Delivery of bevacizumab to atheromatous porcine carotid tissue using echogenic liposomes.

Ultrasound is both a valuable diagnostic tool and a promoter of beneficial tissue bioeffects for the treatment of cardiovascular disease. Vascular effects can be mediated by mechanical oscillations of circulating microbubbles that may also encapsulate and shield therapeutic agents in the bloodstream. Here, the effect of color-Doppler ultrasound exposure on bevacizumab-loaded liposome delivery into the vascular bed was assessed in atheromatous porcine carotids. Bevacizumab, an anti-angiogenic antibody to vascular endothelial growth factor (VEGF-A), was loaded into echogenic liposomes (BEV-ELIP) and confirmed to be immunoreactive. BEV-ELIP flowing within the lumen were exposed to color-Doppler ultrasound at three acoustic pressures for 3.5 min during treatment at physiologic temperature and fluid pressure. To confirm the presence of bubble activity, cavitation was detected within the lumen by a single-element passive cavitation detector. After treatment, the artery was fixed at physiologic pressure and subjected to immunohistochemical analysis to assess the penetration of bevacizumab within the carotid wall. The results suggest that other factors may more strongly influence the deposition of bevacizumab into carotid tissue than color-Doppler ultrasound and cavitation. In both sets of arteries, preferential accumulation of bevacizumab occurred in locations associated with atheroma progression and neointimal thickening: fibrous tissue, necrotic plaque and areas near macrophage infiltration. The delivery of bevacizumab to carotid vascular tissue correlated with the properties of the tissue bed, such as permeability, or affinity for growth-factor binding. Future investigations using this novel therapeutic strategy may focus on characterizing the spatial extent of delivery and bevacizumab colocalization with biochemical markers of atheroma.

Three-dimensional finite element analysis of residual stress in arteries.

Calculation of residual stress in arteries, using the analytical approach has been quite valuable in our understanding of its critical role in vascular mechanics. Stresses are calculated at the central section of an infinitely long tube by imposing a constant axial stretch while deforming the artery from the stress-free state to its unloaded state. However, segments used to perform opening-angle measurements have finite lengths. Further, the stress-free artery configuration is assumed to be circular. Experiments show that they are slightly noncircular. The numerical approach to residual stress calculation can allow us to study both these issues. Using 3D cylindrical geometries and an isotropic material model, we investigated how segment length can affect residual stress calculations and identified the appropriate segment length for experiments. Further, we recorded and used the true noncircular stress-free state of an artery segment, computed the residual stress distribution, and compared it to that from a similar, but circular segment. Our findings suggest that segment length must be ten times the wall thickness for it to be "long" enough. We also found that the circularity assumption may be a reasonable approximation for typical arteries.

A method for in-vivo analysis for regional arterial wall material property alterations with atherosclerosis: preliminary results.

Atherosclerosis is a diffuse arterial disease developing over many years and resulting in a complicated three-dimensional arterial morphology. The arterial wall material properties have been demonstrated to show regional alterations with atheroma development and growth. We present a mechanical analysis of diseased arterial segments reconstructed from intravascular ultrasound images in order to quantitatively identify regional alterations in the elastic constants with atherosclerotic lesions. We employ a finite element and a displacement sensitivity analysis to divide the arterial segment into regions with different material properties and use an optimization algorithm to identify the elastic constants in these regions. The results with regional variations identified with this method correlated qualitatively with the extent and location of atherosclerotic lesions identified by visual inspection of the affected arteries. The optimized elastic modulus in regions affected by early atherosclerotic lesions ranged from 90.9 to 93.0 kPa where as the corresponding magnitudes in normal arterial segments ranged from 97.9 to 101.0 kPa. This method can be potentially employed to identify the extent and location of atherosclerotic lesions in a systematic analysis and may potentially be used for the early detection of lesion growth.

Validation of transesophageal echocardiography to determine physiologic coronary flow.

UNLABELLED: Human studies have suggested that Doppler transesophageal echocardiography (TEE) can determine normal physiologic coronary blood flow (CBF) and alterations in CBF due to proximal flow-limiting stenoses. However, assessment of CBF by Doppler TEE has not been validated. To determine if true estimation of CBF could be obtained with Doppler TEE, seven mongrel dogs (weight range 28 kg-36 kg) were evaluated. Simultaneous CBF determinations by Doppler TEE and epicardial electromagnetic flow (EMF) and/or epicardial Doppler flow (EDF) probes were compared. Measurements were obtained at baseline and following varying degrees of proximal coronary occlusion, which produced reactive hyperemia. RESULTS: Consistent Doppler flow waveforms were obtainable by Doppler TEE in 34 different measurements during perturbations: Mean for TEE Flow (ml/min) = 85, EMF or EDF Flow (ml/min) = 53; Standard Deviation for TEE Flow (ml/min) = 45, EMF or EDF Flow (ml/min) = 38; Minimum for TEE Flow (ml/min) = 42, EMF or EDF Flow (ml/min) = 11; and Maximum for TEE Flow (ml/min) = 174, EMF or EDF Flow (ml/min) = 130. TEE Flow (ml/min) = 1.1 EMF/EDF flow + 26.3. There was a general trend towards overestimation of CBF by Doppler TEE. This study demonstrates that Doppler TEE is a promising method for obtaining measurements of CBF over the physiologic range.

Improving ultrasound reflectivity and stability of echogenic liposomal dispersions for use as targeted ultrasound contrast agents.

Targeted echogenic liposome dispersions for ultrasonic enhancement of vasoactive and pathological components of endothelium and atherosclerosis have recently been developed. The component lipids required for acoustic and targeting properties include phosphatidylcholine, phosphatidylethanolamine phosphatidylglycerol (PG), and cholesterol (CH), initially in a 60:8:2:30 mol % ratio. Component lipids, lyophilization, sugars, and freezing conditions were varied to optimize acoustic ultrasound reflectivity and acoustic stability. Echogenic liposome dispersions were made by using the dehydration-rehydration process. The lipid concentrations were varied (CH in the range 1 to 40 mol % and PG from 1 to 16 mol %). Variations in type and concentration of sugars were examined. The effect of freezing conditions and re-lyophilization was examined. Ultrasound reflectivity was assessed by using a 20-MHz intravascular ultrasound catheter and computer-assisted videodensitometry. Ultrasound reflectivity was optimized at a CH concentration of 10 mol %; PG concentration variation had essentially no effect on initial values of echogenicity. Optimal acoustic stability was observed with concentrations of 10-15 mol % CH and with a PG concentration greater than 4 mol %. Preparations made with 0.2 M mannitol were more ultrasound reflective than those made with lactose, trehalose, and sucrose. Re-lyophilization and freezing temperatures below -20 degrees C increased ultrasound reflectivity. We optimized the ultrasound properties of echogenic liposomal dispersions, the conditions of which provide some insight into the underlying lipid structures responsible. The preparations developed are now more stable and acoustically reflective than our previous preparations. This advances the development of echogenic lipid dispersions as targeted ultrasound contrast agents for use in general ultrasound as well as cardiovascular imaging.

Pulsatile flow simulation in arterial vascular segments with intravascular ultrasound images.

Previous studies have indicated a correlation between local variation in wall shear stress in arterial blood flow and atheroma development. The purpose of this study was to analyze the hemodynamics in vascular segments from morphologically realistic three-dimensional (3D) reconstruction, and to compare the computed wall shear stress in a compliant vascular segment model and the corresponding rigid walled model. Cross-sectional images of the segments of femoral and carotid arteries in five Yucatan miniswine were obtained using intravascular ultrasound (IVUS) imaging and the segment geometry was reconstructed at different times in the cardiac cycle. The actual measured wall motion from the reconstruction was employed to specify the moving boundaries for simulation of physiological distensibility. Velocity profiles and wall shear stress were computed using unsteady computational fluid dynamics analysis. The computed results revealed that the maximum wall shear stress in the compliant model was approximately 4-17 percent less than that in the rigid model if the wall motion is larger than 10 percent. Our analysis demonstrates that inaccuracies due to inflow velocity profile can be minimized by the extension of the model upstream. The phase angle between the diameter change and wall shear is affected by the local changes in geometry of the arteries. These simulations can be potentially used to analyze the effect of regional wall motion changes in the presence of atherosclerotic lesions on the local fluid dynamics and to correlate the same with subsequent growth of the lesions.

