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Etoposide sensitizes neuroblastoma cells expressing caspase 8 to TRAIL.

Kim HR, Lee MW, Kim DS, Jo HY, Lee SH, Chueh HW, Jung HL, Yoo KH, Sung KW, Koo HH - Cell Biol Int Rep (2010) (2012)

Bottom Line: Although TRAIL-R2 expression increased in IMR-32 cells in response to etoposide treatment, cell death was not increased by concurrent treatment with TRAIL compared with etoposide alone, because the cells lacked caspase 8 expression.Restoration of caspase 8 expression by exposure to IFNγ (interferon γ) sensitizes IMR-32 cells to TRAIL.Moreover, pretreatment with etoposide increased TRAIL-induced apoptosis in caspase 8 restored IMR-32 cells through activation of a caspase cascade that included caspases 8, 9 and 3.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatrics, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea.

ABSTRACT
TRAIL [TNF (tumour necrosis factor)-related apoptosis-inducing ligand] is a promising agent for clinical use since it kills a wide range of tumour cells without affecting normal cells. We provide evidence that pretreatment with etoposide significantly enhanced TRAIL-mediated apoptosis via up-regulation of DR5 (death receptor 5 or TRAIL-R2) expression in the caspase 8 expressing neuroblastoma cell line, SK-N-MC. In addition, sequential treatment with etoposide and TRAIL increased caspases 8, 9 and 3 activation, Mcl-1 cleavage and Bid truncation, which suggests that the ability of etoposide and TRAIL to induce apoptosis is mediated through activation of an intrinsic signalling pathway. Although TRAIL-R2 expression increased in IMR-32 cells in response to etoposide treatment, cell death was not increased by concurrent treatment with TRAIL compared with etoposide alone, because the cells lacked caspase 8 expression. Restoration of caspase 8 expression by exposure to IFNγ (interferon γ) sensitizes IMR-32 cells to TRAIL. Moreover, pretreatment with etoposide increased TRAIL-induced apoptosis in caspase 8 restored IMR-32 cells through activation of a caspase cascade that included caspases 8, 9 and 3. These results indicate that the etoposide-mediated sensitization of neuroblastoma cells to TRAIL is associated with an increase in TRAIL-R2 expression and requires caspase 8 expression. These observations support the potential use of a combination of etoposide and TRAIL in future clinical trials.

No MeSH data available.


Related in: MedlinePlus

Etoposide treatment induces cell death and increases expression of TRAIL-R2 in neuroblastoma cell linesIMR-32 and SK-N-MC cells were plated in 96-well plates and either (a) treated for 48 h with the indicated concentrations of etoposide or (b) exposed to 500 ng/ml etoposide for 24, 48 or 72 h. Cell viability was assessed using the Alamar Blue assay, which quantifies mitochondrial activity. The results are presented as cell death percentage against vehicle-treated control (means±S.D. from three independent experiments). (c) Surface expression of TRAIL-R1, TRAIL-R2, TRAIL-R3 and TRAIL-R4 in untreated cells (solid line) or cells treated for 6, 12 or 24 h with 500 ng/ml etoposide (broken line) was analysed by flow cytometry using receptor specific antibodies. Control antibody staining appears as shaded peaks.
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Figure 1: Etoposide treatment induces cell death and increases expression of TRAIL-R2 in neuroblastoma cell linesIMR-32 and SK-N-MC cells were plated in 96-well plates and either (a) treated for 48 h with the indicated concentrations of etoposide or (b) exposed to 500 ng/ml etoposide for 24, 48 or 72 h. Cell viability was assessed using the Alamar Blue assay, which quantifies mitochondrial activity. The results are presented as cell death percentage against vehicle-treated control (means±S.D. from three independent experiments). (c) Surface expression of TRAIL-R1, TRAIL-R2, TRAIL-R3 and TRAIL-R4 in untreated cells (solid line) or cells treated for 6, 12 or 24 h with 500 ng/ml etoposide (broken line) was analysed by flow cytometry using receptor specific antibodies. Control antibody staining appears as shaded peaks.

Mentions: Etoposide treatment induced cell death in both IMR-32 and SK-N-MC cells in a dose- and time-dependent manner (Figures 1a and 1b). To explore the effect of etoposide treatment on the expression of TRAIL receptors in neuroblastoma cells, the expression levels of TRAIL receptors using flow cytometry were analysed. The number of TRAIL-R2 expressing cells gradually increased over 24 h of exposure to etoposide, while the number of cells expressing other TRAIL receptors slightly increased after 24 h treatment (Figure 1c).


