Comet assay reveals DNA strand breaks induced by ultrasonic cavitation in vitro

The induction of DNA strand breaks in cultured Chinese hamster ovary (CHO) cells was investigated in suspensions directly exposed to 2.17-MHz ultrasound. Production of hydrogen peroxide, a DNA-damaging sonochemical, by inertial cavitation was enhanced by the use of argon-and-oxygen-bubbled media and cell survival was improved by establishing standing waves and minimizing tube rotation. Viable cells were separated from the suspension after exposure and kept on ice for evaluation with the single-cell gel electrophoresis (comet) assay. With this assay, DNA damage from as little as 2-mumol/L hydrogen peroxide treatment for 30 min could be detected, and cell survival as low as 2-5% after ultrasound exposure was adequate for assay. An ultrasound dose-response trend was noted for increasing pressure amplitude up to 0.82 MPa (free field) and increasing exposure duration up to 4 min. The cells were able to repair some of the strand breaks when warmed to 28 degrees C for 30 min. The effect was not eliminated by addition of catalase, which indicates that the DNA damage was not due to the action of residual H2O2 alone. The results confirm the hypothesis of DNA damage in cells surviving inertial cavitation.