Rodney Greene

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.

Up-regulation of vascular endothelial growth factor mrna and angiogenesis after transmyocardial laser revascularization

BACKGROUND Angiogenesis has been proposed as a potential mechanism whereby transmyocardial laser revascularization (TMLR) has provided clinical relief of angina. Experimental work has found histologic evidence supporting this, as well as an improved response when angiogenic growth factors have been added to TMLR. The purpose of this study was to demonstrate that the molecular response to TMLR was an increase in the production of endogenous vascular endothelial growth factor to promote angiogenesis. METHODS Ameroid constrictors were placed on the proximal circumflex artery in 12 domestic pigs. After a chronic ischemic zone was established the animals were randomly divided into two groups. In the TMLR group the ischemic zone was treated with carbon dioxide laser. In the control group the ischemic zone was untreated. Six weeks later the animals were sacrificed, and sections from the ischemic zone and the nonischemic zone were submitted for immunohistochemical, histologic, and molecular analysis. Messenger RNA was obtained from northern blot analysis after being probed with vascular endothelial growth factor. RESULTS There was a twofold increase in the vascular endothelial growth factor messenger RNA in the ischemic zone of the TMLR group compared with the control group. Additionally, there was a threefold increase in the number of new blood vessels in the ischemic zone of the TMLR group compared with the control group. CONCLUSIONS Transmyocardial laser revascularization promotes angiogenesis by upregulation of vascular endothelial growth factor. The resulting angiogenesis could be the principle mechanism for the clinical efficacy of TMLR.

Optimal pleurodesis: a comparison study.

With the resurgence of thoracoscopy, there is renewed interest in less invasive methods of pleurodesis. We wished to compare in an animal model a variety of methods suggested in reports. The purpose of the study was to rank the effectiveness of each procedure against the standard mechanical pleurodesis. Twenty-five mongrel dogs (weight, 25 to 35 kg) underwent bilateral thoracotomy. Each animal was randomly assigned to receive two of the following methods of pleurodesis: tetracycline, talc, mechanical abrasion, neodymium: yttrium-aluminum garnet (Nd:YAG) laser (Hereus Inc, E. Rutherford, NJ) photocoagulation, and argon beam coagulator (ABC) (Beacon Lab, Bloomfield, CO) electrocoagulation of the parietal pleura. At evaluation at 30 +/- 2 days, the efficacy of pleurodesis was graded on a scale of 0 to 4, with 0 representing a complete absence of pleural symphysis and 4 the adhesion of more than one lobe to both the chest wall and mediastinum. Mean grade and standard deviation of each method were: talc, 3.0 +/- 0.67; mechanical, 3.0 +/- 0.82; tetracycline, 2.3 +/- 1.4; ABC, 1.5 +/- 0.97; and Nd:YAG laser, 0.7 +/- 0.95. Both the talc and mechanical methods were superior to either the Nd:YAG laser or the ABC (p < 0.01). In this study, neither the Nd:YAG laser nor the ABC proved efficacious in producing pleurodesis. Talc poudrage is the only method of pleural symphysis comparable with mechanical abrasion.

Myocardial functional recovery after fibroblast growth factor 2 gene therapy as assessed by echocardiography and magnetic resonance imaging

