Microbubble adhesion to healthy and diseased vascular cells - the role of surface charge

Abstract

Medical ultrasound (US) imaging is an established and powerful tool for diagnostic imaging. Combined with contrast enhancing agents called microbubbles, its imaging performance has been improved, as well as offering a potential tool for molecular imaging and targeted drug delivery and therapeutics. Microbubbles (MBs) consist of micron-sized, inert gas cores stabilised by a lipid monolayer or protein and suspended in aqueous dispersion. These non-targeted blood perfusion agents were found to persist in circulation, sticking to sites of inflammation or injury in the vasculature. This non-targeted MB retention in the vasculature is not fully understood and this project investigates the role of surface charge on MB-cellular adhesive behaviour. The adhesion of current clinical MBs, SonoVue™, Definity™, Optison™, and an experimental agent, BR38, with healthy and diseased vascular phantoms were observed in vitro. Human umbilical vein endothelial cells (HUVECs) were used to mimic the healthy vasculature and TNFα-activated HUVECs mimicked the diseased vasculature. Some degree of preferential binding for the TNFα-activated versus non-activated HUVECs was observed, indicating a surface component to this interaction. We hypothesised that surface charge plays a role in this interaction. MB surface charge was characterised with laser Doppler electrophoresis (LDE), the most widely-used method for determining particle surface charge. However, MB buoyancy compromised the reliability of this method and hence a micro-electrophoretic technique to determine MB surface charge was developed. MB electrophoretic movement was tracked using an in-house algorithm which was able to determine MB surface charge with at least a 10-fold improvement in relative standard deviations when compared to the LDE method. Our findings suggest an electrostatic role in MB adhesion to TNFα-activated HUVECs for phospholipid-stabilised MBs but not for protein-stabilised MBs. In conclusion, while surface charge may play a role in MB adhesion, there may be other unknown factors which can contribute to MB-cellular interaction.