Multiple coronary artery fistulae communicating through a sinusoid and emptying into the left ventricle.

This case report describes the occurrence of multiple coronary artery fistulae emptying into the left ventricle and includes a small communication into the left atrium. The initial diagnosis of a coronary artery fistula was made by standard and nonstandard transthoracic two-dimensional echocardiogram and Doppler interrogation. Later, multiple coronary fistulae communicating through a sinusoid and draining into the left-sided chambers were confirmed by angiography.

Gadofluorine-enhanced magnetic resonance imaging of carotid atherosclerosis in Yucatan miniswine.

OBJECTIVE: The aim of this study was to determine whether gadofluorine, a paramagnetic magnetic resonance imaging (MRI) contrast agent, selectively enhances carotid atherosclerotic plaques in Yucatan miniswine. METHODS: Atherosclerotic plaques were induced in the left carotid arteries (LCA) of Yucatan miniswine (n=3) by balloon denudation and high cholesterol diet. T1-weighted MRI was performed before and 24 hours after gadofluorine injection (at a dose of 100 micromol/kg) to assess the enhancement of the balloon-injured LCA wall relative to healthy, uninjured right carotid artery (RCA) wall. Histopathology was performed to verify the presence and composition of the atherosclerotic plaques imaged with MRI. RESULTS: Gadofluorine was found to enhance LCA atherosclerotic lesions relative to RCA wall by 21% (P<0.025) 24 hours after contrast injection. Enhancement of healthy LCA wall relative to healthy RCA wall was not observed. CONCLUSION: Gadofluorine selectively enhances carotid atherosclerotic plaques in Yucatan miniswine. Gadofluorine appears to be a promising MR contrast agent for detection of atherosclerotic plaques in vivo.

Regional material property alterations in porcine femoral arteries with atheroma development.

We have developed a novel methodology that permits assessment of regional vascular mechanical property alterations in the presence of atheroma in vivo employing a Yucatan miniswine model with induced lesions. Femoral arteries were imaged with intravascular ultrasound. Image data were segmented and, following three-dimensional reconstruction, underwent finite element and sensitivity analysis with optimization to identify regions with altered vascular mechanical properties. All regions were compared to histological analysis. In 12 animals with 8 weeks of endothelial cell denudation and high cholesterol diet (induced atherosclerosis), the elastic modulus initially decreased with early lesion development and then increased with increasing fibrosis-(elastic modulus-all values x10(4)Pa-mean+/-SEM) histologically normal (non-denuded control segment) elements 9.73+/-0.01, fatty elements 9.53+/-0.01, fibrofatty elements 9.41+/-0.03, and fibrous elements 9.68+/-0.02 (all p<0.001 vs. normal elements). Wall thickness, however, increased with atheroma formation. These data demonstrate decreasing vascular material properties with early lesions, followed by an increase as lesions progress. This methodology permits determination of areas with early atheroma development, follow atheroma progression, and potentially evaluate interventions aimed at decreasing atheroma load and normalizing vascular material properties.

Porcine carotid arterial material property alterations with induced atheroma: an in vivo study.

OBJECTIVE: A novel methodology has been developed to evaluate regional alterations in arterial wall material properties with induced atheroma in an animal model. METHODS: Atheromatous lesions (fatty, fibro-fatty, and fibrous) were induced in the carotid arteries of a Yucatan miniswine model by endothelial cell denudation and high cholesterol diet. The images at base line and 8 weeks after denudation were obtained using intravascular ultrasound (IVUS) imaging along with hemodynamic data. Finite element analysis (FEA) along with optimization was employed to assess regional alterations in elastic modulus in the presence of atheroma confirmed by histology. RESULTS: In animals with 8 weeks of induced atherosclerosis, the elastic modulus increased-(elastic modulus-all values x 10(4) Pa, mean+/-S.D.) normal elements (9.34+/-0.36) compared to abnormal elements (9.52+/-0.36) (p<0.05 versus normal elements). Wall thickness increased with atheroma formation. These data demonstrate stiffening vascular wall elastic modulus with lesion progression. This is different from the behavior of femoral arteries, where the elastic modulus decreases with early stages of atheroma development followed by an increase as lesions progress. CONCLUSIONS: This methodology permits determination of areas with early atheroma development, follow atheroma progression, and potentially evaluate interventions aimed at decreasing atheroma load and normalizing vascular material properties.

Computer three-dimensional reconstruction of the atrioventricular conduction system.

The human atrioventricular conduction system (AVCS), which includes the AV node and its approaches, AV bundle (penetrating, branching, and bifurcating parts), and the bundle branches, is a curved complex structure that has not been reconstructed in three dimensions using computer technology. Microscopic slides of every 40(th) serial section (cut at 7 micron level) of the AVCS were digitized into 600 dots/inch color images. External outlines of each section were manually segmented using commercially available three-dimensional rendering software (Rhinoceros). The AVCS was traced from light microscopy and superimposed onto the external outlines. To account for inherent errors in histological slide preparation, an optimization procedure was used to align external outlines of all sections. The optimal rotation and translation of each section was established by maximizing area of overlap between adjacent sections. A sequential one-dimensional minimization algorithm was used for optimization. Rotation and translation values were then used to align external outlines and the superimposed conduction system, reconstructing the AVCS in three-dimensions. To validate the method, the algorithm was applied to a digitized image transformed with known translations and rotations. The validation procedure demonstrated that each test image aligned in translations and to within 0.01 degree in rotations. Spatial error determined by resolution of the digitized images was +/-0.5/600 inch (+/-21 microns). Three-dimensional reconstruction of every 40th serial section clearly demonstrated the complex curved shape of the AVCS. Three-dimensional reconstruction of the human and canine AVCS permits accurate pathological and electrophysiological correlation of the conduction system.

Alteration in fluid mechanics in porcine femoral arteries with atheroma development.

Although the relationship between the local wall shear stresses (WSS) and atheroma development has been previously studied, the effect of variable regional wall distensibility with early atherosclerotic lesions and its effect on local hemodynamics have not been well investigated. We induced early atherosclerotic lesion development in one femoral artery in a Yucatan miniswine model with the contralateral artery serving as control. Eight weeks following atheroma formation, hemodynamic and intravascular ultrasound image data were obtained. Using the measured regional arterial distension as the moving boundary condition, unsteady laminar incompressible computational analysis was performed on the reconstructed arterial segments. With the development of early atherosclerotic lesions, there was increased wall distensibility and an increase in the computed normalized wall shear stress amplitude (NWSA). Alterations in the local fluid mechanics and mass transport at these sites may need to be considered in our understanding of the continued growth of these lesions.

Intravascular ultrasound molecular imaging of atheroma components in vivo.

OBJECTIVES: Our purpose was to quantitate and confirm specific echogenic immunoliposome (ELIP) atheroma component enhancement in vivo. BACKGROUND: Targeted ELIPs for ultrasonic detection and staging of active molecular components of endothelium and atheroma have been developed. METHODS: In Yucatan miniswine, the endothelium was injured from one femoral and one carotid artery, and animals were fed a high-cholesterol diet for two months to create various stages of atheroma. Arteries were imaged with intravascular ultrasound (IVUS) 5 and 10 min after ELIP injection (5-mg dose). Anti-intercellular adhesion molecule-1 (ICAM-1), anti-vascular cell adhesion molecule-1 (VCAM-1), anti-fibrin, anti-fibrinogen, and anti-tissue factor (TF) conjugated ELIPs were used, and immunohistochemistry (IHC) confirmed the presence or absence of molecular expression. Two blinded observers determined if each segment was enhanced by ELIP. Three-dimensional image reconstruction and videodensitometric analysis determined the mean gray-scale (MGS) change of the luminal border. RESULTS: To determine endothelial injury component enhancement, anti-fibrinogen ELIP enhanced exposed fibrin in all arteries (MGS increased 22 +/- 5%; 6 arteries; 2 animals). To determine enhancement of molecular components in atherosclerotic arteries, observers detected enhancement 5 min after anti-VCAM, anti-ICAM, anti-TF, anti-fibrin, and anti-fibrinogen conjugated ELIPs. Furthermore, ELIP enhanced atheroma MGS by 39 +/- 18% (n = 8). The IHC staining confirmed the expression of respective molecular targets in all enhanced segments. CONCLUSIONS: It was shown that ELIPs specifically enhance endothelial injury/atheroma components. This allows better characterization of the type and extent of active atheroma components and may allow more directed therapy.

