The accumulation of microlesions induced by ultrasound interaction with contrast microbubbles in the myocardium potentially represents a new method of tissue reduction therapy. Anesthetized rats were treated in a heated water bath with 1.5 MHz focused ultrasound pulses triggered once every 4 heartbeats from the electrocardiogram (ECG) during infusion of microbubble contrast agent. Treatment was guided by an 8 MHz B-mode imaging transducer, which also was used to provide estimates of the left ventricular echogenicity (LVE) as a possible predictor of efficacy during treatment. Strategies to reduce prospective clinical treatment durations were tested, including pulse modulation to simulate a theranostic scanning strategy, and an increased agent infusion rate over shorter durations. Sources of variability, including ultrasound path variation and venous catheter placement, also were investigated. ECG premature complexes (PCs) were monitored and Evans-blue stained cardiomyocyte scores (SCSs) were obtained from frozen sections. The LVE reflected variations in the infused microbubble concentration, but failed to predict efficacy. Comparing suspensions of varied microbubble sizes revealed that LVE was dominated by larger bubbles, while efficacy appeared to be dependent on smaller sizes. Simulated scanning was as effective as the normal fixed-beam treatment, and high agent infusion allowed reduced treatment duration. The success of these theranostic strategies may increase the prospects for realistic clinical translation of myocardial cavitation enabled therapy.

Myocardial contrast echocardiography at enhanced therapeutic parameters may be a novel means of tissue reduction therapy, as for hypertrophic cardiomyopathy. Dahl/SS rats were anesthetized and treated by high amplitude pulsed ultrasound guided by 10 MHz ultrasound images. Contrast microbubbles were infused via the tail vein during intermittent pulse-burst exposure at 4 MPa. A sham group, a low impact group (group A, 5 cycle pulses with Gaussian modulation and 1:4 trigger for 5 min) and a high impact group (group B, 10 cycle pulses with 4 ms square modulation and 1:8 trigger for 10 min) were tested. Higher exposure used in group B yielded more substantial injury than lower exposure in group A. Treated rats in both group A and B had significant increases in wall thickness measured by echocardiography the next day, which returned to normal by the end of 6 weeks. Six weeks after ultrasound exposure, heart tissue samples showed tissue fibrosis in Masson’s trichrome stained histology. Maturation of lesions involved fibrosis replacement, preserving structural tissue integrity. This study showed that myocardial injury noted previously progresses into permanent loss of myocardial tissue that may be sufficient for possible hypertrophic cardiomyopathy therapy. More research is needed to define the treatment parameters required for symptomatic relief for hypertrophic cardiomyopathy.

Intermittent high intensity ultrasound pulses with circulating contrast agent microbubbles can induce scattered cavitation myocardial microlesions of potential value for tissue reduction therapy. Here, computer-aided histological evaluation of the effective treated volume was implemented to optimize ultrasound pulse parameters, exposure duration, and contrast agent dose. Rats were treated with 1.5 MHz focused ultrasound bursts and Evans blue staining indicates lethal cardiomyocytic injury. Each heart was sectioned to provide samples covering the entire exposed myocardial volume. Both brightfield and fluorescence images were taken for up to 40 tissue sections. Tissue identification and microlesion detection were first done based on 2D images to form microlesion masks containing the outline of the heart and the stained cell regions. Image registration was then performed on the microlesion masks to reconstruct a volume-based model according to the morphology of the heart. The therapeutic beam path was estimated from the 3D stacked microlesions, and finally the total microlesion volume, here termed macrolesion, was characterized along the therapeutic beam axis. Radially symmetric fractional macrolesions were characterized via stepping disks of variable radius determined by the local distribution of microlesions. Treated groups showed significant macrolesions of a median volume of 87.3 μL, 2.7 mm radius, 4.8 mm length, and 14.0% lesion density compared to zero radius, length, and lesion density for sham. The proposed radially symmetric lesion model is a robust evaluation for Myocardial Contrast Enabled Therapy (MCET). Future work will include validating the proposed method with varying acoustic exposures and optimizing involved parameters to provide macrolesion characterization.

Phase-change perfluorohexane nanodroplets (PFHnDs) are a new class of recondensable submicrometer-sized contrast agents that have potential for contrast-enhanced and super-resolution ultrasound imaging with an ability to reach extravascular targets. The PFHnDs can be optically triggered to undergo vaporization, resulting in spatially stationary, temporally transient microbubbles. The vaporized PFHnDs are hyperechoic in ultrasound imaging for several to hundreds of milliseconds before recondensing to their native, hypoechoic, liquid nanodroplet state. The decay of echogenicity, i.e., the dynamic behavior of the ultrasound signal from optically triggered PFHnDs in ultrasound imaging, can be captured using high frame rate ultrasound imaging. We explore the possibility to manipulate the echogenicity dynamics of optically triggered PFHnDs in ultrasound imaging by changing the phase of the ultrasound imaging pulse. Specifically, the ultrasound imaging system was programmed to transmit two imaging pulses with inverse polarities. We show that the imaging pulse phase can affect the amplitude and the temporal behavior of PFHnD echogenicity in ultrasound imaging. The results of this study demonstrate that the ultrasound echogenicity is significantly increased (about 78% improvement) and the hyperechoic timespan of optically triggered PFHnDs is significantly longer (about 4 times) if the nanodroplets are imaged by an ultrasound pulse starting with rarefactional pressure versus a pulse starting with compressional pressure. Our finding has direct and significant implications for contrast-enhanced ultrasound imaging of droplets in applications such as super-resolution imaging and molecular imaging where detection of individual or low-concentration PFHnDs is required.

BackgroundMyocardial cavitation-enabled therapy (MCET) has been proposed as a means to achieve minimally invasive myocardial reduction using ultrasound to produce scattered microlesions by cavitating contrast agent microbubbles.MethodsRats were treated using burst mode focused ultrasound at 1.5 MHz center frequency and varying envelope and pressure amplitudes. Evans blue staining indicated lethal cardiomyocytic injury. A previously developed quantitative scheme, evaluating the histologic treatment results, provides an insightful analysis for MCET treatment parameters. Such include ultrasound exposure amplitude and pulse modulation, contrast agent dose, and infusion rate.ResultsThe quantitative method overcomes the limitation of visual scoring and works for a large dynamic range of treatment impact. Macrolesions are generated as an accumulation of probability driven microlesion formations. Macrolesions grow radially with radii from 0.1 to 1.6 mm as the ultrasound exposure amplitude (peak negative) increases from 2 to 4 MPa. To shorten treatment time, a swept beam was investigated and found to generate an acceptable macrolesion volume of about 40 μL for a single beam position.ConclusionsUltrasound parameters and administration of microbubbles directly influence lesion characteristics such as microlesion density and macrolesion dimension. For lesion generation planning, control of MCET is crucial, especially when targeting larger pre-clinical models.