Yumiko Iwase

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

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.

Sonodynamically-induced cytotoxicity by rose bengal derivative and microbubbles in isolated sarcoma 180 cells

It is known that the combination of ultrasound and sonodynamic sensitizer (SDS) is effective in noninvasive tumor treatment, referred to as sonodynamic therapy (SDT). Microbubbles have been used in ultrasound therapy as well. The purpose of this paper is to clarify the effect of microbubbles on SDT. Sarcoma 180 cells were suspended in air-saturated phosphate-buffered saline and exposed to ultrasound with the SDS rose bengal derivative (RBD) in standing wave mode in the presence and absence of microbubbles [sonazoid (SZ)]. The ultrasonically induced cytotoxicity with RBD and SZ was about 20 times higher than without either, and about 80% of the SZ microbubbles were destructed by ultrasonic exposure in as short as five seconds. Since microbubbles induce significant cytotoxicity even with short duration, low intensity ultrasound, the application of microbubbles in SDT shows promise in anti-tumor treatment.

Anticancer Effects of Functionalized Carbon Nanotubes Combined with Ultrasound Irradiation through Reactive Oxygen Species Generation

The sonodynamically induced antitumor effect of functionalized carbon nanotubes (CNT) combined with ultrasound through reactive oxygen species generation was investigated. Both in vitro and in vivo antitumor effects were tested in combination with ultrasound at 2 MHz. The rate of ultrasonically induced damage on isolated sarcoma 180 cells was enhanced by twice with 80 mM CNT. This enhancement was significantly inhibited by histidine, which may suggest it was mediated by ultrasonically induced oxidation. The coadministraion of 25 mg/kg and ultrasonic exposure suppressed the growth of implanted colon 26 tomors at an intensity with which ultrasound alone showed only a slight antitumor effect. In addition, the combination of CNT and ultrasound substantially enhanced singlet oxygen generation. Sonodynamically induced cell damage and singlet oxygen generation were significantly suppressed by histidine and NaN3. These results indicate singlet oxygen plays an important role in sonodynamically induced antitumor effect. In conclusion, the presented results suggest that CNT is a potential sensitizer for sonodynamic tumor treatment.

Antitumor effect of sonodynamically activated pyrrolidine tris-acid fullerene

In this study, the sonodynamically induced antitumor effect of pyrrolidine tris-acid fullerene (PTF) was investigated. Sonodynamically induced antitumor effects of PTF by focused ultrasound were investigated using isolated sarcoma-180 cells and mice bearing ectopically-implanted colon 26 carcinoma. Cell damage induced by ultrasonic exposure was enhanced by 5-fold in the presence of 80 µM PTF. The combined treatment of ultrasound and PTF suppressed the growth of the implanted colon 26 carcinoma. Ultrasonically induced 2,2,6,6-tetramethyl-4-piperidone-1-oxyl (4oxoTEMPO) production in the presence and absence of PTF was assessed, and it was shown that 80 µM PTF enhanced 4oxoTEMPO production as measured by ESR spectroscopy. Histidine, a reactive oxygen scavenger, significantly reduced cell damage and 4oxoTEMPO generation caused by ultrasonic exposure in the presence of PTF. These results suggest that singlet oxygen is likely to be involved in the ultrasonically induced cell damage enhanced by PTF.

Apoptosis induction by aluminum phthalocyanine tetrasulfonate-based sonodynamic therapy in HL-60 cells

The present study aims to investigate sonodynamically-induced apoptosis using the phthalocyanine, chloroaluminum phthalocyanine tetrasulfonate (AlPcTS). HL-60 cells were exposed to ultrasound for up to 3 min in the absence and presence of AlPcTS. Apoptosis was analyzed by cell morphology, DNA fragmentation, and caspase-3 activity. Electron spin resonance was used to measure reactive oxygen species. The number of apoptotic cells showing membrane blebbing and cell shrinkage after combined treatment (ultrasound and AlPcTS) was significantly higher than following other treatments, including ultrasound alone and AlPcTS alone. Furthermore, DNA ladder formation, caspase-3 activation and enhanced nitroxide generation were observed in cells treated with ultrasound and AlPcTS. Sonodynamically induced apoptosis, caspase-3 activation, and nitroxide generation were significantly suppressed by histidine. The significant reduction by histidine indicated that ultrasonically generated reactive oxygen species, such as singlet oxygen, is an important mediator of sonodynamically-induced apoptosis.

Involvement of Reactive Oxygen Species in Sonodynamically Induced Apoptosis by 4-Formyloximeetylidene-3-Hydroxyl-2-Vinyl-euterio-Porphynyl (IX)-6-7-Diaspartic Acid (ATX-S10)

Background: In this study, we investigated the induction of apoptosis by ultrasound in the presence of the photochemically active chlorine, 4-formyloximeetylidene-3-hydroxyl-2-vinyl-deuterio-porphynyl (IX)-6-7-diaspartic acid (ATX-S10). Methods: HL-60 cells were exposed to ultrasound for up to 3 min in the presence and absence of ATX-S10, and the induction of apoptosis was examined by analyzing cell morphology, DNA fragmentation, and caspase-3 activity. Results: Cells treated with 80μM ATX-S10 and ultrasound clearly showed membrane blebbing and cell shrinkage, whereas significant morphologic changes were not observed in cells exposed to either ultrasound or ATX-S10 alone. Also, DNA ladder formation and caspase-3 activation were observed in cells treated with both ultrasound and ATX-S10 but not in cells treated with ultrasound or ATX-S10 alone. In addition, the combination of ATX-S10 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. Conclusions: These results indicate that the combination of ultrasound and ATX-S10 induces 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. General significance: The results reported in this paper are experimental, but they significantly support the possibility of sonodynamic treatment for cancer using the induction of apoptosis.