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Abstracts
16. High Energy Shock Waves and 5-Aminolevulinic Acid for Sonodynamic Therapy:
Effects on SK-N-BE and SH-SY5Y Neuroblastoma Cell Lines
Roberto Frairia(1), Loredana Serpe(2), Roberto Canaparo(3), Gian Paolo Zara(3), Mario Eandi(3), Laura Berta(1)
Institution:
1 Dept. of Clinical Pathophysiology
2 Dept. of Drug Science and Technology
3 Dept. of Anatomy, Pharmacology and Forensic Medicine, University of Torino, School of Medicine, Torino, Italy
Device and producing company: Piezoson 100, Richard Wolf
Introduction: An appealing form of treatment for solid tumors is sonodynamic therapy based on the ability of ultrasound to
generate acoustic cavitation and to activate a tumor-localizing sonosensitizer agent (such as porphyrin compounds) like
5-aminolevulinic acid (ALA). High Energy Shock Waves (HESW), generated by a piezoelectric device, are able to induce
acoustic cavitation, which results in a concentration of energy sufficient to generate a sonoluminescent emission, which
is able to cause electronic excitation of porphyrins by energy transfer and to initiate a photochemical process resulting
in cytotoxic reactive oxygen species (ROS). For this reason, we have investigated the ability of HESW to activate ALA in
human neuroblastoma SK-N-BE and SH-SY5Y cell lines.
Methods: SK-N-BE and SH-SY5Y cells were exposed to ALA (50-300 µg/ml) for 24 h and then to HESWs (0.22-
0.43 mJ/mm2; 500-1000 pulses): viable cell growth was determined at days 1, 3 and 7 after HESW treatment. Cell
cytotoxicity was measured with WST-1 proliferation assay and cell death was evaluated by flow cytometric analysis. The
relationship between sonodynamic treatment and production of ROS was evaluated by flow cytometric analysis with
dichlorofluorescein diacetate. Furthermore, mRNA expression of different genes involved in apoptosis through ROS
production was evaluated by quantitative SYBR Green real time RT-PCR, and fluorescence microscopic examination was
carried out to highlight ROS production and cell death.
Results: Sonodynamic treatment was able to induce a significant decrease in cell growth compared to untreated cells at 72
h in both SK-N-BE and SH-SY5Y cells: up to 35% and 50%, respectively. Exposure of ALA pre-incubated cells to HESW
significantly increased ROS production with different onset and extent in SK-N-BE and SH-SY5Y cells, and the apoptotic
rate was significantly increased at 24 h in both cell lines.
Discussion: We have identified different treatment schedules of ALA and HESW to produce the best cytotoxic rate in the
two cell lines studied. These findings are in agreement with our previous reports, indicating that shock waves have a
sudden effect in enhancing cytotoxic activities of compounds defined as sonosensitizers in different cell lines.
Conclusion: Our results show that HESW are able to activate porphyrin compounds in neuroblastoma cell lines by acoustic
cavitation obtaining significant in vitro cytotoxicity through ROS production.
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