that the inhibition of caspase-3 diminishes the expression of apoptotic hallmarks with no effect on cell survival, that the morphology observed in the presence of Ac-DEVD-CHO is an apoptosis-related structure, and that the cell death observed is a programmed cell death independent of caspase-3. The development of this mode of cell death was slower than that of apoptosis by 4 h.
Nakano
Hisako
H
Department of Laboratory Animal Science, Tokyo Metropolitan Institute of Medical Science, 3-18-22 Honkomagome, Bunkyo-ku, Tokyo 113-8613, Japan. nakano@rinshoken.or.jp
Shinohara
Kunio
K
D017209:Apoptosis / Q000528:radiation effects*; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism*; D016923:Cell Death / Q000528:radiation effects; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology*; D006801:Humans; D007700:Kinetics; D007938:Leukemia; D009842:Oligopeptides / Q000494:pharmacology; D013997:Time Factors; D014407:Tumor Cells, Cultured; D014965:X-Rays
D015853:Cysteine Proteinase Inhibitors; D009842:Oligopeptides; C099158:acetyl-aspartyl-glutamyl-valyl-aspartal; C096429:L 709049; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases