Imaging cardiac hypertrophy in hypertrophic cardiomyopathy and its differential diagnosis.

PURPOSE OF REVIEW: The aim of this study was to review imaging of myocardial hypertrophy in hypertrophic cardiomyopathy (HCM) and its phenocopies. The introduction of cardiac myosin inhibitors in HCM has emphasized the need for careful evaluation of the underlying cause of myocardial hypertrophy. RECENT FINDINGS: Advances in imaging of myocardial hypertrophy have focused on improving precision, diagnosis, and predicting prognosis. From improved assessment of myocardial mass and function, to assessing myocardial fibrosis without the use of gadolinium, imaging continues to be the primary tool in understanding myocardial hypertrophy and its downstream effects. Advances in differentiating athlete's heart from HCM are noted, and the increasing rate of diagnosis in cardiac amyloidosis using noninvasive approaches is especially highlighted due to the implications on treatment approach. Finally, recent data on Fabry disease are shared as well as differentiating other phenocopies from HCM. SUMMARY: Imaging hypertrophy in HCM and ruling out other phenocopies is central to the care of patients with HCM. This space will continue to rapidly evolve, as disease-modifying therapies are under investigation and being advanced to the clinic.

In vivo pulsed-field ablation in healthy vs. chronically infarcted ventricular myocardium: biophysical and histologic characterization.

AIMS: Data on ventricular pulsed-field ablation (PFA) are sparse in the setting of chronic myocardial infarction (MI). The objective of this study was to compare the biophysical and histopathologic characteristics of PFA in healthy and MI swine ventricular myocardium. METHODS AND RESULTS: Myocardial infarction swine (n = 8) underwent coronary balloon occlusion and survived for 30 days. We then performed endocardial unipolar, biphasic PFA of the MI border zone and a dense scar with electroanatomic mapping and using an irrigated contact force (CF)-sensing catheter with the CENTAURI System (Galaxy Medical). Lesion and biophysical characteristics were compared with three controls: MI swine undergoing thermal ablation, MI swine undergoing no ablation, and healthy swine undergoing similar PFA applications that included linear lesion sets. Tissues were systematically assessed by gross pathology utilizing 2,3,5-triphenyl-2H-tetrazolium chloride staining and histologically with haematoxylin and eosin and trichrome. Pulsed-field ablation in healthy myocardium generated well-demarcated ellipsoid lesions (7.2 ± 2.1 mm depth) with contraction band necrosis and myocytolysis. Pulsed-field ablation in MI demonstrated slightly smaller lesions (depth 5.3 ± 1.9 mm, P = 0.0002), and lesions infiltrated into the irregular scar border, resulting in contraction band necrosis and myocytolysis of surviving myocytes and extending to the epicardial border of the scar. Coagulative necrosis was present in 75% of thermal ablation controls but only in 16% of PFA lesions. Linear PFA resulted in contiguous linear lesions with no gaps in gross pathology. Neither CF nor local R-wave amplitude reduction correlated with lesion size. CONCLUSION: Pulsed-field ablation of a heterogeneous chronic MI scar effectively ablates surviving myocytes within and beyond the scar, demonstrating promise for the clinical ablation of scar-mediated ventricular arrhythmias.

Myocardial contrast echocardiography assessment of perfusion abnormalities in hypertrophic cardiomyopathy.