Structural evaluation of porcine heart valve prostheses with radiofrequency ultrasound.

BACKGROUND: Bioprosthetic heart valve use is limited by progressive degeneration. Early degenerative changes are often occult, making assessment of tissue integrity difficult. Ultrasound tissue characterization may detect alterations in tissue structure and allow early detection of leaflet degeneration. METHODS: Using a modified echocardiographic unit (Acuson), radiofrequency (RF) integrated backscatter amplitude (IBA) (integral/RF/dt) was measured in 38 leaflets from nine explanted and six control porcine valves. Regions of interest in each leaflet were studied using four ultrasound frequencies. Radiographic gray scale mean and leaflet thickness were measured at each region of interest. Percent collagen and mineral were calculated for each region of interest using color-image processing of histologic sections and compared to IBA. RESULTS: IBA values for control vs. explanted leaflets were (mean value+/-standard deviation): 8.2+/-4.69 dB vs. -4.7+/-4.64 dB at 7.0 MHz; -5.8+/-4.34 dB vs. -3.1+/-5.34 dB at 5.0 MHz; -3.8+/-3.38 dB vs. -2.1+/-3.18 dB at 3.5 MHz; and -9.0+/-4.58 dB vs. -7.1+/-4.25 dB at 2.5 MHz. Collagen content was 27.7+/-8.50% vs. 33.2+/-10.90%, mineral content was 0.1+/-0.10% vs. 2.1+/-4.30%, and radiographic gray scale mean was 150.6+/-1.96 vs. 145.3+/-5.14 for control vs. explanted leaflets, respectively. For control and explanted leaflets IBA, collagen content, mineral content, and radiographic gray scale mean were different (control vs. explanted P<0.05). Leaflet thickness was also noted to be different between the two groups. IBA was different among explanted leaflets with low, medium, and high mineral content. CONCLUSION: IBA was found to be a useful technique to differentiate normal from explanted porcine prosthetic valves in vitro. This method may be useful in the serial assessment of bioprosthetic leaflet degenerative properties in vivo.

The validation of volumetric real-time 3-dimensional echocardiography for the determination of left ventricular function.

The objective of this study was to validate a real-time 3-dimensional echocardiography (3DE) technique for the determination of left ventricular (LV) volume and ejection fraction (EF). In 10 mongrel dogs, an electromagnetic flow (EMF) probe was placed on the aorta, and the thorax was closed. Transthoracic imaging was performed during multiple hemodynamic conditions (n = 58) with simultaneous measurement of stroke volume (SV) with the use of EMF. From the volumetric data set, LV volumes were manually traced off-line by 2 independent observers with an apical rotation method (6 planes) and a conventional method (biplane) in a subset of conditions. This tracing technique was also evaluated in 18 human subjects in whom the calculated EF values were compared with values derived by multigated radionuclide angiography (MUGA). Excellent correlation and close limits of agreement were noted between SV measured by 3DE and EMF (r = 0.93) in dogs. In comparison with EMF-derived SV, 3DE provided better correlation than the biplane method (r = 0.93 versus r = 0.61). Interobserver and intraobserver variabilities were comparable (r = 0.94 and r = 0.94, respectively). In a comparison of MUGA-derived EF values and those obtained by 3DE in human subjects, 3DE provided better correlation than the biplane method (r = 0.94 versus r = 0.85). Real-time 3DE accurately measures left ventricular volumes transthoracically over a wide range of hemodynamic conditions in dogs and accurately determines EF in humans.

