Collapse of the Inner Mitochondrial Transmembrane Potential Is Not Required for Apoptosis of HL60 Cells

Finucane, Deborah M.; Waterhouse, Nigel J.; Amarante-Mendes, Gustavo P.; Cotter, Thomas G.; Green, Douglas R.

doi:10.1006/excr.1999.4527

Cited by 122 publications

(98 citation statements)

References 72 publications

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“…Furthermore, at the time of cytochrome c release (15 min post-PDT) only a small fraction of the cells receiving one of the lower doses of PDT had lost ∆Ψ m . These data are in agreement with the previous conclusion (Finucane et al, 1999a(Finucane et al, , 1999bGoldstein et al, 2000) that dissipation of ∆Ψ m is not required for the release of cytochrome c from mitochondria. However, the possibility of a transient depolarization prior to cytochrome c release without affecting the overall mitochondrial ∆Ψ m and JC-1 labelling cannot be ruled out.…”

Section: Discussionsupporting

confidence: 93%

“…First, time-course studies have demonstrated that release of cytochrome c can precede membrane depolarization (Yang et al, 1997;Bossy-Wetzel et al, 1998). And second, the release of cytochrome c can occur in the absence of mitochondrial depolarization (Kluck et al, 1997;Eskes et al, 1998;Finucane et al, 1999aFinucane et al, , 1999bGoldstein et al, 2000). Alternative models (Martinou et al, Dissociation of mitochondrial depolarization from cytochrome c release during apoptosis induced by photodynamic therapy 2000) have therefore been proposed that depend on the creation of pores or channels in the outer membrane large enough for the passage of cytochrome c. Pro-apoptotic Bcl-2 family proteins, such as Bax, may participate in the channel formation.…”

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confidence: 99%

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Dissociation of mitochondrial depolarization from cytochrome c release during apoptosis induced by photodynamic therapy

2001

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Photodynamic therapy (PDT) with the phthalocyanine photosensitizer Pc 4 induces rapid apoptosis in mouse lymphoma (LY-R) cells, initiating with the release of cytochrome c from mitochondria. It has been proposed that the opening of the mitochondrial membrane permeability transition pores, which results in the dissipation of the mitochondrial membrane potential (Δψ m ), is essential for the escape of cytochrome c from mitochondria into the cytosol as well as for apoptotic cell death. Therefore, we have assessed the correlation between the loss of Δψ m and the release of cytochrome c following PDT. Treatment of LY-R cells with 300 nM Pc 4 and 60, 90 or 120 mJ/cm 2 of red light resulted in apoptosis of 80–90% of the cells, accompanied by >20-fold elevation in caspase-3-like activity within one h. At all 3 doses of PDT employed here, the majority of the cytochrome c was released from mitochondria at 15 min after irradiation, as determined by an immunohistochemical method. In contrast, the loss of Δψ m following PDT, as monitored by the uptake of JC-1 or Rh-123, depended on the PDT dose and the post-treatment time. In spite of the release of cytochrome c at 15 min after each of the 3 doses, a corresponding loss of Δψ m was observed only for those cells that received the highest dose of PDT. Virtually all cells that received one of the lower doses of PDT (300 nM Pc 4 plus 60 or 90 mJ/cm 2 ) maintained normal Δψ m . Hence, our results support the conclusion that the release of cytochrome c from mitochondria resulting from Pc 4-PDT-induced photodamage is independent of the loss of Δψ m . Therefore, it is important to consider a range of doses of this or other apoptotic stimuli in deciphering the relationship of metabolic responses that contribute to apoptosis. © 2001 Cancer Research Campaign http://www.bjcancer.com

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Section: Discussionsupporting

confidence: 93%

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confidence: 99%

Dissociation of mitochondrial depolarization from cytochrome c release during apoptosis induced by photodynamic therapy

2001

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“…However, our results suggest that DOXinduced apoptosis in HL-60 is independent of the loss of mitochondrial membrane potential (DC m ). Indeed, recent reports show that in HL-60 cells, collapse of DC m is not part of the central apoptotic machinery, and is not required for the release of cytochrome c and AIF and caspase activation (Finucane et al, 1999;Li et al, 2000). Our data resemble those reported for etoposide-induced apoptosis (Hishita et al, 2001) in P39 cells, a myelodysplastic syndrome cell line, in which no change in DC m was detected and caspase-3 was activated by a cytochrome c-independent, but lysosomal enzyme-dependent, pathway.…”

Section: Discussionmentioning

confidence: 99%

Proteinase-3, a serine protease which mediates doxorubicin-induced apoptosis in the HL-60 leukemia cell line, is downregulated in its doxorubicin-resistant variant

