MITOCHONDRIAL SPECIFIC ROS HYPEROXIDATION VIA PEROXIREDOXIN III HAS IMPORTANT ROLES ON ARSENIC TRIOXIDE INDUCED APOPTOSIS IN ACUTE PROMYELOCYTIC LEUKEMIA
(Abstract release date: 05/19/16)
EHA Library. Seong C. 06/09/16; 134543; PB1643
Prof. Dr. Chu-Myong Seong
Contributions
Contributions
Abstract
Abstract: PB1643
Type: Publication Only
Background
Despite that the Arsenic trioxide (ATO) is an effective therapeutic drug for acute promyelocytic leukemia (APL), some APL cells are resistant to ATO treatment. Previous studies reported that the apoptotic processes by ATO treatment was due to the accumulation of ROS in the cells and its balance of redox enzymes. However, not only the mechanisms of apoptosis via reactive oxygen species (ROS) and peroxiredoxin (PRX) but its resistance during ATO treatment remain elusive.
Aims
Aims of current study are to elucidate that the upregulation of ROS production and the changes of redox enzyme may be major players of anti-leukemia effect in APL-derived NB4 cells during ATO treatment and to find ways to potentiate of the anti-leukemic effects by maniplating ROS and its redox enzymes.
Methods
NB4, one of the human acute promyelocytic leukemia cell lines, was treated with 2 μM arsenic trioxide to induce apoptosis for 16-48 hours in RPMI-1640 medium supplemented with 10% FBS in CO2 humidified atmosphere at 37°C. Apoptosis was measured by staining with 7-amino-actinomycin D (7-AAD) with flow cytometry. 2, 7-dichlrodihydro-fluorescein-diacetate (H2DCF-DA) and MitoSOX Red were used to detect cellular and mitochondrial levels of ROS. SO2 form for PRX I, PRX II, and PRX III were detected by western blot assay. Steady state level of sulfiredoxin (SRX) and caspase 3, 9 were also studied by western blot analysis. To evaluate of the effect of SRX depletion, NB4 cells were transfected with small interfering RNA (siRNA).
Results
Intracellular ROS of NB4 cells was increased significantly after 16 hour of ATO treatment but decreased after 24 hour of ATO treatment. Mitochondrial ROS of NB4 cells was increased significantly after 39 hour of ATO treatment. Apoptosis of NB4 cell after ATO treatment was increased as time elapsed (24% on 16hr, 26% on 24hr, 48% on 39hr, and 60% on 48hr). Increased cysteine sulfinic acid (Cys–SO2H) of PRX III, oxidized form, was observed as a hyperoxidation reaction in NB4 cells after ATO treatment in concordance with mitochondrial ROS increment of NB4 cells. Increased expressions of cleaved caspase-9 and cleaved caspase-3 were also observed during NB4 cell apoptosis by ATO treatment. Meanwhile, SRX expression was increased in NB4 cells after ATO treatment which was rather unexpected observation. Down regulation of SRX by siRNA promoted ROS generation and apoptosis in ATO-treated NB4 cells.
Conclusion
These findings suggest that ATO-induced anti-leukemic activity can be down regulated by an enhancing oxidized PRX III reduction after ATO-induced SRX activation and that down regulation of SRX can enhance the apoptosis in ATO-treated NB4 cells. This result might provide the insights for finding novel ways in the development of strategies, which may potentiate ATO-induced apoptosis in APL cells.
Session topic: E-poster
Keyword(s): Acute promyelocytic leukemia, Arsenic trioxide, Reactive oxygen species
Type: Publication Only
Background
Despite that the Arsenic trioxide (ATO) is an effective therapeutic drug for acute promyelocytic leukemia (APL), some APL cells are resistant to ATO treatment. Previous studies reported that the apoptotic processes by ATO treatment was due to the accumulation of ROS in the cells and its balance of redox enzymes. However, not only the mechanisms of apoptosis via reactive oxygen species (ROS) and peroxiredoxin (PRX) but its resistance during ATO treatment remain elusive.
Aims
Aims of current study are to elucidate that the upregulation of ROS production and the changes of redox enzyme may be major players of anti-leukemia effect in APL-derived NB4 cells during ATO treatment and to find ways to potentiate of the anti-leukemic effects by maniplating ROS and its redox enzymes.
