Acoustic radiation force for vascular cell therapy: in vitro validation

Abstract

Cell-based therapeutic approaches are attractive for the restoration of the protective endothelial layer in arteries affected by atherosclerosis or following angioplasty and stenting. We have recently demonstrated a novel technique for the delivery of mesenchymal stem cells (MSCs) that are surface-coated with cationic lipid microbubbles (MBs) and displaced by acoustic radiation force (ARF) to a site of arterial injury. The objective of this study was to characterize ultrasound parameters for effective acoustic-based delivery of cell therapy. In vitro experiments were performed in a vascular flow phantom where MB-tagged MSCs were delivered toward the phantom wall using ARF generated with an intravascular ultrasound catheter. The translation motion velocity and adhesion of the MB-cell complexes were analyzed. Experimental data indicated that MSC radial velocity and adhesion to the vessel phantom increased with the time-averaged ultrasound intensity up to 1.65 W/cm², after which no further significant adhesion was observed. Temperature increase from baseline near the catheter was 5.5 ± 0.8°C with this setting. Using higher time-averaged ultrasound intensities may not significantly benefit the adhesion of MB-cell complexes to the target vessel wall (p = NS), but could cause undesirable biologic effects such as heating to the MB-cell complexes and surrounding tissue. For the highest time-averaged ultrasound intensity of 6.60 W/cm², the temperature increase was 11.6 ± 1.3°C.

Figure 1

Experimental setup using a vessel phantom. The negatively charged fluorescent MSCs were labeled with cationic microbubbles. Application of acoustic radiation force caused marginalization and adhesion of the cells to the vessel wall. The region of interest (2 mm × 2 mm) included the lower half portion of the phantom.

Figure 2

Quantification of microbubble attachment of MSCs using flow cytometry. (A) Front scatter cross-section (FSC) and side scatter cross-section (SSC) profile (left column) and the distribution of SSC (right column) of fluorescently labeled HMSC alone. (B) FSC/SSC profile (left column) and the distribution of SSC (right column) of fluorescently labeled HMSC, after mixing with cationic microbubbles (1:40 HMSC to MB ratio). In the FSC/SSC profile plots, the fluorescent events of HMSCs were shown in green, while the microbubbles were shown in black. HMSC association with the cationic microbubbles led to increased SSC of the cells relative to HMSCs alone.

Figure 3

The axial velocities within the phantom vessel followed a concentric flow pattern (shear stress at vessel wall 1.1 Pa). The experimental data for axial velocities (solid line) with no ultrasound exposure were well correlated with theoretical values (green dashed line).

Figure 4

Radial velocities of MB-cell complexes towards the vessel wall. Bars grouped together had different tone burst configurations but had the same duty cycle and therefore same time-averaged ultrasound intensity.

Figure 5

Experimental radial velocity of the MSCs plotted against time-averaged ultrasound intensity.

Figure 6

The total number of MB-MSCs adhered on the vessel wall during 10 seconds of ultrasound exposure (solid line) and 5 seconds after the ultrasound was turned off following the US treatment (dotted line).

Figure 7

Flow trajectories of the MB-cell complexes using the measured radial velocity at various ultrasound parameter combinations (axial velocity obtained from the theoretical concentric flow pattern). For simplicity, only MB-cell complexes entering the field of view at 3 different locations (1.35 mm, 1 mm, and 0.6 mm away from the vessel wall) were plotted.

Figure 8

Flow trajectories of MB-cell complexes for various acoustic pressures and duty cycles after maximum intensity persistence was applied to the recorded images.