In vitro characterization of liposomes and Optison by acoustic scattering at 3.5 MHz.

Liposomes are phospholipid vesicles that can encapsulate both gas and fluid. With antibody conjugation, new formulations, known as immunoliposomes, can be targeted to atheroma and other pathologic components and are, thus, being developed as novel diagnostic ultrasound (US) echo contrast agents to enhance atherosclerosis imaging. The majority of these echogenic liposomes range in diameter from 0.25 to 5.0 microm. To quantify the echogenicity of liposome suspensions of varying concentrations, the backscattering coefficient at 3.5 MHz was determined experimentally. The backscattering coefficient was also estimated theoretically as a function of air volume fraction by modeling the encapsulated air as a free air bubble and assuming single bubble scattering. For most of the liposome concentrations examined in this study (on the order of 10(8)/mL), the backscattering coefficient equals or exceeds that of Optison at the human clinical dosage (on the order of 10(4)/mL). Experimental measurement of the decrease in backscattering coefficient shows promise as a sensitive method for determining whether liposomes are left intact or destroyed during imaging; thus, helping to explore their potential as a vehicle for targeted drug delivery. In addition, the attenuation of US through liposome suspensions is negligible at 3.5 MHz relative to the attenuation through Optison (0.25 dB/cm), suggesting that liposomes have a much higher scatter-to-attenuation ratio and could be more efficient as contrast agents.

Left ventricular thrombus enhancement after intravenous injection of echogenic immunoliposomes: studies in a new experimental model.

BACKGROUND: Targeted echogenic immunoliposomes (ELIPs) for ultrasound enhancement of atheroma components have been developed. To date, ELIP delivery has been intra-arterial. To determine whether ELIPs can be given intravenously with enhancement of systemic structures, a left ventricular thrombus (LVT) model was developed. METHODS AND RESULTS: In 6 animals plus 1 dose-ranging animal, the apical coronary arteries were ligated, and an LVT was produced by injecting Hemaseel fibrin adhesive through the apical myocardium. The thrombus was imaged epicardially and transthoracically at 0, 1, 5, and 10 minutes after anti-fibrinogen ELIP injections. The dose of ELIPs was varied. PBS and unconjugated ELIPs were controls. The apical thrombi were easily reproduced and clearly visible with epicardial and transthoracic ultrasound. Enhancement occurred with 2 mg anti-fibrinogen ELIPs and increased with dose. With 8 mg ELIPs, enhancement was different from control within 10 minutes (P<0.05). Rhodamine-labeled anti-fibrinogen ELIPs were seen with fluorescence microscopy of the LVT. Blinded viewing detected enhancement by 10 minutes in all animals after anti-fibrinogen ELIPs. CONCLUSIONS: We describe an easily reproducible LVT model. Anti-fibrinogen ELIPs delivered intravenously, as a single-step process, rapidly enhance the ultrasound image of a systemic target. This allows for future development of ELIPs as a targeted ultrasound contrast agent.

Physical correlates of the ultrasonic reflectivity of lipid dispersions suitable as diagnostic contrast agents.

