Various anticancer drugs cause mitochondrial perturbations in association with apoptosis. Here we investigated the involvement of caspase- and Bcl-2-dependent pathways in doxorubicin-induced mitochondrial perturbations and apoptosis. For this purpose, we set up a novel three-color flow cytometric assay using rhodamine 123, annexin V-allophycocyanin, and propidium iodide to assess the involvement of the mitochondria in apoptosis caused by doxorubicin in the breast cancer cell line MTLn3. Doxorubicin-induced apoptosis was preceded by up-regulation of CD95 and CD95L and a collapse of mitochondrial membrane potential (Atk) occurring prior to phosphatidylserine externalization. This drop in Deltapsi was independent of caspase activity, since benzyloxycarbonyl-Val-Ala-DL-Asp-fluoromethylketone did not inhibit it. Benzyloxycarbonyl-Val-Ala-DLAsp-fluoromethylketone also blocked activation of caspase-8, thus excluding an involvement of the death receptor pathway in Deltapsi dissipation. Furthermore, although overexpression of Bcl-2 in MTLn3 cells inhibited apoptosis, dissipation of Deltapsi was still observed. No decrease in Deltapsi was observed in cells undergoing etoposide-induced apoptosis. Immunofluorescent analysis of Deltapsi and cytochrome c localization on a cell-to-cell basis indicates that the collapse of Deltapsi and cytochrome c release are mutually independent in both normal and Bcl2-overexpressing cells. Together, these data indicate that doxorubicin-induced dissipation of the mitochondrial membrane potential precedes phosphatidylserine externalization and is independent of a caspase- or Bcl2-controlled checkpoint.
机构:
WASHINGTON UNIV, SCH MED, HOWARD HUGHES MED INST, DEPT PATHOL, ST LOUIS, MO 63110 USAWASHINGTON UNIV, SCH MED, HOWARD HUGHES MED INST, DEPT PATHOL, ST LOUIS, MO 63110 USA
Zha, JP
;
Harada, H
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机构:
WASHINGTON UNIV, SCH MED, HOWARD HUGHES MED INST, DEPT PATHOL, ST LOUIS, MO 63110 USAWASHINGTON UNIV, SCH MED, HOWARD HUGHES MED INST, DEPT PATHOL, ST LOUIS, MO 63110 USA
Harada, H
;
Yang, E
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机构:
WASHINGTON UNIV, SCH MED, HOWARD HUGHES MED INST, DEPT PATHOL, ST LOUIS, MO 63110 USAWASHINGTON UNIV, SCH MED, HOWARD HUGHES MED INST, DEPT PATHOL, ST LOUIS, MO 63110 USA
Yang, E
;
Jockel, J
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机构:
WASHINGTON UNIV, SCH MED, HOWARD HUGHES MED INST, DEPT PATHOL, ST LOUIS, MO 63110 USAWASHINGTON UNIV, SCH MED, HOWARD HUGHES MED INST, DEPT PATHOL, ST LOUIS, MO 63110 USA
Jockel, J
;
Korsmeyer, SJ
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机构:
WASHINGTON UNIV, SCH MED, HOWARD HUGHES MED INST, DEPT PATHOL, ST LOUIS, MO 63110 USAWASHINGTON UNIV, SCH MED, HOWARD HUGHES MED INST, DEPT PATHOL, ST LOUIS, MO 63110 USA
机构:
Karolinska Inst, Inst Environm Med, Div Toxicol, S-17177 Stockholm, SwedenKarolinska Inst, Inst Environm Med, Div Toxicol, S-17177 Stockholm, Sweden
Zhivotovsky, B
;
Orrenius, S
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机构:Karolinska Inst, Inst Environm Med, Div Toxicol, S-17177 Stockholm, Sweden
Orrenius, S
;
Brustugun, OT
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机构:Karolinska Inst, Inst Environm Med, Div Toxicol, S-17177 Stockholm, Sweden
Brustugun, OT
;
Doskeland, SO
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机构:Karolinska Inst, Inst Environm Med, Div Toxicol, S-17177 Stockholm, Sweden
机构:
WASHINGTON UNIV, SCH MED, HOWARD HUGHES MED INST, DEPT PATHOL, ST LOUIS, MO 63110 USAWASHINGTON UNIV, SCH MED, HOWARD HUGHES MED INST, DEPT PATHOL, ST LOUIS, MO 63110 USA
Zha, JP
;
Harada, H
论文数: 0引用数: 0
h-index: 0
机构:
WASHINGTON UNIV, SCH MED, HOWARD HUGHES MED INST, DEPT PATHOL, ST LOUIS, MO 63110 USAWASHINGTON UNIV, SCH MED, HOWARD HUGHES MED INST, DEPT PATHOL, ST LOUIS, MO 63110 USA
Harada, H
;
Yang, E
论文数: 0引用数: 0
h-index: 0
机构:
WASHINGTON UNIV, SCH MED, HOWARD HUGHES MED INST, DEPT PATHOL, ST LOUIS, MO 63110 USAWASHINGTON UNIV, SCH MED, HOWARD HUGHES MED INST, DEPT PATHOL, ST LOUIS, MO 63110 USA
Yang, E
;
Jockel, J
论文数: 0引用数: 0
h-index: 0
机构:
WASHINGTON UNIV, SCH MED, HOWARD HUGHES MED INST, DEPT PATHOL, ST LOUIS, MO 63110 USAWASHINGTON UNIV, SCH MED, HOWARD HUGHES MED INST, DEPT PATHOL, ST LOUIS, MO 63110 USA
Jockel, J
;
Korsmeyer, SJ
论文数: 0引用数: 0
h-index: 0
机构:
WASHINGTON UNIV, SCH MED, HOWARD HUGHES MED INST, DEPT PATHOL, ST LOUIS, MO 63110 USAWASHINGTON UNIV, SCH MED, HOWARD HUGHES MED INST, DEPT PATHOL, ST LOUIS, MO 63110 USA
机构:
Karolinska Inst, Inst Environm Med, Div Toxicol, S-17177 Stockholm, SwedenKarolinska Inst, Inst Environm Med, Div Toxicol, S-17177 Stockholm, Sweden
Zhivotovsky, B
;
Orrenius, S
论文数: 0引用数: 0
h-index: 0
机构:Karolinska Inst, Inst Environm Med, Div Toxicol, S-17177 Stockholm, Sweden
Orrenius, S
;
Brustugun, OT
论文数: 0引用数: 0
h-index: 0
机构:Karolinska Inst, Inst Environm Med, Div Toxicol, S-17177 Stockholm, Sweden
Brustugun, OT
;
Doskeland, SO
论文数: 0引用数: 0
h-index: 0
机构:Karolinska Inst, Inst Environm Med, Div Toxicol, S-17177 Stockholm, Sweden