Mechanism of dibucaine-induced apoptosis in promyelocytic leukemia cells (HL-60).
Dibucaine, a local anesthetic, inhibited the growth of promyelocytic leukemia cells (HL-60) without inducing arrest of the cell cycle and differentiation to granulocytes. Typical DNA fragmentation and DNA ladder formation were induced in a concentration- and time-dependent manner. The half-maximal concentration of dibucaine required to induce apoptosis was 100 microM. These effects were prevented completely by the pan-caspase inhibitor z-Val-Ala-Asp-(OMe)-fluoromethylketone (z-VAD-fmk), thereby implicating the cysteine aspartase (caspase) cascade in the process. Dibucaine activated various caspases, such as caspase-3, -6, -8, and -9 (-like) activities, but not caspase-1 (-like) activity, and induced mitochondrial membrane depolarization and the release of cytochrome c (Cyt.c) from mitochondria into the cytosol. Processing of pro-caspase-3, -8, and -9 by dibucaine was confirmed by western blot analysis. Bid, a death agonist member of the Bcl-2 family, was processed by caspases following exposure of cells to dibucaine. However, 100 microM dibucaine scarcely inhibited oxidative phosphorylation, but it induced membrane permeability transition in isolated rat liver mitochondria. Taken together, these data suggest that dibucaine induced apoptosis of HL-60 cells through activation of the caspase cascade in conjunction with Cyt.c release induced by a processed product of Bid and depolarization of the mitochondrial membrane potential.
Arita
K
K
Department of Biological Chemistry, Faculty of Agriculture, Yamaguchi University, 753-8515, Yamaguchi, Japan.
Utsumi
T
T
Kato
A
A
Kanno
T
T
Kobuchi
H
H
Inoue
B
B
Akiyama
J
J
Utsumi
K
K
D000779:Anesthetics, Local / Q000494:pharmacology; D017209:Apoptosis; D020169:Caspases / Q000378:metabolism; D002453:Cell Cycle / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D048429:Cell Size / Q000187:drug effects; D003574:Cytochrome c Group / Q000378:metabolism; D053938:DNA Fragmentation / Q000187:drug effects; D003992:Dibucaine / Q000494:pharmacology*; D018922:HL-60 Cells; D006801:Humans; D007425:Intracellular Membranes / Q000187:drug effects / Q000378:metabolism; D015473:Leukemia, Promyelocytic, Acute; D008564:Membrane Potentials / Q000187:drug effects; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism / Q000502:physiology; D008933:Mitochondrial Swelling / Q000187:drug effects; D010084:Oxidation-Reduction / Q000187:drug effects; D010766:Phosphorylation / Q000187:drug effects
D000779:Anesthetics, Local; D003574:Cytochrome c Group; D020169:Caspases; D003992:Dibucaine