Engineered 3D microvascular networks for the study of ultrasound-microbubble-mediated drug delivery.

Localized and targeted drug delivery can be achieved by the combined action of ultrasound, and microbubbles on the tumor microenvironment, likely through sonoporation and other therapeutic mechanisms not well understood. Here, we present a perfusable in vitro model with a realistic 3D geometry to study the interactions between microbubbles and the vascular endothelium in the presence of ultrasound. Specifically, a three-dimensional, endothelial-cell-seeded in vitro microvascular model was perfused with cell culture medium and microbubbles while sonicated by a single-element 1 MHz focused transducer. This setup mimics the in vivo scenario in which ultrasound induces a therapeutic effect in the tumor vasculature in the presence of flow. Fluorescence and bright-field microscopy were employed to assess the microbubble-vessel interactions and the extent of drug delivery and cell death both in real time during treatment as well as after treatment. Propidium iodide was used as the model drug while calcein AM was used to evaluate cell viability. The acoustic parameter sets chosen for this work were: 1) acoustic pressure: 1.4 MPa, pulse length: 500 cycles, duty cycle: 5%, and 2) acoustic pressure: 0.4 MPa, pulse length: 1000 cycles, duty cycle: 20%. Enhanced drug delivery and cell death were observed in both cases while the higher pressure setting had a more pronounced effect. By introducing physiological flow to the in vitro microvascular model and examining the PECAM-1 expression of the endothelial cells within it, we demonstrated that our model is a good mimic of the in vivo vasculature and is therefore a viable platform to provide mechanistic insights into ultrasound-mediated drug delivery.