Etoposide sensitizes neuroblastoma cells expressing caspase 8 to TRAIL.

Kim HR, Lee MW, Kim DS, Jo HY, Lee SH, Chueh HW, Jung HL, Yoo KH, Sung KW, Koo HH - Cell Biol Int Rep (2010) (2012)

Etoposide treatment induces cell death and increases expression of TRAIL-R2 in neuroblastoma cell linesIMR-32 and SK-N-MC cells were plated in 96-well plates and either (a) treated for 48 h with the indicated concentrations of etoposide or (b) exposed to 500 ng/ml etoposide for 24, 48 or 72 h. Cell viability was assessed using the Alamar Blue assay, which quantifies mitochondrial activity. The results are presented as cell death percentage against vehicle-treated control (means±S.D. from three independent experiments). (c) Surface expression of TRAIL-R1, TRAIL-R2, TRAIL-R3 and TRAIL-R4 in untreated cells (solid line) or cells treated for 6, 12 or 24 h with 500 ng/ml etoposide (broken line) was analysed by flow cytometry using receptor specific antibodies. Control antibody staining appears as shaded peaks.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
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getmorefigures.php?uid=PMC3475444&req=5

Figure 1: Etoposide treatment induces cell death and increases expression of TRAIL-R2 in neuroblastoma cell linesIMR-32 and SK-N-MC cells were plated in 96-well plates and either (a) treated for 48 h with the indicated concentrations of etoposide or (b) exposed to 500 ng/ml etoposide for 24, 48 or 72 h. Cell viability was assessed using the Alamar Blue assay, which quantifies mitochondrial activity. The results are presented as cell death percentage against vehicle-treated control (means±S.D. from three independent experiments). (c) Surface expression of TRAIL-R1, TRAIL-R2, TRAIL-R3 and TRAIL-R4 in untreated cells (solid line) or cells treated for 6, 12 or 24 h with 500 ng/ml etoposide (broken line) was analysed by flow cytometry using receptor specific antibodies. Control antibody staining appears as shaded peaks.
Mentions: Etoposide treatment induced cell death in both IMR-32 and SK-N-MC cells in a dose- and time-dependent manner (Figures 1a and 1b). To explore the effect of etoposide treatment on the expression of TRAIL receptors in neuroblastoma cells, the expression levels of TRAIL receptors using flow cytometry were analysed. The number of TRAIL-R2 expressing cells gradually increased over 24 h of exposure to etoposide, while the number of cells expressing other TRAIL receptors slightly increased after 24 h treatment (Figure 1c).

Bottom Line: Although TRAIL-R2 expression increased in IMR-32 cells in response to etoposide treatment, cell death was not increased by concurrent treatment with TRAIL compared with etoposide alone, because the cells lacked caspase 8 expression.Restoration of caspase 8 expression by exposure to IFNγ (interferon γ) sensitizes IMR-32 cells to TRAIL.Moreover, pretreatment with etoposide increased TRAIL-induced apoptosis in caspase 8 restored IMR-32 cells through activation of a caspase cascade that included caspases 8, 9 and 3.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatrics, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea.

ABSTRACT
TRAIL [TNF (tumour necrosis factor)-related apoptosis-inducing ligand] is a promising agent for clinical use since it kills a wide range of tumour cells without affecting normal cells. We provide evidence that pretreatment with etoposide significantly enhanced TRAIL-mediated apoptosis via up-regulation of DR5 (death receptor 5 or TRAIL-R2) expression in the caspase 8 expressing neuroblastoma cell line, SK-N-MC. In addition, sequential treatment with etoposide and TRAIL increased caspases 8, 9 and 3 activation, Mcl-1 cleavage and Bid truncation, which suggests that the ability of etoposide and TRAIL to induce apoptosis is mediated through activation of an intrinsic signalling pathway. Although TRAIL-R2 expression increased in IMR-32 cells in response to etoposide treatment, cell death was not increased by concurrent treatment with TRAIL compared with etoposide alone, because the cells lacked caspase 8 expression. Restoration of caspase 8 expression by exposure to IFNγ (interferon γ) sensitizes IMR-32 cells to TRAIL. Moreover, pretreatment with etoposide increased TRAIL-induced apoptosis in caspase 8 restored IMR-32 cells through activation of a caspase cascade that included caspases 8, 9 and 3. These results indicate that the etoposide-mediated sensitization of neuroblastoma cells to TRAIL is associated with an increase in TRAIL-R2 expression and requires caspase 8 expression. These observations support the potential use of a combination of etoposide and TRAIL in future clinical trials.

No MeSH data available.


Related in: MedlinePlus