Impact of myocardial contrast echocardiography on vascular permeability: an in vivo dose response study of delivery mode, pressure amplitude and contrast dose

Abstract

An in vivo rat model of myocardial contrast echocardiography (MCE) was defined and used to examine the dose range response of microvascular permeabilization and premature ventricular contractions (PVCs) with respect to method of imaging, peak rarefactional pressure amplitude (PRPA) and agent dose. A left ventricular short axis view was obtained on anesthetized rats at 1.7 MHz using a diagnostic ultrasound system with simultaneous ECG recording. Evans blue dye, a marker for microvascular leakage, and a bolus of Optison® were injected IV. Counts of PVCs were made from video tape during the 3 min of MCE. Hearts were excised 5 min after imaging and petechial hemorrhages, Evans blue colored area and Evans blue content were determined. No PVCs or microvascular leakage were seen in rats imaged without contrast agent followed by contrast agent injection without imaging. When PVCs were detected during MCE, petechial hemorrhages and Evans blue leakage were also found in the myocardium. Triggering 1:4 at end-systole produced the most PVCs per frame and most microvascular leakage, followed by end-systole 1:1, continuous scanning and end-diastole triggering 1:1. All effects increased with increasing Optison® dosage in the range 25 to 500 μL kg⁻¹. Ultrasound PRPA was important, with apparent thresholds for PVCs at 1.0 MPa and for petechiae at 0.54 MPa. PVCs, petechial hemorrhages and microvascular leakage in the myocardium occur as a result of MCE in rats. (E-mail: [email protected])

Introduction

The echocardiographic contrast phenomenon was described 34 y ago (Gramiak and Shah, 1968) and has since been developed for detection of intracardiac shunts or abnormalities of venous connections, enhanced delineation of endocardial borders and assessment of myocardial perfusion Weyman et al., 1979, Fraker et al., 1979, Van Hare and Silverman, 1989, Crouse and Kramer, 1991, Masugata et al., 2000. Many commercial ultrasound contrast agents (USCAs) have been developed and evaluated in clinical trials for both safety and efficacy Goldberg et al., 1994, Cohen et al., 1998, Feinstein et al., 1990, Kitzman et al., 2000, Grayburn et al., 1998, Main et al., 1997, which have rarely shown side-effects. Presently, several commercial USCA are approved for clinical use for left ventricular opacification, while other agents and applications are under development and evaluation Mulvagh et al., 2000, Goldberg et al., 2001.

The key to the invention of USCA is the coincidence that the sizes of gas bodies (gas bubbles stabilized by shells or skins) that can pass through the microcirculation are the same sizes that can produce strong echoes at acoustic frequencies in the range of diagnostic ultrasound. This strong interaction can destabilize the gas bodies leading to nucleation of acoustic cavitation. Cavitation nucleation was demonstrated in vitro soon after the introduction of commercial agents (Miller and Thomas, 1995) and recently has been shown to occur in the myocardium (Porter et al. 2001). Acoustic cavitation is a well-known mechanism for micro-scale bioeffects (AIUM, 2000). Since cavitation nuclei capable of strong interaction with diagnostic ultrasound do not normally exist within the circulation, use of USCA introduces an unnatural potential for cavitational bioeffects of diagnostic ultrasound on the microcirculation (AIUM, 2000). Bioeffects of diagnostic ultrasound reported to occur with USCA include capillary leakage in vivo in rat spinotrapezius muscle (Skyba et al. 1998) and in mouse abdominal muscle (Miller and Quddus, 2000). The full range of research into this potential for microscale bioeffects with USCA-augmented ultrasound has been the subject of several reviews (AIUM, 2000; Miller, 2000).

More recent research has emphasized potential bioeffects of myocardial contrast echocardiography (MCE). Capillary rupture has been seen in an isolated rabbit heart preparation and associated with a depression of left ventricular contractile function (Ay et al. 2001). Microbubble destruction within the myocardium has been shown to cause mild elevations of troponin-T, but this was not associated with left ventricular dysfunction (Chen et al. 2002). Physiological effects of ultrasound on the heart have also been noted. Using ECG monitoring, cardiac arrhythmia has been shown for relatively high-power ultrasound without USCA (MacRobbie et al. 1997). Premature ventricular contractions (PVCs) have been noted for diagnostic ultrasound in a multicenter study of the USCA SHU-454 (Rovai et a l. 1991). PVC induction with diagnostic ultrasound has been specifically studied and reported to occur in humans with an experimental USCA (van der Wouw et al. 2000). PVCs together with histologically definable damage to the myocardium has been reported for relatively high intensity focused ultrasound with USCA (Zachary et al. 2002).

Many questions remain concerning the relation of the experimental USCA dosage and ultrasound exposure to clinical conditions and the relationship of PVCs to microvascular damage. This present research tested the hypothesis that petechial hemorrhage and microvascular leakage might occur in the myocardium concomitantly with PVC induction during diagnostic MCE. To approximate clinical scanning conditions, a commercial USCA was used at clinically relevant IV (i.e., not intraarterial) doses in the intact rat (i.e., not isolated hearts) with a calibrated diagnostic ultrasound system. For these conditions, PVCs, myocardial petechiae and microvascular leakage of a marker dye were induced by MCE in rats. The dose range response with respect to ultrasound contrast dose, ultrasound peak rarefactional pressure amplitude (PRPA) and mode of ultrasound delivery was evaluated. Results indicate a well defined dose response, with apparent thresholds for effects.