Functional comparison of transmyocardial revascularization by mechanical and laser means.

BACKGROUND: As a result of the clinical benefit observed in angina patients treated by transmyocardial revascularization (TMR) with a laser, interest in mechanical TMR has been renewed. Although the injury induced by mechanical TMR is similar to laser TMR, the resultant impact on myocardial contractility is unknown. The purpose of this study was to determine whether mechanical TMR improves ventricular function as compared with laser TMR in chronically ischemic myocardium. METHODS: After establishing an area of chronic myocardial ischemia, 25 domestic pigs were randomized to treatment by: excimer laser (group I), a hot needle (50 degrees C) (group II), a normothermic needle (group III), an ultrasonic needle (40 KHz) (group IV), or no treatment (group V). All devices create a transmural channel of the same diameter; 22 +/- 1 transmural channels were created in each animal. Regional myocardial contractility was assessed by measuring ventricular wall thickening at rest and with dobutamine stress echocardiography. Six weeks after revascularization, the animals were restudied at rest and with stress. Postsacrifice and histologic analysis of angiogenesis and TMR effects was then assessed. RESULTS: Laser TMR provided significant recovery of ischemic myocardial function. This improvement in contractility after laser TMR was a 75% increase over the baseline function of the ischemic zone (p < 0.01). Mechanical TMR provided no significant improvement in function posttreatment. In fact, TMR achieved with an ultrasonic needle demonstrated a 40% worsening of the contractility versus the pretreatment baseline (p < 0.05). Histologic analysis demonstrated a significant increase in new blood vessels in the ischemic zone after laser TMR, which was not demonstrated for any of the other groups (p < 0.05). Additionally, evaluation of the mechanical TMR channels demonstrated significant scarring, which correlated with the functional results. CONCLUSIONS: Using devices to create an injury analogous to the laser, mechanical TMR failed to improve the function of chronically ischemic myocardium. Only laser TMR significantly improved myocardial function.

Echocardiographic Doppler evaluation of left ventricular diastolic filling in older, highly trained male endurance athletes.

Previously published data have suggested that endurance training does not retard the normative aging impairment of early left ventricular diastolic filling (LVDF). Those studies, suggesting no effect of exercise training, have not examined highly trained endurance athletes or their LVDF responses after exercise. We therefore compared LVDF characteristics in a group of older highly trained endurance athletes (n = 12, mean age 69 years, range 65-75) and a group of sedentary control subjects (n = 12, mean age 69 years, range 65-73) with no cardiovascular disease. For all subjects, M-mode and Doppler echocardiographic data were obtained at rest. After baseline studies, subjects underwent graded, maximal cardiopulmonary treadmill exercise testing using a modified Balke protocol. Breath-by-breath respiratory gas analysis and peak exercise oxygen consumption (VO(2)max) measurements were obtained. Immediately after exercise and at 3-6 minutes into recovery, repeat Doppler echocardiographic data were obtained for determination of LVDF parameters. VO(2)max (44 +/-6.3 vs 27+/-4.2 ml/kg/min, P<0.001), oxygen consumption at anaerobic threshold (35+/-5.4 vs 24+/-3.8 ml/kg/min, P<0.001), exercise duration (24+/-3 vs 12+/-6 minutes, P<0.001), and left ventricular mass index (61+/-13 vs 51+/-7.8 kg/m(2), P<0.05) were greater in endurance athletes than in sedentary control subjects, whereas body mass index was lower (22+/-1.7 vs 26+/-3.4 kg/m(2), P<0.001). No differences in any of the LVDF characteristics were observed between the groups with the exception of a trend toward a lower atrial filling fraction at rest in the endurance athlete group versus the control subjects (P = 0.07). High-intensity endurance exercise training promotes exceptional peak exercise oxygen consumption and cardiovascular stamina but does not appear to alter normative aging effects on left ventricular diastolic function.

