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Involvement of reactive oxygen species in sonodynamically induced apoptosis using a novel porphyrin derivative.

Yumita N, Iwase Y, Nishi K, Komatsu H, Takeda K, Onodera K, Fukai T, Ikeda T, Umemura S, Okudaira K, Momose Y - Theranostics (2012)

Bottom Line: Sonodynamically induced apoptosis, caspase-3 activation, and nitroxide generation were significantly suppressed by histidine.The significant reduction in sonodynamically induced apoptosis, nitroxide generation, and caspase-3 activation by histidine suggests active species such as singlet oxygen are important in the sonodynamic induction of apoptosis.These experimental results support the possibility of sonodynamic treatment for cancer using the induction of apoptosis.

View Article: PubMed Central - PubMed

Affiliation: 1. School of Pharmacy, Yokohama College of Pharmacy, 601, Matano-cho, Totsuka-ku, Yokohama, Kanagawa 245-0066, Japan;

ABSTRACT
In this study, we investigated the induction of apoptosis by ultrasound in the presence of the novel porphyrin derivative DCPH-P-Na(I). HL-60 cells were exposed to ultrasound for up to 3 min in the presence and absence of DCPH-P-Na(I), and the induction of apoptosis was examined by analyzing cell morphology, DNA fragmentation, and caspase-3 activity. Reactive oxygen species were measured by means of ESR and spin trapping technique. Cells treated with 8 μM DCPH-P-Na(I) and ultrasound clearly showed membrane blebbing and cell shrinkage, whereas significant morphologic changes were not observed in cells exposed to either ultrasound or DCPH-P-Na(I) alone. Also, DNA ladder formation and caspase-3 activation were observed in cells treated with both ultrasound and DCPH-P-Na(I) but not in cells treated with ultrasound or DCPH-P-Na(I) alone. In addition, the combination of DCPH-P-Na(I) and the same acoustical arrangement of ultrasound substantially enhanced nitroxide generation by the cells. Sonodynamically induced apoptosis, caspase-3 activation, and nitroxide generation were significantly suppressed by histidine. These results indicate that the combination of ultrasound and DCPH-P-Na(I) induced apoptosis in HL-60 cells. The significant reduction in sonodynamically induced apoptosis, nitroxide generation, and caspase-3 activation by histidine suggests active species such as singlet oxygen are important in the sonodynamic induction of apoptosis. These experimental results support the possibility of sonodynamic treatment for cancer using the induction of apoptosis.

No MeSH data available.


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A) Fraction of HL60 cells not stained with Trypan blue following ultrasonic exposure in the presence and absence of DCPH-P-Na(I). ▲, 8 μM DCPH-P-Na(I) alone; ○, ultrasound alone; ■, 8 μM DCPH-P-Na(I) + ultrasound. Values represent the means ± S.D. of four independent experiments. B) Analysis of cellular morphology by phase contrast microscopy after a 4-h incubation under the following conditions: (a) No treated (b) 8 μM DCPH-P-Na(I) alone, (c) ultrasound alone, and (d) 8 μM DCPH-P-Na(I) + ultrasound. C) Fraction of apoptotic HL-60 cells after a 3-min exposure to ultrasound in the presence and absence of DCPH-P-Na(I).□, control; ■, 8 μM DCPH-P-Na(I) alone; ○, ultrasound alone; ●, 8 μM DCPH-P-Na(I) + ultrasound. Values represent the means ± S.D. of four independent experiments. The asterisk symbol indicates significant difference from untreated control at P < 0.05. D) Apoptotic fraction of HL-60 cells as a function of ultrasonic intensity.○, ultrasound alone; ●, 8 μM DCPH-P-Na(I) + ultrasound. Values represent the means ± S.D. of four independent experiments.
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Figure 3: A) Fraction of HL60 cells not stained with Trypan blue following ultrasonic exposure in the presence and absence of DCPH-P-Na(I). ▲, 8 μM DCPH-P-Na(I) alone; ○, ultrasound alone; ■, 8 μM DCPH-P-Na(I) + ultrasound. Values represent the means ± S.D. of four independent experiments. B) Analysis of cellular morphology by phase contrast microscopy after a 4-h incubation under the following conditions: (a) No treated (b) 8 μM DCPH-P-Na(I) alone, (c) ultrasound alone, and (d) 8 μM DCPH-P-Na(I) + ultrasound. C) Fraction of apoptotic HL-60 cells after a 3-min exposure to ultrasound in the presence and absence of DCPH-P-Na(I).□, control; ■, 8 μM DCPH-P-Na(I) alone; ○, ultrasound alone; ●, 8 μM DCPH-P-Na(I) + ultrasound. Values represent the means ± S.D. of four independent experiments. The asterisk symbol indicates significant difference from untreated control at P < 0.05. D) Apoptotic fraction of HL-60 cells as a function of ultrasonic intensity.○, ultrasound alone; ●, 8 μM DCPH-P-Na(I) + ultrasound. Values represent the means ± S.D. of four independent experiments.

Mentions: We first exposed HL-60 cells to an ultrasound (6 W/cm2) in the presence and absence of 8 μM DCPH-P-Na(I) and examined their integrity by staining with Trypan blue. Figure 3A shows the fraction of unstained HL-60 cells vs. the duration of exposure. The results show that the unstained (intact) fraction decreased exponentially with the duration of exposure. Following exposure to ultrasound for 3 min in the presence and absence of DCPH-P-Na(I), the fraction of unstained cells was 80% and 56%, respectively. DCPH-P-Na(I) alone did not cause cell damage.