BACKGROUND: Perfusion defects during stress can occur in hypertrophic cardiomyopathy (HCM) from either structural or functional abnormalities of the coronary microcirculation. In this study, vasodilator stress myocardial contrast echocardiography (MCE) was used to quantify and spatially characterize hyperemic myocardial blood flow (MBF) deficits in HCM. METHODS: Regadenoson stress MCE was performed in patients with septal-variant HCM (n = 17) and healthy control subjects (n = 15). The presence and spatial distribution (transmural diffuse, patchy, subendocardial) of perfusion defects was determined by semiquantitative analysis. Kinetic analysis of time-intensity data was used to quantify MBF, microvascular flux rate (ß), and microvascular blood volume. In patients undergoing septal myectomy (n = 3), MCE was repeated > 1 years after surgery.  RESULTS: In HCM subjects, perfusion defects during stress occurred in the septum in 80%, and in non-hypertrophied regions in 40%. The majority of septal defects (83%) were patchy or subendocardial, while 67% of non-hypertrophied defects were transmural and diffuse. On quantitative analysis, hyperemic MBF was approximately 50% lower (p < 0.001) in the hypertrophied and non-hypertrophied regions of those with HCM compared to controls, largely based on an inability to augment ß, although hypertrophic regions also had blood volume deficits. There was no correlation between hyperemic MBF and either percent fibrosis on magnetic resonance imaging or outflow gradient, yet those with higher degrees of fibrosis (≥ 5%) or severe gradients all had low septal MBF during regadenoson. Substantial improvement in hyperemic MBF was observed in two of the three subjects undergoing myectomy, both of whom had severe pre-surgical outflow gradients at rest. CONCLUSION: Perfusion defects on vasodilator MCE are common in HCM, particularly in those with extensive fibrosis, but have a different spatial pattern for the hypertrophied and non-hypertrophied segments, likely reflecting different contributions of functional and structural abnormalities. Improvement in hyperemic perfusion is possible in those undergoing septal myectomy to relieve obstruction.  TRIAL REGISTRATION: ClinicalTrials.gov NCT02560467.

Cardiac allograft vasculopathy diagnosed by vasodilator myocardial contrast echocardiography perfusion imaging.

Cardiac allograft vasculopathy (CAV) remains a common long-term complication of cardiac transplantation. While invasive coronary angiography is considered the gold standard, it is also invasive and lacks sensitivity to detect early, distal CAV. Although vasodilator stress myocardial contrast echocardiography perfusion imaging (MCE) is used in the detection of microvascular disease in non-transplant patients, there is little data guiding its use in transplant recipients. Herein is a case series of four heart transplant recipients that had vasodilator stress MCE performed in addition to invasive coronary angiography for CAV surveillance. MCE at rest and after regadenason was performed using a continuous infusion of lipid-shelled microbubbles. We describe a case of normal microvascular function, diffuse microvascular dysfunction, patchy sub-endocardial perfusion defects and a focal sub-endocardial perfusion defect. Cardiac allograft vasculopathy can be heralded by several different perfusion patterns on MCE in patients after orthotopic heart transplant. The varying prognoses and potential interventions for these different patterns require further investigation.

Resting coronary flow drives the daily pattern in coronary flow reserve in patients with chest pain without obstructive epicardial stenosis.

Objectives: Ischemia with no obstructive coronary artery disease (INOCA) is a risk factor for major adverse cardiovascular events and is characterized by abnormal coronary microvascular tone. In patients with INOCA, adverse cardiovascular events most commonly occur in the morning compared to other times of the day and night. Materials and methods: We tested whether coronary microvascular function varies diurnally with attenuation in the morning in patients with symptomatic coronary artery disease without significant (>50%) epicardial stenosis. We evaluated data from 17 patients studied in the AM (700-1159 h) and 11 patients in the PM (1200-1800 h). Coronary microvascular function was measured using perfusion contrast imaging at rest and after infusion of intravenous regadenoson. We calculated microvascular flow reserve as the ratio of hyperemic to resting flow. Along with independent sample t-tests, we performed bootstrapping procedures to test mean differences between AM and PM groups, using the bias-corrected and accelerated method with 5,000 bootstrapped samples. Results and conclusion: The AM and PM groups were matched for demographic and existing risk factors. Coronary microvascular flow reserve was ∼33% higher in the AM compared to the PM (P = 0.025, BCa 95% CI [0.25, 1.64]; Hedge's g = 0.89, 95% CI [0.11, 1.66]) as a result of significantly lower resting flow (∼50%) in the AM compared to the PM (P = 0.03, M Diff = -56.65, BCa 95% CI [-118.59, -2.12]; Hedge's g = -0.86, 95% CI [-1.60, -0.06]). Our observations are of clinical value and can influence diagnosis and treatment in the clinic based on the time of day of measurements.

Multi-modality imaging assessment of microbubbles and cerebral emboli in left ventricular pulsed field ablation.

