Gene transfer with echo-enhanced contrast agents: comparison between Albunex, Optison, and Levovist in mice--initial results

PURPOSE: To determine if commercially available echo-enhanced microbubble contrast agents could be used to increase gene transfection efficiency by means of relatively low-intensity ultrasound-mediated microbubble destruction in skeletal muscles. MATERIALS AND METHODS: Three types of ultrasound microbubble contrast agents (0.01 mL of albumin [Albunex] and human albumin [Optison] and 10 mg/mL of SH U 508A [Levovist]) were each separately mixed with the reporter plasmid DNA (25 microg) encoding green fluorescent protein (GFP) prior to intramuscular injection into the quadriceps muscle of a mouse thigh bilaterally (seven mice per contrast agent). One of the muscle sites that was injected with plasmid DNA was irradiated with low-intensity therapeutic ultrasound (1 MHz) at an intensity of 2.0 W/cm2 for 2 minutes. Mice were sacrificed 7 days after ultrasound treatment for gene expression assay. The number of GFP-expressing muscle fibers was counted. Statistical significance was determined with a two-tailed Student t test. P <.05 was considered to indicate statistically significant difference. RESULTS: Muscle tissue exposed to ultrasound with air-filled Albunex or Levovist microbubbles revealed no difference in the number of GFP-expressing muscle fibers compared with the control non-ultrasound-exposed muscle. Albumin-coated octafluoropropane gas-filled Optison microbubbles showed a 10-fold increase in the number of GFP-expressing fibers (P <.05). CONCLUSION: Low-intensity ultrasound with echo-enhanced Optison induced efficient gene transfer unlike that with Albunex or Levovist.     

Enhancement of gas‐filled microbubble R2* by iron oxide nanoparticles for MRI

Gas‐filled microbubbles have the potential to become a unique intravascular MR contrast agent due to their magnetic susceptibility effect, biocompatibility, and localized manipulation via ultrasound cavitation. However, microbubble susceptibility effect is relatively weak when compared with other intravascular MR susceptibility contrast agents. In this study, enhancement of microbubble susceptibility effect by entrapping monocrystalline iron oxide nanoparticles (MIONs) into polymeric microbubbles was investigated at 7 T in vitro. Apparent T₂ enhancement (ΔR₂*) induced by microbubbles was measured to be 79.2 ± 17.5 sec^?1 and 301.2 ± 16.8 sec^?1 for MION‐free and MION‐entrapped polymeric microbubbles at 5% volume fraction, respectively. ΔR₂* and apparent transverse relaxivities (r₂*) for MION‐entrapped polymeric microbubbles and MION‐entrapped solid microspheres (without gas core) were also compared, showing the synergistic effect of the gas core with MIONs. This is the first experimental demonstration of microbubble susceptibility enhancement for MRI application. This study indicates that gas‐filled polymeric microbubble susceptibility effect can be substantially increased by incorporating iron oxide nanoparticles into microbubble shells. With such an approach, microbubbles can potentially be visualized with higher sensitivity and lower concentrations by MRI. Magn Reson Med, 2010. © 2009 Wiley‐Liss, Inc.     

Air microbubbles as MR susceptibility contrast agent at 1.5 Tesla.

Air microbubbles have been investigated recently at high magnetic field strength (2 Tesla or greater) as potential MR susceptibility contrast agents. We used a phantom to measure their susceptibility at 1.5 T to clarify their usefulness for this purpose. The phantom, filled with fresh Levovist suspension at 4 different doses (67 to 125 mg/mL), was continuously scanned with a gradient-echo technique at a temporal resolution of 10 s. The transverse relaxation increase (R2*) by microbubbles demonstrated a time course of exponential decay at each dose (time-constant, 39 to 57 s). The dependency of R2* on microbubble volume fraction was linear, with a slope of 89 s-1 per percentage microbubble volume fraction. Our study represents the first step towards applying microbubbles as susceptibility contrast agents at 1.5 T.     

Prolonging the ultrasound signal enhancement from thrombi using targeted microbubbles based on sulfur-hexafluoride-filled gas

RATIONALE AND OBJECTIVES: The objective of this study is to develop and characterize new microbubbles based on lipids and sulfur hexafluoride (SF6) for targeting thrombi as an improved ultrasound contrast agent. MATERIALS AND METHODS: Bioconjugate ligands were inserted into the lipid-coated membranes of SF6 gas microbubbles, and their physicochemical properties were determined. Diagnostic efficacies of SF6-filled microbubbles and the contrast agent SonoVue (Bracco Imaging, Geneve, Switzerland) were compared in dogs. RESULTS: Suspensions of lyophilized powder were reconstituted by injecting saline containing 3.1 x 10(8) SF6 microbubbles/mL with a mean diameter of 4.4 microm. More than 90% of microbubbles had diameters between 1 and 10 microm. After reconstitution, echogenicity and microbubble characteristics were unchanged for 8 hours. Targeted microbubbles increased the echogenicity of thrombi significantly and provided a longer period of optimal signal enhancement compared with nontargeted microbubbles. CONCLUSIONS: Our thrombus-targeting microbubble contrast agent shows high echogenicity and stability and thereby enhances the visualization of intravascular thrombi and prolongs the duration of the diagnostic window.     