Involvement of reactive oxygen species in sonodynamically induced apoptosis using a novel porphyrin derivative.

Yumita N, Iwase Y, Nishi K, Komatsu H, Takeda K, Onodera K, Fukai T, Ikeda T, Umemura S, Okudaira K, Momose Y - Theranostics (2012)

A) Fraction of HL60 cells not stained with Trypan blue following ultrasonic exposure in the presence and absence of DCPH-P-Na(I). ▲, 8 μM DCPH-P-Na(I) alone; ○, ultrasound alone; ■, 8 μM DCPH-P-Na(I) + ultrasound. Values represent the means ± S.D. of four independent experiments. B) Analysis of cellular morphology by phase contrast microscopy after a 4-h incubation under the following conditions: (a) No treated (b) 8 μM DCPH-P-Na(I) alone, (c) ultrasound alone, and (d) 8 μM DCPH-P-Na(I) + ultrasound. C) Fraction of apoptotic HL-60 cells after a 3-min exposure to ultrasound in the presence and absence of DCPH-P-Na(I).□, control; ■, 8 μM DCPH-P-Na(I) alone; ○, ultrasound alone; ●, 8 μM DCPH-P-Na(I) + ultrasound. Values represent the means ± S.D. of four independent experiments. The asterisk symbol indicates significant difference from untreated control at P < 0.05. D) Apoptotic fraction of HL-60 cells as a function of ultrasonic intensity.○, ultrasound alone; ●, 8 μM DCPH-P-Na(I) + ultrasound. Values represent the means ± S.D. of four independent experiments.
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Figure 3: A) Fraction of HL60 cells not stained with Trypan blue following ultrasonic exposure in the presence and absence of DCPH-P-Na(I). ▲, 8 μM DCPH-P-Na(I) alone; ○, ultrasound alone; ■, 8 μM DCPH-P-Na(I) + ultrasound. Values represent the means ± S.D. of four independent experiments. B) Analysis of cellular morphology by phase contrast microscopy after a 4-h incubation under the following conditions: (a) No treated (b) 8 μM DCPH-P-Na(I) alone, (c) ultrasound alone, and (d) 8 μM DCPH-P-Na(I) + ultrasound. C) Fraction of apoptotic HL-60 cells after a 3-min exposure to ultrasound in the presence and absence of DCPH-P-Na(I).□, control; ■, 8 μM DCPH-P-Na(I) alone; ○, ultrasound alone; ●, 8 μM DCPH-P-Na(I) + ultrasound. Values represent the means ± S.D. of four independent experiments. The asterisk symbol indicates significant difference from untreated control at P < 0.05. D) Apoptotic fraction of HL-60 cells as a function of ultrasonic intensity.○, ultrasound alone; ●, 8 μM DCPH-P-Na(I) + ultrasound. Values represent the means ± S.D. of four independent experiments.
Mentions: We first exposed HL-60 cells to an ultrasound (6 W/cm2) in the presence and absence of 8 μM DCPH-P-Na(I) and examined their integrity by staining with Trypan blue. Figure 3A shows the fraction of unstained HL-60 cells vs. the duration of exposure. The results show that the unstained (intact) fraction decreased exponentially with the duration of exposure. Following exposure to ultrasound for 3 min in the presence and absence of DCPH-P-Na(I), the fraction of unstained cells was 80% and 56%, respectively. DCPH-P-Na(I) alone did not cause cell damage.

Bottom Line: Sonodynamically induced apoptosis, caspase-3 activation, and nitroxide generation were significantly suppressed by histidine.The significant reduction in sonodynamically induced apoptosis, nitroxide generation, and caspase-3 activation by histidine suggests active species such as singlet oxygen are important in the sonodynamic induction of apoptosis.These experimental results support the possibility of sonodynamic treatment for cancer using the induction of apoptosis.

View Article: PubMed Central - PubMed

Affiliation: 1. School of Pharmacy, Yokohama College of Pharmacy, 601, Matano-cho, Totsuka-ku, Yokohama, Kanagawa 245-0066, Japan;

ABSTRACT
In this study, we investigated the induction of apoptosis by ultrasound in the presence of the novel porphyrin derivative DCPH-P-Na(I). HL-60 cells were exposed to ultrasound for up to 3 min in the presence and absence of DCPH-P-Na(I), and the induction of apoptosis was examined by analyzing cell morphology, DNA fragmentation, and caspase-3 activity. Reactive oxygen species were measured by means of ESR and spin trapping technique. Cells treated with 8 μM DCPH-P-Na(I) and ultrasound clearly showed membrane blebbing and cell shrinkage, whereas significant morphologic changes were not observed in cells exposed to either ultrasound or DCPH-P-Na(I) alone. Also, DNA ladder formation and caspase-3 activation were observed in cells treated with both ultrasound and DCPH-P-Na(I) but not in cells treated with ultrasound or DCPH-P-Na(I) alone. In addition, the combination of DCPH-P-Na(I) and the same acoustical arrangement of ultrasound substantially enhanced nitroxide generation by the cells. Sonodynamically induced apoptosis, caspase-3 activation, and nitroxide generation were significantly suppressed by histidine. These results indicate that the combination of ultrasound and DCPH-P-Na(I) induced apoptosis in HL-60 cells. The significant reduction in sonodynamically induced apoptosis, nitroxide generation, and caspase-3 activation by histidine suggests active species such as singlet oxygen are important in the sonodynamic induction of apoptosis. These experimental results support the possibility of sonodynamic treatment for cancer using the induction of apoptosis.

No MeSH data available.


Related in: MedlinePlus