BACKGROUND: Pulsed field ablation (PFA) may have a superior safety profile compared to other technologies, but it has the potential to cause gaseous microbubbles (MB), which may be associated with cerebral emboli. Limited relative safety data has been published regarding PFA in the left ventricle (LV). METHODS: Healthy and chronic myocardial infarction (MI) swine underwent PFA (monopolar, biphasic, 25 Amps) in the LV using an irrigated focal catheter under intra-cardiac echocardiography (ICE) guidance for MB monitoring. Two control swine received air MBs through the lumen of the ablation catheter. Swine underwent brain MRI before and after PFA (or control air MB injection). Gross pathology and histology of brains with abnormal MRI findings were performed. RESULTS: Four healthy and 5 chronic MI swine underwent 124 left ventricular PFA applications. No PFA-related MB formation was noted on ICE. Both control swine developed multiple acute emboli in the thalamus and caudate on DWI, ADC, and FLAIR brain MRI images in response to air MB injection. Of the 9 PFA swine, there were no abnormalities on ADC or FLAIR images. There was one hyperintense focus in the left putamen on the DWI trace image, but the absence of ADC or FLAIR affirmation suggested it was artifact. Gross pathology and histopathology of this region did not detect any abnormalities. CONCLUSIONS: Focal monopolar biphasic PFA of both healthy and chronically infarcted left ventricular myocardium does not generate any MB or cerebral emboli observable on ICE and brain MRI.

Increased Susceptibility for Adverse Reactions to Ultrasound Enhancing Agents in Sickle Cell Disease.

BACKGROUND: Pain-related adverse events (AEs) to ultrasound enhancing agents (UEAs) have been reported in patients with sickle cell disease (SCD). The aims of this study were to characterize the scope of these AEs in the SCD population and to investigate potential mechanisms on the basis of pathways involved in SCD vaso-occlusive crisis (VOC) and pain. METHODS: The prevalence and classification of AEs were analyzed from two clinical trials in which high-dose Definity infusions were used in patients with SCD (n = 55) or matched control subjects (n = 43) to study muscle or myocardial microvascular perfusion. Because complement (C') activation can trigger VOC in SCD, C' activation and surface adhesion of C' proteins on lipid UEAs were studied in vitro. C'-mediated UEA attachment to bone marrow immune cells was assessed using flow cytometry in a murine SCD model (Townes mice). Blood from patients receiving Definity was obtained to measure specific lysophospholipid metabolites of lipids in Definity thought to mediate SCD pain. RESULTS: Moderate or greater AEs, all of which were nociceptive (back or bone pain), occurred in one control subject and nine SCD subjects (2% vs 16%, P = .02). Patients with SCD who had AEs tended to have more severe manifestations of SCD. Three of the subjects with SCD had previously received Definity without complications. In patients with SCD, four AEs were classified as severe in intensity and as serious AEs on the basis of need for medical intervention. AEs were described to be similar to SCD-related pain, but there was no evidence for VOC, hemolysis, hypotension, or hypoxemia. At baseline, markers of C' activation were greater in patients with SCD than control subjects. However, after administration of lipid UEAs, SCD and control subjects were similar with regard to C' activation response, anaphylatoxin production, bone marrow microbubble retention, and production of lysophospholipids. There was a trend toward increased deposition of C3b and C3bi on lipid UEAs exposed to serum from patients with SCD. CONCLUSIONS: Patients with SCD are particularly susceptible to nociceptive AEs when given Definity at high doses. The mechanism for these AEs remains unclear but most are not related to the triggering of classic VOC.

Contrast Ultrasound Assessment of Skeletal Muscle Recruitable Perfusion after Permanent Left Ventricular Assist Device Implantation: Implications for Functional Recovery.

