Ultrasound well below the intensity threshold of cavitation can promote efficient uptake of small drug model molecules in fibroblast cells

Ultrasound (US) induced enhancement of plasma membrane permeability is a hugely promising tool for delivering exogenous vectors at the specific biological site in a safe and efficient way. In this respect, here we report effects of membrane permeability alteration on fibroblast-like cells undergoing very low-intensity of US. The change in permeability was pointed out in terms of high uptake efficiency of the fluoroprobe calcein, thus resembling internalization of small cell-impermeable model drugs, as measured by fluorescence microscopy and flow cytometry. Fluorescence evidences moreover suggests that the higher the time of exposure, the larger will be the size of molecules can be internalized. The uptake events were related to the cell viability and also with structural changes occurring at membrane level as revealed by infrared spectroscopy and preliminary membrane fluidity and atomic force microscopy (AFM) investigation. Thus, the question of whether the uptake of cell-impermeable molecules is consistent with the presence of disruptions on the cell membrane (sonopore formation) has been addressed. In this framework, our findings may constitute experimental evidence in support of sub-cavitation sonoporation models recently proposed, and they may also provide some hints towards the actual working condition of medical US dealing with the optimum risk to benefit therapeutic ratio.