Quantitative and qualitative analysis of in vivo Doppler signal enhancement of FS-069.

RATIONALE AND OBJECTIVES: In vivo lifetime of ultrasound (US) contrast agents is still limited and thus a cause for ongoing investigations of new substances. The purpose of this study was to determine the time intensity changes of the Doppler signals obtained within the femoral vein after two different doses of a new microsphere-based ultrasound contrast agent. METHODS: Twenty-four healthy male volunteers (mean age, 29; average weight, 76 kg) were included in this study. All underwent a triplex Doppler US examination after an intravenous bolus injection of 0.3 mL and 1.0 mL Optison. To examine the signal enhancement characteristics of this contrast agent the audio signal of the pulsed-wave spectral Doppler US was measured quantitatively using an audio analyzer, whereas velocity-encoded color Doppler examinations were scored qualitatively (score 0-3). RESULTS: The mean maximal enhancement of the audio signal at a dose of 1.0 mL FS-069 was significantly higher than with a bolus of 0.3 mL FS-069 (29 +/- 2 dB vs. 26 dB +/- 2 dB, P < 0.001). The time-intensity curves after each bolus injection yielded an early peak (one minute after the injection) followed by constantly decreasing signal intensities. The scoring of the velocity-encoded color Doppler US revealed an optimal enhancement (score 2) for 3 minutes and 20 seconds (0.3 mL Optison) and for 6 minutes (1.0 mL Optison), respectively. CONCLUSIONS: This study showed the capability of triplex Doppler ultrasound signal enhancement after Optison. 1.0 mL Optison proved to be the more appropriate dose for an optimal signal enhancement than 0.3 mL Optison.     

脂质囊泡超声造影剂的质量评价及猪肝彩色多普勒显影研究

目的 对自制的脂质囊泡超声造影剂进行质量评价,研究不同剂量与正常猪肝彩色多普勒显影效果间的相互关系.方法 显微镜观察脂质囊泡的形态、激光粒度测定仪测定粒径及粒度分布,用血细胞分析仪统计不同放置时间条件下和配制浓度粒径在2-8μm范围内的囊泡浓度以及光照和温度对囊泡浓度和平均粒径的影响.进行造影剂剂量与正常猪肝实质彩色多...     

Mechanism of hepatic parenchyma-specific contrast of microbubble-based contrast agent for ultrasonography: microscopic studies in rat liver

OBJECTIVE: The objective of this study was to elucidate the mechanism of hepatic parenchyma-specific contrast of Sonazoid (microbubble contrast agent) using microscopic techniques. MATERIALS AND METHODS: Sonazoid was intravenously injected into rats to investigate the microbubble dynamics and distribution within hepatic microcirculation in exteriorized liver using intravital microscopy and to observe dose dependency of ultrasound hepatic contrast effect. In vitro and in vivo uptake of microbubbles by Kupffer cells was examined using confocal laser scanning microscopy. RESULTS: Intravital observation demonstrated freely flowing microbubbles in the sinusoid and some microbubbles co-localized with Kupffer cells. The microbubbles internalized in Kupffer cells were identified with reflected light by confocal laser scanning microscopy. The percentage of Kupffer cells taking up microbubbles was about 1% at clinical dose at which the homogeneous hepatic contrast was observed. CONCLUSIONS: The hepatic parenchyma-specific contrast by Sonazoid is due to distribution of the microbubbles in Kupffer cells.     

MR perfusion imaging using encapsulated laser-polarized 3He.

In this work, the use of a new carrier agent for intravascular laser-polarized 3He imaging is reported. Lipid-based helium microbubbles were investigated. Their average diameter of 3 microm, which is smaller than that of the capillaries, makes it possible to conduct in vivo studies. The NMR relaxation parameters T1, T2, and T2* of a microbubble suspension were measured as 90 s, 300 ms, and 4.5 ms, respectively, and in vivo images of encapsulated 3He with signal-to-noise ratios (SNRs) larger than 30 were acquired. Dynamic cardiac images and vascular images of encapsulated 3He were obtained in rats using intravenous injections of microbubble suspensions. Excellent preservation of 3He polarization through the lung capillaries and heart cavities was observed. The first images of 3He microbubble distributions in the lungs were obtained. Additionally, the potential of this technique for lung perfusion assessment was validated through an experimental embolism model with the visualization of perfusion defects.     