Development of echogenic, plasmid-incorporated, tissue-targeted cationic liposomes that can be used for directed gene delivery.

RATIONALE AND OBJECTIVES: Echogenic antibody-conjugated anionic liposomes have been developed that allow directed tissue targeting and acoustic enhancement. These are not efficient for gene delivery. A cationic formulation that allows directed gene delivery while retaining acoustic properties may provide more efficient transfection. METHODS: Cationic liposomes were prepared and acoustic reflectivity was determined. Anti-fibrinogen-conjugated liposomes were laid on fibrin-coated slides and adherence was quantified using fluorescence techniques. Liposomes were combined with a reporter gene and plated on cell cultures. Human umbilical vein endothelial cells were stimulated to upregulate intercellular adhesion molecule-1 (ICAM-1) and were treated with anti-ICAM-1-conjugated liposomes, and gene expression was quantified. RESULTS: Cationic liposomes retained their acoustic reflectivity and demonstrated specific adherence to fibrin under flow conditions. Significant transfection of human umbilical vein endothelial cells was demonstrated, with higher gene expression seen with specific antibody-conjugated liposomes. CONCLUSIONS: Novel acoustic cationic liposomes have been developed that can be antibody conjugated for site-specific adherence and directed cell modification. This presents exciting potential for a vector that allows tissue enhancement and targeted gene delivery.

In vivo targeting of acoustically reflective liposomes for intravascular and transvascular ultrasonic enhancement.

OBJECTIVES: The purpose of this study was to target acoustically reflective liposomes to atherosclerotic plaques in vivo for ultrasound image enhancement. BACKGROUND: We have previously demonstrated the development of acoustically reflective liposomes that can be conjugated for site-specific acoustic enhancement. This study evaluates the ability of liposomes coupled to antibodies specific for different components of atherosclerotic plaques and thrombi to target and enhance ultrasonic images in vivo. METHODS: Liposomes were prepared with phospholipids and cholesterol using a dehydration/ rehydration method. Antibodies were thiolated for liposome conjugation with N-succinimidyl 3-(2-pyridyldithio) propionate resulting in a thioether linkage between the protein and the phospholipid. Liposomes were conjugated to antifibrinogen or anti-intercellular adhesion molecule-1 (anti-ICAM-1). In a Yucatan miniswine model, atherosclerosis was developed by crush injury of one carotid and one femoral artery and ingestion of a hypercholesterolemic diet. After full plaque development the arteries were imaged (20-MHz intravascular ultrasound catheter and 7.5-MHz transvascular linear probe) after injection of saline, unconjugated liposomes and antibody conjugated liposomes. RESULTS: Conjugated liposomes retained their acoustically reflective properties and provided ultrasonic image enhancement of their targeted structures. Liposomes conjugated to antifibrinogen attached to thrombi and fibrous portions of the atheroma, whereas liposomes conjugated to anti-ICAM-1 attached to early atheroma. CONCLUSIONS: Our data demonstrate that this novel acoustic agent can provide varying targeting with different antibodies with retention of intravascular and transvascular acoustic properties.

In vitro targeting of acoustically reflective immunoliposomes to fibrin under various flow conditions.