BACKGROUND: In heart failure with reduced ejection fraction (HFrEF), abnormal regulation of skeletal muscle perfusion contributes to reduced exercise tolerance. The aim of this study was to test the hypothesis that improvement in functional status after permanent left ventricular assist device (LVAD) implantation in patients with HFrEF is related to improvement in muscle perfusion during work, which was measured using contrast-enhanced ultrasound (CEUS). METHODS: CEUS perfusion imaging of calf muscle at rest and during low-intensity plantar flexion exercise (20 W, 0.2 Hz) was performed in patients with HFrEF (n = 22) at baseline and 3 months after placement of permanent LVADs. Parametric analysis of CEUS data was used to quantify muscle microvascular blood flow (MBF), blood volume index, and red blood cell flux rate. For subjects alive at 3 months, comparisons were made between those with New York Heart Association functional class I or II (n = 13) versus III or IV (n = 7) status after LVAD. Subjects were followed for a median of 5.7 years for mortality. RESULTS: Echocardiographic data before and after LVAD placement and LVAD parameters were similar in subjects classified with New York Heart Association functional class I-II versus functional class III-IV after LVAD. Skeletal muscle MBF at rest and during exercise before LVAD implantation was also similar between groups. After LVAD placement, resting MBF remained similar between groups, but during exercise those with New York Heart Association functional class I or II had greater exercise MBF (111 ± 60 vs 52 ± 38 intensity units/sec, P = .03), MBF reserve (median, 4.45 [3.95 to 6.80] vs 2.22 [0.98 to 3.80]; P = .02), and percentage change in exercise MBF (median, 73% [-28% to 83%] vs -45% [-80% to 26%]; P = .03). During exercise, increases in MBF were attributable to faster microvascular flux rate, with little change in blood volume index, indicating impaired exercise-mediated microvascular recruitment. The only clinical or echocardiographic feature that correlated with post-LVAD exercise MBF was a history of diabetes mellitus. There was a trend toward better survival in patients who demonstrated improvement in muscle exercise MBF after LVAD placement (P = .05). CONCLUSIONS: CEUS perfusion imaging can quantify peripheral vascular responses to advanced therapies for HFrEF. After LVAD implantation, improvement in functional class is seen in patients with improvements in skeletal muscle exercise perfusion and flux rate, implicating a change in vasoactive substances that control resistance arteriolar tone.

Augmentation of Pulmonary Perfusion by Conducted Effects of a Pulmonary Artery Ultrasound Catheter.

Ultrasound (US) generated by catheters used clinically for US-facilitated thrombolysis can release shear-dependent vasodilators from endothelial and red blood cells. We hypothesized that catheter-based US in the pulmonary artery (PA) decreases downstream vascular resistance and increases pulmonary blood flow. In rhesus macaques, a U.S. Food and Drug Administration-approved multi-element US catheter was placed in a pulmonary artery. Comprehensive echocardiography was performed (i) at baseline, (ii) during hypoxemia (12% FIO2) to increase pulmonary vascular resistance (PVR) and (c) 15 min after initiating US during hypoxemia. Reduced FIO2 produced intended reductions in oxygen saturation (69 ± 3%) and PaO2 (34 ± 5 mm Hg), yet on echocardiography, hypoxemia did not create the intended model, with only modest hypoxia-related increases in PA systolic pressure (24 ± 4 to 28 ± 4 mm Hg, p = 0.05) and no significant change in PVR or multiparametric right ventricular (RV) function. Although US did not further change total PVR, on 99mTc-macroalbumin aggregate single-photon-emission computed tomography imaging, lung perfusion was significantly higher in the lung ipsilateral to the US catheter versus the contralateral control lung (133 ± 48 cpm vs. 103 ± 43 × 103 cpm, p = 0.01). We conclude that PA catheter-based US increases regional lung perfusion, most likely from vasodilators that are conducted downstream.

Systemic polidocanol from intravenous or pressurized intrauterine administration produces reversible cardiovascular toxicity.

Objective: Fatal allergic responses and cardiac arrhythmias have been reported with the intravenous (IV) administration of polidocanol. We sought to identify the physiologic mechanism of systemic cardiovascular response after transcervical (TC) and IV administration of polidocanol. Methods: We continuously monitored blood pressure (BP) and heart rate using an arterial line during IV and intraperitoneal (IP) administration of polidocanol solution (PS) and polidocanol doxycycline solution in female rats and TC and IP administration of polidocanol foam (PF) and PDF (TC only) in female baboons. We performed TC procedures using a catheter with (pressurized) and without (nonpressurized) balloon inflation. Baboons also underwent monitoring during IV PS administration with and without pretreatment with antihistamines. We performed cardiac echo and electrocardiograms during selected experiments. We defined a refractory hypotension as a sustained decrease of more than 30% from baseline that prevented delivery of the target dose. Results: We found a dose-related increase in the proportion of baboons that developed refractory hypotension during TC administration of 5% PDF and PF, an effect confined to pressurized administration. The infusion of 0.5% PS in rats induced a rapid and dramatic refractory hypotension. The inclusion of doxycycline did not improve or deteriorate these outcomes, and doxycycline solution or saline (control) alone did not affect BP. All five female baboons that received up to 20 mL of 1% PS (200 mg) developed refractory hypotension. Pretreatment with diphenhydramine, ranitidine, or both did not block the refractory hypotension induced by IV administration of 1% PS (100 mg). In contrast, only one of the six female baboons treated with IP PF 400 mg developed a decrease of more than 30% in BP, and this response was not sustained. Cardiac echocardiography done in four baboons during TC treatment demonstrated a decrease in cardiac output as the physiologic mechanism of hypotension. We did not observe important changes on the electrocardiograms. Conclusions: Adverse cardiovascular effects of polidocanol treatment occur owing to a direct myocardial effect of polidocanol and not as a result of a hypersensitivity reaction. Pressurized TC administration of PF results in refractory hypotension owing to endometrial vascular uptake of polidocanol and not as a result of uptake from peritoneal surfaces.

