High intensity focused ultrasound-induced gene activation in sublethally injured tumor cells in vitro

Cultured human cervical cancer (HeLa) and rat mammary carcinoma (R3230Ac) cells were transfected with vectors encoding green fluorescent protein (GFP) under the control of hsp70B promoter. Aliquots of $10-μl$ transfected cells $(5×107cells∕ml)$ were placed in $0.2-ml$ thin-wall polymerase chain reaction tubes and exposed to $1.1-MHz$ high intensity focused ultrasound (HIFU) at a peak negative pressure $P−=2.68MPa$⁠. By adjusting the duty cycle of the HIFU transducer, the cell suspensions were heated to a peak temperature from $50to70°C$ in $1–10s$⁠. Exposure dependent cell viability and gene activation were evaluated. For a $5-s$ HIFU exposure, cell viability dropped from 95% at $50°C$ to 13% at $70°C$⁠. Concomitantly, gene activation in sublethally injured tumor cells increased from 4% at $50°C$ to 41% at $70°C$⁠. A similar trend was observed at $60°C$ peak temperature as the exposure time increased from $1to5s$⁠. Further increase of exposure duration to $10s$ led to significantly reduced cell viability and lower overall gene activation in exposed cells. Altogether, maximum HIFU-induced gene activation was achieved at $60°C$ in $5s$⁠. Under these experimental conditions, HIFU-induced gene activation was found to be produced primarily by thermal rather than mechanical stresses.