Quantitative evaluation of the mortality rate of HeLa cells induced by microbubble vibration and collapse under pulsed ultrasound irradiation

Drug delivery systems (DDS) using ultrasound and microbubbles have been proposed to enhance drug efficacy and reduce undesirable side effects. While several researchers have reported the vibrational characteristics of lipid-coated microbubbles under ultrasound irradiation and the effects on cell damage, it is necessary to quantitatively evaluate the effects to establish the safety criteria of ultrasound DDS. In this paper, the effects of the microbubble behavior on adhesive HeLa cells under ultrasound irradiation were evaluated. Microbubbles coated with a phospholipid shell were fabricated and two size distributions of bubbles were prepared. The HeLa cells were exposed to pulsed ultrasound at 1 MHz with microbubbles in a water tank, and the cell mortality rate was measured quantitatively under fluorescent observation. The collapse of the microbubbles with a resonance size of approximately 2 μm enhanced the cell mortality rate. Greater sound pressure amplitude produced a higher collapsing rate of the microbubble and cell mortality rate, implying the destruction of microbubbles with resonance size generated a shock wave or microjet, inducing cell damage. While the cell mortality rate decreased exponentially with the increase in distance between cells and microbubbles, a smaller distance induced a higher cell mortality rate: 84% of the HeLa cells died within 2.5 μm of microbubbles produced by a pulsed ultrasound with 1.0 MPa at 1 MHz.