Ibrutinib Inhibits BMX-Dependent Endothelial VCAM-1 Expression In Vitro and Pro-Atherosclerotic Endothelial Activation and Platelet Adhesion In Vivo.

Introduction: Inflammatory activation of the vascular endothelium leads to overexpression of adhesion molecules such as vascular cell adhesion molecule-1 (VCAM-1), contributing to the pro-thrombotic state underpinning atherogenesis. While the role of TEC family kinases (TFKs) in mediating inflammatory cell and platelet activation is well defined, the role of TFKs in vascular endothelial activation remains unclear. We investigated the role of TFKs in endothelial cell activation in vitro and in a nonhuman primate model of diet-induced atherosclerosis in vivo. Methods and Results: In vitro, we found that ibrutinib blocked activation of the TFK member, BMX, by vascular endothelial growth factors (VEGF)-A in human aortic endothelial cells (HAECs). Blockade of BMX activation with ibrutinib or pharmacologically distinct BMX inhibitors eliminated the ability of VEGF-A to stimulate VCAM-1 expression in HAECs. We validated that treatment with ibrutinib inhibited TFK-mediated platelet activation and aggregation in both human and primate samples as measured using flow cytometry and light transmission aggregometry. We utilized contrast-enhanced ultrasound molecular imaging to measure platelet GPIbα and endothelial VCAM-1 expression in atherosclerosis-prone carotid arteries of obese nonhuman primates. We observed that the TFK inhibitor, ibrutinib, inhibited platelet deposition and endothelial cell activation in vivo. Conclusion: Herein we found that VEGF-A signals through BMX to induce VCAM-1 expression in endothelial cells, and that VCAM-1 expression is sensitive to ibrutinib in vitro and in atherosclerosis-prone carotid arteries in vivo. These findings suggest that TFKs may contribute to the pathogenesis of atherosclerosis and could represent a novel therapeutic target.

Echocardiographic Ischemic Memory Molecular Imaging for Point-of-Care Detection of Myocardial Ischemia.

BACKGROUND: Noninvasive molecular imaging of recent ischemia can potentially be used to diagnose acute coronary syndrome (ACS) with high accuracy. OBJECTIVES: The authors hypothesized that bedside myocardial contrast echocardiography (MCE) ischemic memory imaging could be achieved with phosphatidylserine microbubbles (MBPS) that are retained in the microcirculation via ischemia-associated endothelial activation. METHODS: A dose-finding study was performed in healthy volunteers (n = 17) to establish optimal MBPS dosing. Stable patients with ACS (n = 30) and confirmed antecedent but resolved myocardial ischemia were studied within 2 hours of coronary angiography and percutaneous coronary intervention (PCI) when indicated. MCE molecular imaging was performed 8 minutes after intravenous administration of MBPS. MCE perfusion imaging was used to assess the status of the postischemic microcirculation. RESULTS: Based on dose-finding studies, 0.10 or 0.15 mL of MBPS based on body mass was selected. In patients with ACS, all but 2 underwent primary PCI. MCE molecular imaging signal intensity was greater in the postischemic risk area vs remote territory (median [95% CI]: 56 [33-66] vs 8 [2-17] IU; P < 0.001) with a receiver-operating characteristic curve C-statistic of 0.94 to differentiate post-ischemic from remote territory. Molecular imaging signal in the risk area was not related to type of ACS (unstable angina: 3; non-ST-segment elevation myocardial infarction: 14; ST-segment elevation myocardial infarction: 13), peak troponin, time to PCI, post-PCI myocardial perfusion, GRACE (Global Registry of Acute Coronary Events) score, or HEART score. CONCLUSIONS: Molecular imaging with point-of-care echocardiography and MBPS can detect recent but resolved myocardial ischemia. This bedside technique requires only minutes to perform and appears independent of the degree of ischemia. (Ischemic Memory Imaging With Myocardial Contrast Echocardiography; NCT03009266).