We have previously demonstrated the development of acoustically reflective liposomes as a novel ultrasound contrast agent, that can be conjugated to antibodies for site specific acoustic enhancement of pathologically altered vascular tissue. The liposomes are echogenic due to the lipid composition, without gas entrapment, and have a size of less than one micron (Alkan-Onyuksel et al., 1996). When conjugated to anti-fibrinogen antibodies, the liposomes have the ability to attach to fibrin coated surfaces and thrombi in vitro as demonstrated by scanning electron microscopy and ultrasound imaging (Demos et al., 1997a). Anti-fibrinogen liposomes were shown to attach to fibrous atheroma and thrombi in a Yucatan miniswine model of induced atherosclerosis whereas liposomes conjugated to anti-intercellular adhesion molecule-1 (anti-ICAM-1) were demonstrated to target early stage atherosclerotic plaques (Demos et al., 1997b). The purpose of this study is to evaluate the binding characteristics of anti-fibrinogen liposomes in vitro under a variety of flow conditions in order to optimize the targeting ability of the immunoliposomes. Radiolabeled anti-fibrinogen liposomes were applied to fibrin coated filter paper and placed in a flow circuit under controlled flow conditions. Flow conditions were altered to study the effects of different shear stresses, temperature, plasma flow and pulsatile flow on the retention of liposomes to fibrin after set time periods. The retention of liposomes conjugated to polyclonal and monoclonal antibodies as well as Fab fragments made from monoclonal antibodies were compared. The binding characteristics of liposomes conjugated to different quantities of polyclonal antibodies were analyzed. At physiological shear stress of 1.5 N/m2 (15 dynes/cm2) over 70% of the liposomes remained attached to fibrin after two hours. A smaller and greater portion of the liposomes remained attached at higher and lower shear stresses respectively. Plasma components and temperature had no effect on liposomal retention whereas pulsatile flow resulted in a slight reduction in binding. Monoclonal antibodies showed a slight trend of reduced retention to fibrin over time as compared with polyclonal antibodies and Fab fragments. The quantity of antibody conjugated to the liposomes plays a role in liposome retention as demonstrated by the reduction in liposome retention caused by reducing the quantity of antibody conjugated to the liposomes. Anti-fibrinogen liposomes were retained to the fibrin surface to a large extent under all flow conditions likely to occur in vivo and therefore can provide site specific ultrasound contrast for a long enough time period to allow for imaging after injection.

Left ventricular functional improvement after transmyocardial laser revascularization.

BACKGROUND: Transmyocardial laser revascularization has been used to treat patients with end-stage coronary artery disease that is not amenable to standard revascularization. Although there is evidence of angina relief and quality of life enhancement, there is little information concerning improvement in myocardial contractility. The purpose of this study was to determine whether transmyocardial laser revascularization improves myocardial function in chronically ischemic myocardium. METHODS: In a model of chronic ischemia by Ameroid occlusion of the circumflex artery, domestic pigs (n = 8) were treated with transmyocardial laser revascularization. Before laser treatment, segmental contraction was assessed at rest and with dobutamine stress echocardiography. Myocardium subtended by the occlusion was compared with that remote from the occlusion. Six weeks after transmyocardial laser revascularization, the animals were restudied at rest and with stress, and then sacrificed. Sham-treated control animals (n = 4) underwent the same procedures but were not treated with transmyocardial laser revascularization. Control animals did not demonstrate significant recovery of function. RESULTS: Transmyocardial laser revascularization improved resting function in chronically ischemic myocardium by 100%. CONCLUSIONS: Transmyocardial laser revascularization significantly improves the function of chronically ischemic myocardium. These data may help explain the mechanisms by which transmyocardial laser revascularization is clinically effective.

A hypothesis regarding vascular acoustic emission accompanying arterial injury induced by balloon angioplasty.

Stress-induced structural damage is often accompanied by sound release. This behavior is known as acoustic emission (AE). We hypothesize that vascular injury such as that produced by balloon angioplasty is associated with AE. Postmortem human peripheral arterial specimens were randomly partitioned into test (n = 10) and control segments (n = 10). Test segments were inserted into a pressurization circuit and subjected to two consecutive hydrostatic pressurizations. Amplitude, frequency, and energy content of the AE signals released during pressurization were quantified. Test and matched control segments subsequently underwent identical histological processing. Pressure-induced tissue trauma was estimated via computerized histomorphometric analysis of the resulting slides (n = 100). Vascular acoustic emission (VAE) signals exhibited an amplitude range of +/- 5.0 mu bars and were observed to occur during periods of increasing intraluminal pressure. The VAE signal power within the monitored bandwidth was concentrated below 350 Hz. More than 25 times as much VAE energy was released during the first pressurization as during the second: 1,855 +/- 513.8 mJ vs. 73 +/- 44.9 mJ (mean +/- SEM, p < 0.006). Estimates of circumferential intimal wall stress at AE onset averaged 170 kPa, slightly below reported values of arterial tissue rupture strength. Histomorphometric estimates of tissue trauma was greater for the test than their matched control segments (p < 0.0001). These preliminary data suggest that detectable acoustic energy is released by vascular tissue subjected to therapeutic stress levels. Histological analysis suggest that the underlying source of sound energy may be related to tissue trauma, independent of histological preparation artifacts. From this preliminary work, we conclude that VAE may be a fundamental property accompanying vascular tissue trauma, which may have applications to improving balloon angioplasty outcomes.

