Dynamics and Mechnisms of Ultrasound and Microbubble Mediated Intracellular Plasmid DNA Uptake

Ultrasound and microbubble has shown great potential in gene therapy as this technology can achieve noninvasive and effective intracellular gene delivery and transfection in vitro and in vivo. However, the underlying mechanism of how plasmid DNA intracellular uptake occurs is not fully understood. This study aims to reveal the dynamic process of ultrasound and microbubble mediated plasmid DNA intracellular uptake and unveil the mechanisms involved on single cell level. Plasmid DNA with radius of 35.1±10.6nm can achieve intracellular uptake through transient membrane pore with radius of 86.5±45nm in theorical calculation. The spatial correlation between bubble and trans-membrane plasmid DNA demonstrates that transient pore is an effective route for plasmid DNA uptake. Based on time-resolved bubble dynamics, shear stress on cell membrane are estimated under both high pressure, short pulse and low pressure, long pulse conditions. The endocytosis inhibition study reveals that endocytosis plays an important role under both ultrasound conditions. Some plasmid DNA in cytoplasm not correlated with bubbles or membrane vesicles suggest that other global and fast mechanism may be involved under both ultrasound conditions. In sum, we conclude that the plasmid DNA intracellular uptake is a multi-mechanisms involved, bubble-cell interaction dependent, dynamic process.