High-intensity ultrasound catheter ablation achieves deep mid-myocardial lesions in vivo.

BACKGROUND: Radiofrequency ablation of epicardial and mid-myocardial ventricular arrhythmias is limited by lesion depth. OBJECTIVE: The purpose of this study was to generate deep mid-interventricular septal (IVS) lesions using high-intensity ultrasound (US) from an endocardial catheter-based approach. METHODS: Irrigated US catheters (12 F) were fabricated with 3 × 5 mm transducers of 5.0, 6.5, and 8.0 MHz frequencies and compared in an ex vivo perfused myocardial ablation model. In vivo septal ablation in swine (n = 12) was performed via femoral venous access to the right ventricle. Lesions were characterized by echocardiography, cardiac magnetic resonance imaging, and electroanatomic voltage mapping pre- and post-ablation, and at 30 days. Four animals were euthanized immediately post-ablation to compare acute and chronic lesion histology and gross pathology. RESULTS: In ex vivo models, maximal lesion depth and volume was achieved by 6.5 MHz catheters, which were used in vivo. Lesion depth by gross pathology was similar post-ablation (10.8 mm; 95% confidence interval [CI] 9.9-12.4 mm) and at 30 days (11.2 mm; 95% CI 10.6-12.4 mm) (P = .56). Lesion volume decreased post-ablation to 30 days (from 255 [95% CI 198-440] to 162 [95% CI 133-234] mm3; P = .05), yet transmurality increased from 58% (95% CI 50%-76%) to 81% (95% CI 74%-93%), attributable to a reduction in IVS thickness (from 16.0 ± 1.7 to 10.6 ± 2.4 mm; P = .007). Magnetic resonance imaging confirmed dense septal ablation by delayed enhancement, with increased T1 time post-ablation and at 30 days and increased T2 time only post-ablation. Voltage mapping of both sides of IVS demonstrated reduced unipolar (but not bipolar) voltage along the IVS. CONCLUSION: High-intensity US catheter ablation may be an effective treatment of mid-myocardial or epicardial ventricular arrhythmias from an endocardial approach.

Augmentation of Tissue Perfusion with Contrast Ultrasound: Influence of Three-Dimensional Beam Geometry and Conducted Vasodilation.

BACKGROUND: Cavitation of microbubble contrast agents with ultrasound produces shear-mediated vasodilation and an increase in tissue perfusion. We investigated the influence of the size of the cavitation volume by comparing flow augmentation produced by two-dimensional (2D) versus three-dimensional (3D) therapeutic ultrasound. We also hypothesized that cavitation could augment flow beyond the ultrasound field through release of vasodilators that are carried downstream. METHODS: In 11 rhesus macaques, cavitation of intravenously administered lipid-shelled microbubbles was performed in the proximal forearm flexor muscles unilaterally for 10 min. Ultrasound cavitation (1.3 MHz, 1.5 MPa peak negative pressure) was performed with 2D or 3D transmission with beam elevations of 5 and 25 mm, respectively, and pulsing intervals (PIs) sufficient to allow complete postdestruction refill (5 and 12 sec for 2D and 3D, respectively). Contrast ultrasound perfusion imaging was performed before and after cavitation, using multiplane assessment within and beyond the cavitation field in 1.5-cm increments. Cavitation in the hindlimb of mice using 2D ultrasound at a PI of 1 or 5 sec was performed to examine microvascular flow changes from cavitation in only arteries versus the microcirculation. RESULTS: In primates, the degree of muscle flow augmentation in the center of the cavitation field was similar for 2D and 3D conditions (five- to sixfold increase for both, P < .01 vs baseline). The spatial extent of flow augmentation was only modestly greater for 3D cavitation because of an increase in perfusion with 2D transmission that was detected outside of the cavitation field. In mice, cavitation in the microvascular compartment (PI 5 sec) produced the greatest degree of flow augmentation, yet cavitation in the arterial compartment (PI 1 sec) still produced a three- to fourfold increase in flow (P < .001 vs control). The mechanism for flow augmentation beyond the cavitation zone was investigated by in vitro studies that demonstrated cavitation-related release of vasodilators, including adenosine triphosphate and nitric oxide, from erythrocytes and endothelial cells. CONCLUSIONS: Compared with 2D transmission, 3D cavitation of microbubbles generates a similar degree of muscle flow augmentation, possibly because of a trade-off between volume of cavitation and PI, and only modestly increases the spatial extent of flow augmentation because of the ability of cavitation to produce conducted effects beyond the ultrasound field.