Cardiac memory: a mechanical and electrical phenomenon.

Alterations in repolarization following prolonged periods of ventricular pacing, termed "cardiac memory," have been well documented. Postpacing changes in cardiac function have also been noted in hypertrophic cardiomyopathy. This study was designed to evaluate the effects of ventricular pacing on postpacing diastolic function and its relationship to repolarization changes. Eight subjects (mean age, 76 yr) with permanent pacemakers were enrolled in this study. Each subject was evaluated at a fixed pacing rate with recording of electrocardiographic and echocardiographic data. Seven sets of measurements were performed in the same sequential pattern: 1) after 1 wk of atrial pacing, 2) within 10 min after initiation of atrioventricular sequential pacing (ventricular pacing), 3) within 10 min after termination of 1 h of ventricular pacing, 4) after 1 wk of ventricular pacing, and 5) within 10 min, at 1 h, and at 24 h after termination of ventricular pacing. All subjects had repolarization changes characteristic of cardiac memory only after 1 wk of ventricular pacing. Changes in repolarization parameters were accompanied by changes in peak left ventricular filling rate (dD/dt/D; P = 0.02) and isovolumic relaxation time (IVRT; P = 0.03) that at 24 h approached baseline values. Correlations existed between changes in the Q-T interval and IVRT (r = 0.53, P = 0.007) and between changes in T wave amplitude and dD/dt/D (r = 0.48, P = 0.018) after long-term ventricular pacing. Thus changes in both repolarization and diastolic function persist after cessation of ventricular pacing and lend support to the concept of electrical and mechanical cardiac memory.

In vitro targeting of antibody-conjugated echogenic liposomes for site-specific ultrasonic image enhancement.

Tissue-specific ultrasonic enhancement can be used for the detection and characterization of atherosclerosis. We have previously demonstrated the generation of inherently echogenic (acoustically reflective) liposomes solely by varying lipid composition and controlling the method of production. In this study, echogenic liposomes composed of phosphatidylcholine (PC), 4-(p-maleimidophenyl) butyryl phosphatidylethanolamine (MPB-PE), phosphatidylglycerol (PG), and cholesterol were conjugated to human gamma globulin to determine the effect of antibody conjugation on liposomal acoustic reflectivity. The liposomes remained highly echogenic following antibody conjugation. Echogenic liposomes were also conjugated to rabbit antihuman fibrinogen to study their ability to target fibrin. Antibody-conjugated liposomes were targeted to fibrin-coated filter paper and slides, thrombi made in vitro, and segments of atheroma in an animal model of atherosclerosis. Liposomes were detected by scanning electron microscopy, radiolabeling, and imaging with intravascular ultrasound. Electron microscopy revealed attachment of antibody-conjugated liposomes to fibrin on slides and to the fibrous plaques of the arterial segments, whereas unconjugated liposomes did not attach. Similarly, conjugated liposomes did not attach to normal arteries, indicating their binding to the arterial segment is directed towards a component of the fibrous plaque. Ultrasound imaging of the thrombi demonstrated surface attachment of the acoustic conjugated liposomes. 125I-Labeled liposomes conjugated to rabbit anti-human were targeted to fibrin-coated paper. Counting specifically bound radioactivity showed that > 84% of applied liposomes remained attached to the fibrin after washing with saline. These results demonstrate the potential of acoustically reflective liposomes for site-specific targeting and acoustic enhancement.