Gordon

Rastegar

et al. 2002

Oncogene

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We report here that expression of proteinase 3 (PR3), a serine protease, is down-regulated in the HL60/ADR multidrug resistant variant of the human myelogenous leukemia cell line HL-60, and that down-regulation of PR3 is associated with doxorubicin (DOX) resistance in these cells. To determine whether PR3 is involved in DOX-induced apoptosis in HL-60 cells, and whether its loss causes resistance to DOX, we inhibited PR3 expression by an anti-sense PR3 oligodeoxynucleotide and showed that inhibition of PR3 expression results in a significant reduction in DOX-induced DNA fragmentation and increased resistance to DOX-induced apoptosis. Our results revealed that PR3-mediated DOX-induced apoptosis in HL-60 cells is independent of the loss of mitochondrial membrane potential (DC m ) and activation of the caspase-8 and -9 pathways. Moreover, while PR3 is involved in the cleavage of caspase-3, PR3-mediated DOX-induced DNA fragmentation and apoptosis were not prevented by a specific inhibitor of caspase-3. These data suggest that activation of caspase-3 alone is not sufficient to trigger PR3-mediated DOX-induced apoptosis. Treatment with an anti-PR3 oligomer significantly decreased reactive oxygen species (ROS) generation in cells treated with low concentrations of DOX, revealing a role for PR3 in enhancing production of DOX-induced ROS. Moreover, DOX-induced apoptosis at 0.001 -0.01 mM was only inhibited in HL-60 cells pre-treated with the antioxidant N-acetyl-cysteine in the absence of anti-PR3, revealing that DOX-induced apoptosis in these cells is PR3-and ROS-dependent. Our results show that PR3 is involved in DOX-induced ROS-dependent apoptosis and that its loss is associated with resistance to DOX in HL-60 cells.

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“…In many systems, Bax is required for the release of the mitochondrial intermembrane proteins and the induction of apoptosis (Finucane et al, 1999;Kim et al, 2001;Li et al, 2001). To assess the role of Bax in this pathway in the case of Pc 4-PDT, we first studied the relationship between Bax activation (translocation from cytosol to mitochondria) and an early step of the pathway, cytochrome c release, by immunocytochemistry in MCF-7c3 cells following a PDT dose of 200 nM Pc 4 and 200 mJ cm À1 (LD 90 dose).…”

Section: Bax Is Translocated From the Cytosol To Mitochondria During mentioning

confidence: 99%

“…The membrane insertion and oligomerisation of Bax is essential for the release of cytochrome c and apoptosis, as evidenced by the blockage of apoptosis in Bax mutants that have lost the capacity for mitochondrion insertion due to deletion of the mitochondrion-targeting C-terminus (Nechushtan et al, 1999). In addition, overexpression of Bax protein in mammalian cells results in the induction of apoptosis through the release of cytochrome c and activation of caspases (Rosse et al, 1998;Finucane et al, 1999;Gross et al, 1999), and purified Bax protein is capable of triggering the release of cytochrome c from isolated mitochondria (Xiang et al, 1996;Vander Heiden et al, 1997;Eskes et al, 1998;Jurgensmeier et al, 1998;Marzo et al, 1998;Narita et al, 1998). Although the mechanism by which Bax triggers cytochrome c release is not clear, studies in cell-free systems have shown that Bax interacts either with the mitochondrial permeability transition (PT) pore components, the voltage-dependent anion channel and the adenine nucleotide translocator (Marzo et al, 1998), or with cardiolipin of the outer membrane (Kuwana et al, 2002) to form megachannels that allow the passage of cytochrome c.…”

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confidence: 99%

Bax is essential for mitochondrion-mediated apoptosis but not for cell death caused by photodynamic therapy

et al. 2003

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The role of Bax in the release of cytochrome c from mitochondria and the induction of apoptosis has been demonstrated in many systems. Using immunocytochemical staining, we observed that photodynamic therapy (PDT) with the photosensitiser Pc 4 induced Bax translocation from the cytosol to mitochondria, and the release of cytochrome c from mitochondria as early signalling for the intrinsic pathway of apoptosis in human breast cancer MCF-7c3 cells. To test the role of Bax in apoptosis, MCF-7c3 cells were treated with Bax antisense oligonucleotides, which resulted in as much as a 50% inhibition of PDT-induced apoptosis. In the second approach, Bax-negative human prostate cancer DU-145 cells were studied. Following PDT, the hallmarks of apoptosis, including the release of cytochrome c from mitochondria, loss of mitochondrial membrane potential, caspase activation, and chromatin condensation and fragmentation, were completely blocked in these cells. Restoration of Bax expression in DU-145 cells restored apoptosis, indicating that the resistance of DU-145 cells to PDT-induced apoptosis is due to the lack of Bax rather than to another defect in the apoptotic machinery. However, despite the inhibition of apoptosis, the Bax-negative DU-145 cells were as photosensitive as Bax-replete MCF7c3 cells, as determined by clonogenic assay. Thus, for Pc 4-PDT, the commitment to cell death occurs prior to Bax activation.

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Collapse of the Inner Mitochondrial Transmembrane Potential Is Not Required for Apoptosis of HL60 Cells

Cited by 122 publications

References 72 publications

Dissociation of mitochondrial depolarization from cytochrome c release during apoptosis induced by photodynamic therapy

Dissociation of mitochondrial depolarization from cytochrome c release during apoptosis induced by photodynamic therapy

Proteinase-3, a serine protease which mediates doxorubicin-induced apoptosis in the HL-60 leukemia cell line, is downregulated in its doxorubicin-resistant variant

Bax is essential for mitochondrion-mediated apoptosis but not for cell death caused by photodynamic therapy

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