Treatment of Limb Ischemia with Conducted Effects of Catheter-Based Endovascular Ultrasound.

Ultrasound (US) is known to stimulate endogenous shear-dependent pathways, and can lower microvascular resistance through mediators that are conducted downstream from US exposure. We hypothesized that endovascular US, already in use for thrombolysis in humans, can improve tissue perfusion in the setting of acute limb ischemia through downstream-conducted effects. Models of severe peripheral arterial disease were developed in mice and in rhesus macaques. An endovascular US catheter (2.3 MHz, 0.5-1.1 MPa) was used to expose the limb adductor in mice for 10 min or the femoral artery distal to stenosis in macaques for 15 min. Quantitative contrast-enhanced ultrasound perfusion imaging was performed to assess flow augmentation in the adductor muscle of mice and the calf muscle of macaques. Microvascular blood flow in the ischemic limb relative to the contralateral control limb was reduced to 22 ± 8% in mice and 36 ± 20% in macaques. US produced immediate 2.3- and 3-fold increases (p < 0.05) in the murine and macaque ischemic limbs, respectively. In macaques, perfusion in the ischemic limb was increased to a normal level. We conclude that non-cavitating US produced by endovascular catheters that are used to enhance thrombolysis in humans can reduce vascular resistance and increase limb perfusion in the setting of acute ischemia.

Arterial Platelet Adhesion in Atherosclerosis-Prone Arteries of Obese, Insulin-Resistant Nonhuman Primates.

Background Platelet-endothelial interactions are thought to contribute to early atherogenesis. These interactions are potentiated by oxidative stress. We used in vivo molecular imaging to test the hypothesis that platelet-endothelial interactions occur at early stages of plaque development in obese, insulin-resistant nonhuman primates, and are suppressed by NADPH-oxidase-2 inhibition. Methods and Results Six adult rhesus macaques fed a Western-style diet for a median of 4.0 years were studied at baseline and after 8 weeks of therapy with the NADPH-oxidase-2-inhibitor apocynin (50 mg/kg per day). Six lean control animals were also studied. Measurements included intravenous glucose tolerance test, body composition by dual-energy X-ray absorptiometry, carotid intimal medial thickness, carotid artery contrast ultrasound molecular imaging for platelet GPIbα (glycoprotein- Ibα) and vascular cell adhesion molecule-1, and blood oxidative markers on mass spectrometry. Compared with lean controls, animals on a Western-style diet were obese (median body mass: 16.0 versus 8.7 kg, P=0.003; median truncal fat: 49% versus 20%, P=0.002), were insulin resistant (4-fold higher insulin-glucose area under the curve on intravenous glucose tolerance test, P=0.002), had 40% larger carotid intimal medial thickness (P=0.004), and exhibited oxidative signatures on proteomics. In obese but not lean animals, signal enhancement on molecular imaging was significantly elevated for GPIbα and vascular cell adhesion molecule-1. The signal correlated modestly with intimal medial thickness but not with the degree of insulin resistance. Apocynin significantly (P<0.01) reduced median signal for GPIbα by >80% and vascular cell adhesion molecule-1 signal by 75%, but did not affect intimal medial thickness, body mass, or intravenous glucose tolerance test results. Conclusion In nonhuman primates, diet-induced obesity and insulin resistance leads to platelet-endothelial adhesion at early atherosclerotic lesion sites, which is associated with the expression of pro-inflammatory adhesion molecules. These responses appear to be mediated, in part, through oxidative pathways.