BACKGROUND Although it has been shown that gene therapy is capable of inducing neovascularization in ischemic myocardium, the functional significance of such therapeutic angiogenesis remains less certain. The purpose of this study was to investigate whether an experimental link could be made between the ability of a novel fibroblast growth factor 2 (FGF2) gene formulation to promote neovascularization, and its ability to restore myocardial function. METHODS Fibroblast growth factor 2 gene was delivered by means of an adenovirus vector formulated in a collagen-based matrix to provide localized and sustained gene activity. Using a model of chronic myocardial ischemia, animals were randomized to either treatment of the ischemic area by injections of adenovirus vector-FGF2 or no treatment. Left ventricular function was assessed by rest and dobutamine stress echocardiography as well as contrast-enhanced and cine magnetic resonance imaging scans. Studies were repeated 6 weeks after treatment. Arteriogenesis was assessed by quantifying the total arteriolar wall area present in treated areas, using anti-alpha-actin immunohistochemistry and subsequent morphometric analyses. RESULTS Echocardiographic results demonstrated a significant restoration of myocardial function in FGF2 gene-treated areas as measured by myocardial wall thickening (0.38 +/- 0.08 cm pretreatment versus 0.76 +/- 0.09 cm posttreatment; p < 0.05). This was demonstrated by comparing the ischemic zones of FGF2 gene-treated versus control-treated animals, as well as by comparing ischemic with nonischemic zones in individual animals This functional improvement was confirmed by cine magnetic resonance imaging, in which 68% (147 of 216) of the treated segments showed improvement in wall motion and there was no change in the untreated segments. Fibroblast growth factor 2 gene treatment also enhanced arteriogenesis within the ischemic zone, as FGF2 gene-treated animals showed a 340% increase in the total arteriolar wall area present versus control-treated animals. CONCLUSIONS The function of ischemic myocardium can be restored by a novel FGF2 gene delivery method using a gene-activated matrix. The increased arteriogenesis as a result of FGF2 gene therapy leads to restoration of this myocardial function.

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.

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.

Computation of vascular flow dynamics from intravascular ultrasound images

Analysis of three-dimensional velocity profiles and wall shear stress distribution in a segment of an artery reconstructed from in vivo imaging data are presented in this study. Cross-sectional images of a segment of the abdominal aorta in dogs were obtained using intravascular ultrasound (IVUS) imaging employing a constant pull back technique. Simultaneous measurement of pressures distal and proximal to the vessel segment along with gated pulsed Doppler velocity measurements were also obtained. The three-dimensional geometry of the vascular segment was reconstructed from the IVUS images during peak forward flow phase, and a computational mesh was constructed from the data. A quasi-steady analysis of incompressible Newtonian fluid was performed with a finite difference general purpose computational analysis program FLOW3D. The velocity at the inlet and pressure at the outlet measured at the corresponding time (time referenced to ECG) were used to specify the boundary conditions for the computational flow model. The computed results compared favorably with previously reported results. The purpose of the present study was to analyze the hemodynamics in vascular segments from morphologically realistic three-dimensional reconstructions. The method can be potentially employed in analyzing the hemodynamics in the region of atherosclerotic plaques at various stages of development and the reactivity of the vessel in response to pharmacological and mechanical interventions.

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.

Validation of transesophageal echocardiography to determine physiologic coronary flow.

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:

Functional consequences and intracoronary localization of alpha-adrenergic stimulation of the canine coronary circulation

Although alpha-adrenergic stimulation can increase coronary vascular resistance, it remains unknown whether the vasoconstriction can override intrinsic coronary regulatory influences to produce ischemia. Methoxamine, 2 to 4 mg, was infused into the circumflex coronary artery of 23 chloralose-anesthetized open chest dogs, and resulted in a 68% increase in coronary vascular resistance. The functional consequence of this increased coronary vascular resistance was assessed by gated radionuclide ventriculography and ST-T wave changes on the electrocardiogram. In six dogs (Group I), aortic pressure changed trivially (less than 5 mm Hg) to allow distinction between direct effects of the flow reduction and indirect effects of increased aortic pressure. In this group, coronary blood flow decreased 33% from a control value of 44 +/- 10 ml/min (p less than 0.001) and left ventricular ejection fraction decreased from 0.54 +/- 0.12 to 0.46 +/- 0.10 (p less than 0.025). In eight dogs (Group II) in which aortic pressure increased by more than 5 mm Hg, left ventricular ejection fraction decreased from 0.46 +/- 0.07 to 0.39 +/- 0.09 (p less than 0.002). Pressure gradients were measured between the aorta and a distal coronary artery branch to calculate small and large vessel resistances separately in four other dogs (Group III). The resistance of small coronary arteries accounted for 92% of the total increase in coronary vascular resistance produced by methoxamine. In five other dogs (Group IV), intracoronary methoxamine, 2 mg, produced ST-T wave changes suggestive of ischemia as it increased coronary vascular resistance by 33%.(ABSTRACT TRUNCATED AT 250 WORDS)