Microbubble potentiated changes in cell permeability and viability

The effect of various combinations of microbubbles and ultrasonic exposure parameters on changes of cell membrane permeability is investigated in an in-vitro cell (KHT-C: murine fibrosarcoma cell line) suspension system. The parameters investigated were peak negative pressure (0.1 to 0.9 MI), frequency (1.0 and 3.5 MHz), exposure time (10 sec to 5 min), and microbubble concentration (Optison, 0 to 10%). The number of cells permeabilised (stained with FITC- dextran, 77 kD) and viable (not stained with propidium iodide) were measured by flow-cytometry. Sonoporation in KHT-C tumour cells was possible. The uptake of FITC-dextran increased with peak negative pressure and exposure time, and decreased with frequency. The addition of microbubbles, induced an approximately four-fold increase in uptake; this effect was strongly dependent on bubble concentration. Cell death was induced by ultrasound alone and enhanced by the addition of microbubbles, especially at lower frequencies. The ratio of cells in which reversible permeability was induced to nonviable cells was calculated as an analogy to a ´therapeutic index´. Though the number of cells permeabilised at 1 MHz is greater than at 3.5 MHz, so is the number of nonviable cells. At 3.5 MHz, bubbles had the effect of increasing sonoporation with less impact on the number of cells killed. This therefore may be an optimum frequency for sonoporation. Future work will continue to characterize the effect of different exposure parameters and bubble combinations; in-vitro and in-vivo model of drug uptake will follow.