Enhancement of Ultrasonically Induced Cytotoxic Effect by Hematoporphyrin in vitro

Abstract

It was the aim of this study to investigate the ultrasonically induced cytotoxic effect of hematoporphyrin (Hp) on isolated sarcoma 180 (S180) cells and to explore the potential biological mechanism of this action. S180 tumor cells suspended in air-saturated phosphate-buffered saline (pH 7.2) were exposed to ultrasound at 1.6 MHz in a standing wave mode for up to 90 s with and without 100 µg/ml Hp. The intracellular Hp concentration was evaluated to determine the optimum timing of ultrasound exposure after its administration with a fluorescence spectrophotometer based on the standard curve. Cell viability was determined by the trypan blue exclusion test. The morphological changes of S180 cells induced by ultrasonic irradiation were evaluated by scanning electron microscope and transmission electron microscope observation. The participation of lipid peroxidation products in the cell damage process was investigated by the malon aldehyde content test. Our experiments suggested that an incubation time of 45 min after the addition of 100 µg/ml Hp was to be chosen as the best time for ultrasound exposure in vitro. The rate of ultrasonically induced cell damage was enhanced by the increase in ultrasound intensity and exposure time in the presence and absence of Hp. At an ultrasound intensity not less than 3 W/cm² and an irradiation time not less than 60 s, a significant synergistic effect of ultrasound combined with Hp was observed, while no cell damage was observed with 100 µg/ml Hp alone. The malon aldehyde content test showed that the lipid peroxidation level significantly increased after sonodynamic therapy treatment. Scanning electron microscope and transmission electron microscope observation indicated that the changes in cell ultra-structure such as cell membrane destruction, mitochondria swelling and chromatin condensation are important factors that induced cell damage in S180 tumor cells. The results indicate that the biological mechanism might be involved in mediating the killing effect on S180 cells, and the ultra-structural changes may account for cell destruction induced by sonodynamic therapy treatment in our experiment mode.