In vitro identification of angioplasty-induced injury by use of vascular acoustic emissions.

BACKGROUND: We have developed a novel method of diagnosing stress-induced vascular injury. This approach uses the sound energy released from atherosclerotic arterial tissue during in vitro balloon angioplasty to characterize type and severity of induced trauma. METHODS AND RESULTS: Thirty-two postmortem human peripheral arterial specimens 1.0 cm long were subjected to in vitro balloon angioplasty with simultaneous acoustic emission monitoring. Specimens were examined before and after angioplasty to ascertain the extent of angioplasty-induced injury. Gross observation was used to identify dissection. A three-dimensional intravascular ultrasound reconstruction technique was used to estimate the luminal surface area of the specimen. Change in luminal surface area (postangioplasty minus preangioplasty) was used to quantify induced injury. The energy content and spectral distribution of the digitally acquired vascular acoustic emission (VAE) signals were computed. Comparisons of angioplasty-induced trauma with VAE signal characteristics were made. Dissection (mural laceration of variable depth) was observed in 15 of 32 specimens. Eleven showed no evidence of induced dissection, and 6 had preexisting intimal disruptions. The energy content of the VAE signals collected from specimens with dissection was greater than that obtained from those in which dissection was absent: 845 +/- 89.4 mJ (mean +/- SEM; n = 15) versus 128 +/- 40.8 mJ (n = 1 l; P < .001). Comparison of induced trauma and VAE signal energy demonstrated a proportional relationship (r = .87, P < .001, n = 32). CONCLUSIONS: VAE signals contain information characterizing type and severity of angioplasty-induced arterial injury. Because vascular injury is related to adverse procedural outcome, development of VAE technology as an adjunct to conventional diagnostic modalities may facilitate optimal balloon angioplasty delivery and postprocedural care.

Intravascular ultrasound. Three-dimensional applications and forward viewing.

This article provides the reader with some idea of the principles and techniques of three-dimensional reconstruction using intravascular imaging data. The article also describes new intravascular ultrasound imaging devices that have the ability to interrogate the arterial wall ahead of the imaging catheter.

Regional vascular mechanical properties by 3-D intravascular ultrasound with finite-element analysis.

A method employing intravascular ultrasound (IVUS) and simultaneous hemodynamic measurements, with resultant finite element analysis (FEA) of accurate three-dimensional IVUS reconstructions (3-DR), was developed to estimate the regional distribution of arterial elasticity. Human peripheral arterial specimens (iliac and femoral, n = 7) were collected postmortem and perfused at three static transmural pressures: 80, 120, and 160 mmHg. At each pressure, IVUS data were collected at 2.0-mm increments through a 20.0-mm segment and used to create an accurate 3-DR. Mechanical properties were determined over normotensive and hypertensive ranges. An FEA and optimization procedure was implemented in which the elemental elastic modulus was scaled to minimize the displacement error between the computer-predicted and actual deformations. The "optimized" elastic modulus (Eopt) represents an estimate of the component element material stiffness. A dimensionless variable (beta), quantifying structural stiffness, was computed. Eopt of nodiseased tissue regions (n = 80) was greater than atherosclerotic regions (n = 88) for both normotensive (Norm) and hypertensive (Hyp) pressurization: Norm, 9.3 +/- 0.98 vs. 3.5 +/- 0.30; Hyp, 11.3 +/- 0.72 vs. 8.5 +/- 0.47, respectively (mean +/- SE x 10(6) dyn/cm2; P < 0.01 vs. nondiseased). No differences in beta between nondiseased and atherosclerotic tissue were noted at Norm pressurization. With Hyp pressurization, beta of atherosclerotic regions were greater than nondiseased regions: 21.5 +/- 2.21 vs. 14.0 +/- 2.11, respectively (P < 0.03). This method provides a means to identify regional in vivo variations in mechanical properties of arterial tissue.