Efficacy Estimation of Microbubble-Assisted Local Sonothrombolysis Using a Catheter with a Series of Miniature Transducers

Abstract

Intravascular ultrasound has good prospects for clinical applications in sonothrombolysis. The catheter-based side-looking intravascular ultrasound thrombolysis (e.g., Ekosonic catheters) used in clinical studies has a high frequency (2 MHz). The lower-frequency ultrasound requires a larger-diameter transducer. In our study, we designed and manufactured a small ultrasound-based prototype catheter that can emit a lower frequency ultrasound (1.1 MHz). In order to evaluate the safety and efficacy of local low-frequency ultrasound-enhanced thrombolysis, a microbubble (MB) was introduced to augment thrombolysis effect of locally delivered low-intensity ultrasound. The results demonstrated that combination of ultrasound and MB realized higher clot lysis than urokinase-only treatment (17.0% ± 1.2% vs. 14.9% ± 2.7%) under optimal ultrasound settings of 1.1 MHz, 0.414 MPa, 4.89 W/cm², 5% duty cycle and MB concentration of 60 μg/mL. When urokinase was added, the fibrinolysis accelerated by MB and ultrasound resulted in a further increased thrombolysis rate that was more than two times than that of urokinase alone (36.7% ± 5.5% vs. 14.9% ± 2.7%). However, a great quantity of ultrasound energy was required to achieve substantial clot lysis without MB, leading to the situation that temperature accumulated inside the clot became harmful. We suggest that MB-assisted local sonothrombolysis be considered as adjuvant therapy of thrombolytic agents.

Keywords: catheter-directed thrombolysis; deep vein thrombosis; microbubble; sonothrombolysis; thrombolysis; ultrasound.

Figure 1

Schematic wiring diagram of the ultrasound core wire.

Figure 2

Ultrasound-based prototype catheter.

Figure 3

Thrombolysis catheter system.

Figure 4

Schematic of thrombolysis experimental system.

Figure 5

Mass reduction of bovine blood clots in static saline. (a) Clot lysis rate of clots prepared at each volume of CaCl₂. Clots were exposed in saline for 60 min without storing at 5 °C in advance (n = 3). (b) Clot lysis rate for various cold preservation times. Clots formed with 70 $\mathsf{\mu }\mathrm{L}$ CaCl₂ were exposed in saline for 60 min (n = 3). (c) Clot lysis rate over different standing times. Clots were prepared with 70 $\mathsf{\mu }\mathrm{L}$ CaCl₂ and stored in 5 °C for at least 24 h before experiment (n = 3, * p < 0.05, ** p < 0.01, *** p < 0.001).

Figure 6

(a) Impedance amplitude and phase curves of ultrasound transducer. (b) Peak negative pressure conversion profile of transducers in one ultrasound-based prototype catheter. Distance of each transducer to the hydrophone is 2 mm. The fitted formula for each transducer is (1) $\mathrm{y}=0.0074\mathrm{x}-0.0243,$ R² = 0.9999; (2) $\mathrm{y}=0.0075\mathrm{x}-0.0518,$R² = 0.9988; (3) $\mathrm{y}=0.0079\mathrm{x}-0.0334,$R² = 0.9992; (4) $\mathrm{y}=0.071\mathrm{x}-0.0199,$R² = 0.9984. (c) Time domain waveform of acoustic pressure.

Figure 7

The relation between voltage squared integral and distance.

Figure 8

The normalized spatial distribution diagram of the sound field.

Figure 9

In vitro US alone experiment result. Clot lysis vs. duty cycle (n = 3, US settings of 1.1 MHz, 50 V_pp input voltage, 500 PRF, 60 min treatment).

Figure 10

The temperature rise of thrombus interior at each duty cycle (at 1.1 MHz, 60 V_pp input voltage, 500 PRF).

Figure 11

Initial in vitro test for MB-assisted local sonothrombolysis. (a) Clot mass reduction in an active group before and after treatment. (b) Clot mass reduction in the control group before and after treatment.

Figure 12

In vitro US + MB experiment results. (a) Clot lysis vs. MB concentration (n = 3, US settings of 1.1 MHz, 60 V_pp input voltage, 500 PRF, 5% duty cycle, 60 min treatment). (b) Clot lysis vs. input voltage (n = 3, US settings of 1.1 MHz, 500 PRF, 5% duty cycle, 60 $\mathsf{\mu }\mathrm{g}/\mathrm{mL}$, MB injected at 1 mL/h, 60 min treatment). (c) Clot lysis vs. duty cycle (n = 3, US settings of 1.1 MHz, 60 V_pp input voltage, 500 PRF, 60 $\mathsf{\mu }\mathrm{g}/\mathrm{mL}$, MB injected at 1 mL/h, 60 min treatment). (d) Clot lysis vs. time (n = 3, US settings of 1.1 MHz, 60 V_pp input voltage, 500 Hz PRF, 60 $\mathsf{\mu }\mathrm{g}/\mathrm{mL}$, MB injected at 1 mL/h) (*** p < 0.001, ** p < 0.01, * p < 0.05).

Figure 13

In vitro US + MB + urokinase experiment result. Clot lysis vs. urokinase concentration (n = 3, US settings of 1.1 MHz, 60 V_pp input voltage, 500 Hz PRF, 60 $\mathsf{\mu }\mathrm{g}/\mathrm{mL}$ MB in combination with 20,000 units/mL urokinase injected at 2 mL/h, 60 min treatment) (*** p < 0.001, * p < 0.05).