The objective of this study was to determine the physical basis of ultrasound (US) reflectivity of echogenic lipid dispersions. These dispersions were made using a process previously described involving sonication of the lipid in water, addition of mannitol, freezing, lyophilization and rehydration. The component lipids were egg phosphatidylcholine, dipalmitoylphosphatidylethanolamine, dipalmitoylphosphatidylglycerol and cholesterol in a molar ratio of 69:8:8:15. Ultrasound reflectivity, as assessed with a 20-MHz intravascular US catheter and analyzed using computer-assisted videodensitometry, was found to be sensitive to variations in ambient pressure; echogenicity was greatly reduced by exposure to 0.5 atm vacuum for 10 s or 1.5 atm pressure for 10 s. Pressure changes of the magnitude that obtain in the arterial circulation had little effect on echogenicity. Vacuum treatment resulted in the release of approximately 100 microL air from a standard preparation of 10 mg lipid in 1 mL. Maximum ultrasonic reflectivity required the presence of 0.1-0.2 mol/L mannitol during the lyophilization step; mere addition of mannitol to the lipid lyophilized in the absence of mannitol produced nonreflective dispersions. Inclusion of sodium phosphate or other electrolytes reduced echogenicity. High echogenicity was associated with the presence of large-volume freeze-dried cakes and fusion of liposomes (which led to a 10 times increase in liposome diameters) during freezing before lyophilization. Lyophilization from water led to liposome fusion, but the cakes were small and US reflectivity was weak. Lyophilization from solutions of cryoprotectants such as trehalose produced large cakes, but little liposome fusion and also led to weak US reflectivity. Filtration through defined pores revealed that approximately 50% of the echogenicity originated from particles smaller than 1 microm and about 2/3 from particles smaller than 3 microm. These results indicate that lyophilization from 0.2 mol/L mannitol solution generates a disrupted array of lipid bilayers that, upon rehydration, fuse and trap small amounts of air distributed among liposome-size particles.

Evaluation of valvular regurgitation severity using digital acquisition of echocardiographic images.

BACKGROUND: Digital acquisition is a technique for storing echocardiographic data that offers advantages over conventional videotape (VT); however, limited information is available on its accuracy for the evaluation of valvular regurgitation. METHODS: We evaluated 102 patients with at least 1 regurgitant lesion. Data were obtained on VT and in 1 cardiac cycle stored digitally (1C). To assess for incremental improvement with acquisition of multiple cycles, digital images were also acquired with 2 (2C) or 3 cardiac cycles (3C). Both digital and VT images were graded for regurgitant severity as absent, trivial, mild, moderate, or severe. Kappa statistics were used to assess agreement. RESULTS: A total of 171 valvular regurgitant lesions (mild or greater) were evaluated. The overall agreement between 1C and VT images was kappa = 0.61. With multiple cycle acquisition, there was no improvement in agreement (kappa = 0.56 and 0.57 for 2C and 3C, respectively). When subgrouped, the level of agreement between 1C and VT was slightly lower for the aortic valve than for the mitral or tricuspid valves (kappa = 0.49, 0.63, 0.64, respectively). CONCLUSION: The 1C technique has substantial agreement and correlation with standard VT for the evaluation of regurgitant lesions with the use of color flow Doppler. The acquisition of multiple cardiac cycles does not provide incremental improvement over single beat acquisition.

A physiologic flow chamber model to define intravascular ultrasound enhancement of fibrin using echogenic liposomes.

RATIONALE AND OBJECTIVES: Echogenic immunoliposomes (ELIP) for enhancement of vasoactive and pathologic components of endothelium and atherosclerosis have been developed. A physiologic flow chamber model has been developed to define intravascular ultrasound enhancement of a fibrin interface. METHODS: A IgG ELIP was used, which nonspecifically associated with fibrin, to demonstrate the suitability of this model. With varying doses of IgG ELIP, the fibrin wells were imaged at 1, 2, 4, 6, and 9 minutes. RESULTS: IgG ELIP enhanced fibrin versus saline (P < 0.005) was visible at 1 minute, lasted at least 9 minutes, and at 6 minutes the interface enhanced 27% +/- 6.1%. Enhancement was caused by increases in interface thickness and brightness. Enhancement increased with dose up to 8 mg lipid (n = 4 per time point). CONCLUSION: This model can quantitate the components of IVUS enhancement of an interface produced by ELIP. This model may allow for further development and understanding of ELIP and other targeted ultrasound contrast agents.