Methods
NB4, one of the human acute promyelocytic leukemia cell lines, was treated with 2 μM arsenic trioxide to induce apoptosis for 16-48 hours in RPMI-1640 medium supplemented with 10% FBS in CO2 humidified atmosphere at 37°C. Apoptosis was measured by staining with 7-amino-actinomycin D (7-AAD) with flow cytometry. 2, 7-dichlrodihydro-fluorescein-diacetate (H2DCF-DA) and MitoSOX Red were used to detect cellular and mitochondrial levels of ROS. SO2 form for PRX I, PRX II, and PRX III were detected by western blot assay. Steady state level of sulfiredoxin (SRX) and caspase 3, 9 were also studied by western blot analysis. To evaluate of the effect of SRX depletion, NB4 cells were transfected with small interfering RNA (siRNA).
Results
Intracellular ROS of NB4 cells was increased significantly after 16 hour of ATO treatment but decreased after 24 hour of ATO treatment. Mitochondrial ROS of NB4 cells was increased significantly after 39 hour of ATO treatment. Apoptosis of NB4 cell after ATO treatment was increased as time elapsed (24% on 16hr, 26% on 24hr, 48% on 39hr, and 60% on 48hr). Increased cysteine sulfinic acid (Cys–SO2H) of PRX III, oxidized form, was observed as a hyperoxidation reaction in NB4 cells after ATO treatment in concordance with mitochondrial ROS increment of NB4 cells. Increased expressions of cleaved caspase-9 and cleaved caspase-3 were also observed during NB4 cell apoptosis by ATO treatment. Meanwhile, SRX expression was increased in NB4 cells after ATO treatment which was rather unexpected observation. Down regulation of SRX by siRNA promoted ROS generation and apoptosis in ATO-treated NB4 cells.
Conclusion
These findings suggest that ATO-induced anti-leukemic activity can be down regulated by an enhancing oxidized PRX III reduction after ATO-induced SRX activation and that down regulation of SRX can enhance the apoptosis in ATO-treated NB4 cells. This result might provide the insights for finding novel ways in the development of strategies, which may potentiate ATO-induced apoptosis in APL cells.
Session topic: E-poster
Keyword(s): Acute promyelocytic leukemia, Arsenic trioxide, Reactive oxygen species
Abstract: PB1643
Type: Publication Only
Background
Despite that the Arsenic trioxide (ATO) is an effective therapeutic drug for acute promyelocytic leukemia (APL), some APL cells are resistant to ATO treatment. Previous studies reported that the apoptotic processes by ATO treatment was due to the accumulation of ROS in the cells and its balance of redox enzymes. However, not only the mechanisms of apoptosis via reactive oxygen species (ROS) and peroxiredoxin (PRX) but its resistance during ATO treatment remain elusive.
Aims
Aims of current study are to elucidate that the upregulation of ROS production and the changes of redox enzyme may be major players of anti-leukemia effect in APL-derived NB4 cells during ATO treatment and to find ways to potentiate of the anti-leukemic effects by maniplating ROS and its redox enzymes.
Methods
NB4, one of the human acute promyelocytic leukemia cell lines, was treated with 2 μM arsenic trioxide to induce apoptosis for 16-48 hours in RPMI-1640 medium supplemented with 10% FBS in CO2 humidified atmosphere at 37°C. Apoptosis was measured by staining with 7-amino-actinomycin D (7-AAD) with flow cytometry. 2, 7-dichlrodihydro-fluorescein-diacetate (H2DCF-DA) and MitoSOX Red were used to detect cellular and mitochondrial levels of ROS. SO2 form for PRX I, PRX II, and PRX III were detected by western blot assay. Steady state level of sulfiredoxin (SRX) and caspase 3, 9 were also studied by western blot analysis. To evaluate of the effect of SRX depletion, NB4 cells were transfected with small interfering RNA (siRNA).