Augmentation of Tissue Perfusion in Patients With Peripheral Artery Disease Using Microbubble Cavitation.

OBJECTIVES: The authors investigated ideal acoustic conditions on a clinical scanner custom-programmed for ultrasound (US) cavitation-mediated flow augmentation in preclinical models. We then applied these conditions in a first-in-human study to test the hypothesis that contrast US can increase limb perfusion in normal subjects and patients with peripheral artery disease (PAD). BACKGROUND: US-induced cavitation of microbubble contrast agents augments tissue perfusion by convective shear and secondary purinergic signaling that mediates release of endogenous vasodilators. METHODS: In mice, unilateral exposure of the proximal hindlimb to therapeutic US (1.3 MHz, mechanical index 1.3) was performed for 10 min after intravenous injection of lipid microbubbles. US varied according to line density (17, 37, 65 lines) and pulse duration. Microvascular perfusion was evaluated by US perfusion imaging, and in vivo adenosine triphosphate (ATP) release was assessed using in vivo optical imaging. Optimal parameters were then used in healthy volunteers and patients with PAD where calf US alone or in combination with intravenous microbubble contrast infusion was performed for 10 min. RESULTS: In mice, flow was augmented in the US-exposed limb for all acoustic conditions. Only at the lowest line density was there a stepwise increase in perfusion for longer (40-cycle) versus shorter (5-cycle) pulse duration. For higher line densities, blood flow consistently increased by 3-fold to 4-fold in the US-exposed limb irrespective of pulse duration. High line density and long pulse duration resulted in the greatest release of ATP in the cavitation zone. Application of these optimized conditions in humans together with intravenous contrast increased calf muscle blood flow by >2-fold in both healthy subjects and patients with PAD, whereas US alone had no effect. CONCLUSIONS: US of microbubbles when using optimized acoustic environments can increase perfusion in limb skeletal muscle, raising the possibility of a therapy for patients with PAD. (Augmentation of Limb Perfusion With Contrast Ultrasound; NCT03195556).

Limb Perfusion During Exercise Assessed by Contrast Ultrasound Varies According to Symptom Severity in Patients with Peripheral Artery Disease.

BACKGROUND: In patients with peripheral artery disease (PAD), the severity of symptoms correlates poorly with ankle-brachial index (ABI). The aim of this study was to test the hypothesis that limb perfusion assessed using contrast-enhanced ultrasound (CEU) during contractile exercise varies according to functional class in patients with PAD, particularly those with ABIs in the 0.4 to 0.6 range whose symptoms vary widely. METHODS: Bilateral quantitative CEU perfusion imaging of the calf was performed in normal control subjects (n = 10) and patients with PAD who had at least one limb with a moderately reduced ABI (0.4-0.6; n = 17). Imaging was performed at rest and immediately after 30 sec of modest periodic (0.3-Hz) plantar flexion (10 W). RESULTS: In patients with PAD, Rutherford symptom classification for each limb varied widely, including in limbs with ABIs of 0.4 to 0.6 (n = 6 with mild or no symptoms, n = 14 with moderate to severe symptoms). CEU perfusion imaging parameters at rest were similar between control subjects and patients with PAD irrespective of ABI. In normal control subjects, limb flow increased on average by > 20-fold after only 30 sec of moderate exercise. In patients with PAD, muscle exercise perfusion for all limbs was reduced compared with control subjects and decreased according to the severity of ABI reduction, primarily from reduced microvascular flux rate. Even limbs with ABIs > 0.9 in patients with PAD had lower exercise perfusion than in control subjects (P = .03). In subjects with PAD, exercise perfusion was lower in those with moderate to severe versus mild symptoms when analyzed for all limbs (median, 30 IU/sec [interquartile range (IQR), 21-52 IU/sec] vs 84 IU/sec [IQR, 36-177 IU/sec]; P = .01) and limbs with ABIs of 0.4 to 0.6 (median, 26 IU/sec [IQR, 14-41 IU/sec] vs 54 IU/sec [IQR, 31-105 IU/sec]; P = .05). CONCLUSIONS: In patients with PAD, CEU exercise perfusion imaging detects differences in limb muscle perfusion that are likely to be responsible for differences in symptom severity and can detect the flow abnormalities from microvascular dysfunction even in limbs with normal ABIs.