Inertial Cavitation Ultrasound with Microbubbles Improves Reperfusion Efficacy When Combined with Tissue Plasminogen Activator in an In Vitro Model of Microvascular Obstruction

Akash Goyal, Francois T H Yu, Mathea G Tenwalde, Xucai Chen, Andrew Althouse, Flordeliza S Villanueva, John J Pacella, Akash Goyal, Francois T H Yu, Mathea G Tenwalde, Xucai Chen, Andrew Althouse, Flordeliza S Villanueva, John J Pacella

Abstract

We have previously reported that long-tone-burst, high-mechanical-index ultrasound (US) and microbubble (MB) therapy can restore perfusion in both in vitro and in vivo models of microvascular obstruction (MVO). Addition of MBs to US has been found to potentiate the efficacy of thrombolytics on large venous thrombi; however, the optimal US parameters for achieving microvascular reperfusion of MVO caused by microthrombi, when combined with tissue plasminogen activator (tPA), are unknown. We sought to elucidate the specific effects of US, with and without tPA, for effective reperfusion of MVO in an in vitro model using both venous and arterial microthrombi. Venous- and arterial-type microthrombi were infused onto a mesh with 40-μm pores to simulate MVO. Pulsed US (1 MHz) was delivered with inertial cavitation (IC) (1.0 MPa, 1000 cycles, 0.33 Hz) and stable cavitation (SC) US (0.23 MPa, 20% duty cycle, 0.33 Hz) regimes while MB suspension (2 × 106 MBs/mL) was infused. The efficacy of sonoreperfusion with these parameters was tested with and without tPA. Sonoreperfusion efficacy was significantly greater for IC + tPA compared with tPA alone, IC, SC and SC + tPA, suggesting lytic synergism between tPA and US for both venous- and arterial-type microthrombi. In contrast to our previous in vitro studies using 1.5 MPa at 5000 US cycles without tPA, the IC regime employed herein used 90% less US energy. These findings suggest an IC regime can be used with tPA synergistically to achieve a high degree of fibrinolysis for both thrombus types.

Keywords: Microbubbles; Microcirculation; Microvascular obstruction; Myocardial infarction; Sonothrombolysis; Ultrasound; Ultrasound contrast agents.

Copyright © 2017 World Federation for Ultrasound in Medicine & Biology. Published by Elsevier Inc. All rights reserved.

Figures

Figure 1
Figure 1
Schematic diagram of the in-vitro model of microvascular obstruction.
Figure 2
Figure 2
Characteristics of thrombi. (a) Photographs of venous and arterial thrombi. (b) Scanning electron microscope (SEM) image (×2000) of venous thrombus. (c) SEM image of arterial thrombus.
Figure 3
Figure 3
PCD data. (a) Representative power spectra of inertial cavitation (IC) regime. (b) Representative power spectra of stable cavitation regime (SC). (c) Comparison of the averaged power spectra during the first 1 ms for IC regime and during the first 10 ms for SC regime.
Figure 4
Figure 4
Time course of upstream pressure during ultrasound and microbubble treatment for venous thrombi (A) and arterial thrombi (B). Values presented as mean ± standard deviation (n=4–5 for each experimental condition).
Figure 5
Figure 5
Lytic index for venous and arterial microthrombi under various conditions (n=4–5 for each experimental condition). Means with the same letter are not statistically different from each other. Lytic index was greater for IC+tPA than all other treatment conditions for venous microthrombi (p<0.05) and than tPA alone and SC for arterial microthrombi (p<0.05). Comparing across microthrombus type, lytic index for IC+tPA was greater for venous type than arterial type (p<0.05).
Figure 6
Figure 6
Early lytic rate for venous and arterial microthrombi under various conditions (n=4–5 for each experimental condition). Means with the same letter are not statistically different from each other. Early lytic rate was greater for IC+tPA for venous microthrombi than all other treatment conditions (p<0.05). No statistical differences were noted across microthrombus type (p>0.05).
Figure 7
Figure 7
Late lytic rate for venous and arterial microthrombi (n=4–5 for each experimental condition). Means with the same letter are not statistically different from each other. Late lytic rate was greater for IC+tPA for venous microthrombi than tPA alone, SC, and IC (p<0.05), while late lytic rate for IC+tPA for arterial microthrombi was greater than tPA alone and SC (p<0.05). Late lytic rate for SC+tPA was greater than SC for venous microthrombi (p<0.05) while not statistically different for arterial microthrombi (p<0.05). As a group, late lytic rate was greater for venous microthrombi than arterial microthrombi (p<0.05), but no specific treatment group was significantly different.

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