Results
Intracellular ROS of NB4 cells was increased significantly after 16 hour of ATO treatment but decreased after 24 hour of ATO treatment. Mitochondrial ROS of NB4 cells was increased significantly after 39 hour of ATO treatment. Apoptosis of NB4 cell after ATO treatment was increased as time elapsed (24% on 16hr, 26% on 24hr, 48% on 39hr, and 60% on 48hr). Increased cysteine sulfinic acid (Cys–SO2H) of PRX III, oxidized form, was observed as a hyperoxidation reaction in NB4 cells after ATO treatment in concordance with mitochondrial ROS increment of NB4 cells. Increased expressions of cleaved caspase-9 and cleaved caspase-3 were also observed during NB4 cell apoptosis by ATO treatment. Meanwhile, SRX expression was increased in NB4 cells after ATO treatment which was rather unexpected observation. Down regulation of SRX by siRNA promoted ROS generation and apoptosis in ATO-treated NB4 cells.
Conclusion
These findings suggest that ATO-induced anti-leukemic activity can be down regulated by an enhancing oxidized PRX III reduction after ATO-induced SRX activation and that down regulation of SRX can enhance the apoptosis in ATO-treated NB4 cells. This result might provide the insights for finding novel ways in the development of strategies, which may potentiate ATO-induced apoptosis in APL cells.
Session topic: E-poster
Keyword(s): Acute promyelocytic leukemia, Arsenic trioxide, Reactive oxygen species
Type: Publication Only
Background
Despite that the Arsenic trioxide (ATO) is an effective therapeutic drug for acute promyelocytic leukemia (APL), some APL cells are resistant to ATO treatment. Previous studies reported that the apoptotic processes by ATO treatment was due to the accumulation of ROS in the cells and its balance of redox enzymes. However, not only the mechanisms of apoptosis via reactive oxygen species (ROS) and peroxiredoxin (PRX) but its resistance during ATO treatment remain elusive.
Aims
Aims of current study are to elucidate that the upregulation of ROS production and the changes of redox enzyme may be major players of anti-leukemia effect in APL-derived NB4 cells during ATO treatment and to find ways to potentiate of the anti-leukemic effects by maniplating ROS and its redox enzymes.
Methods
NB4, one of the human acute promyelocytic leukemia cell lines, was treated with 2 μM arsenic trioxide to induce apoptosis for 16-48 hours in RPMI-1640 medium supplemented with 10% FBS in CO2 humidified atmosphere at 37°C. Apoptosis was measured by staining with 7-amino-actinomycin D (7-AAD) with flow cytometry. 2, 7-dichlrodihydro-fluorescein-diacetate (H2DCF-DA) and MitoSOX Red were used to detect cellular and mitochondrial levels of ROS. SO2 form for PRX I, PRX II, and PRX III were detected by western blot assay. Steady state level of sulfiredoxin (SRX) and caspase 3, 9 were also studied by western blot analysis. To evaluate of the effect of SRX depletion, NB4 cells were transfected with small interfering RNA (siRNA).
Results
Intracellular ROS of NB4 cells was increased significantly after 16 hour of ATO treatment but decreased after 24 hour of ATO treatment. Mitochondrial ROS of NB4 cells was increased significantly after 39 hour of ATO treatment. Apoptosis of NB4 cell after ATO treatment was increased as time elapsed (24% on 16hr, 26% on 24hr, 48% on 39hr, and 60% on 48hr). Increased cysteine sulfinic acid (Cys–SO2H) of PRX III, oxidized form, was observed as a hyperoxidation reaction in NB4 cells after ATO treatment in concordance with mitochondrial ROS increment of NB4 cells. Increased expressions of cleaved caspase-9 and cleaved caspase-3 were also observed during NB4 cell apoptosis by ATO treatment. Meanwhile, SRX expression was increased in NB4 cells after ATO treatment which was rather unexpected observation. Down regulation of SRX by siRNA promoted ROS generation and apoptosis in ATO-treated NB4 cells.
Conclusion
These findings suggest that ATO-induced anti-leukemic activity can be down regulated by an enhancing oxidized PRX III reduction after ATO-induced SRX activation and that down regulation of SRX can enhance the apoptosis in ATO-treated NB4 cells. This result might provide the insights for finding novel ways in the development of strategies, which may potentiate ATO-induced apoptosis in APL cells.
Session topic: E-poster
Keyword(s): Acute promyelocytic leukemia, Arsenic trioxide, Reactive oxygen species
{{ help_message }}
{{filter}}