Microbubble contrast agent with focused ultrasound to create brain lesions at low power levels: MR imaging and histologic study in rabbits

PURPOSE: To evaluate magnetic resonance (MR) imaging-based thermometry for predicting the onset and spatial extent of lesions produced by focused ultrasound combined with a microbubble contrast agent (Optison; GE Healthcare, Milwaukee, Wis) and to compare the resulting induced temperature increase and threshold for damage with those in studies performed without the agent. MATERIALS AND METHODS: The experiments were approved by the animal care committee. Fifty-three locations in the brains of 15 rabbits were sonicated with various exposure parameters by using a 1.5-MHz focused ultrasound transducer. MR imaging was used to map the temperature rise and, along with light microscopy, to examine the lesions. Diameters of isotherms created from thermometry were compared with the resulting lesions by using Bland-Altman analysis and linear regression. The minimum acoustic power necessary for lesion creation was determined, and the apparent temperature threshold for damage was calculated with probit analysis. These thresholds were compared with prior work performed without the contrast agent. The heating induced with the microbubbles was compared with that in sonications performed without them by using a t test. RESULTS: The MR imaging-mapped temperature distributions matched the shape of the lesions. The diameters of isotherms correlated well with diameters measured at contrast material-enhanced MR imaging (mean difference between measurements, 0.0 mm +/- 0.5; R = 0.93). The temperature increase with microbubbles was statistically larger (P < .01) than for sonications performed without microbubbles. In some locations (mostly continuous wave exposures), damage was observed along the ultrasound beam path. The time-averaged acoustic power damage threshold was reduced by 91% for 10-second exposures when compared with earlier studies performed without microbubbles. The probability of producing lesions was 50% at a temperature increase of 5.9 degrees C, 5.5 degrees C lower than was observed earlier without the agent. CONCLUSION: MR imaging-based temperature measurements appeared to correlate with focused ultrasound-induced lesions in the brain when microbubbles were present, even though the temperature appeared to be below the threshold for thermal damage.     

Microbubble-augmented ultrasound declotting of thrombosed arteriovenous dialysis grafts in dogs

PURPOSE: Transcutaneous low-frequency ultrasound (LFUS) can effectively lyse clots in the presence of microbubbles. This study was designed to test the commercially available human albumin microspheres injectable suspension octafluoropropane formulation, Optison, to establish efficacy and assess US parameters of intensity and wave modes in a canine model of a thrombosed arteriovenous (dialysis) graft. MATERIALS AND METHODS: Arteriovenous grafts in five dogs were cannulated, temporarily ligated, and thrombosed. Different declotting techniques were randomized to treat nine groups. Control groups involved direct saline (4.5 mL) clot injection in 0.5-1.0-mL increments. One group underwent peripheral intravenous microbubble injection (13.5 mL). Six groups underwent direct incremental clot injection of 4.5 mL of microspheres with LFUS for 30 minutes in 3-5-minute increments with use of various intensity settings in continuous-wave and pulsed-wave (PW) modes. At each increment, angiography was used to grade flow, declotting, and overall success. RESULTS: One hundred four procedures showed success in all 24 high-intensity PW modes (1.2-2.0 W/cm(2)); only one of 20 control experiments was successful (P <.0001). Medium-intensity modes yielded intermediate success rates. Lowest-intensity direct-injection groups and intravenous and control groups ranked lower. Results at 30 minutes were better than at 15 minutes (P <.0001). CONCLUSIONS: LFUS with direct injection of microbubbles is effective in lysing moderate-sized clots and recanalizing thrombosed arteriovenous grafts. It best succeeds at the higher range of intensity settings tested in PW mode. Further development is justified.     

Effects of water exchange on the measurement of myocardial perfusion using paramagnetic contrast agents.

To investigate the effects of water exchange on quantification of perfusion, data were acquired in isolated hearts (n = 11) and used to develop a model of exchange. Myocardial T1 was measured 3 times/sec during step changes in concentration of intravascular (polylysine-gadolinium-diethylene-triamine-pentaacetic acid) and extracellular (gadoteridol) agents. For the intravascular agent, the change in 1/T1 (deltaR1) was lower than predicted by fast exchange (2.7+/-0.5 vs. 7.8 sec(-1), respectively), and suggested an intra-extravascular exchange rate of 3 Hz. For the extracellular agent, contrast kinetics were similar to those of similarly sized molecules (wash-in time constant 38+/-5 sec), and the data suggested fast interstitial-cellular exchange. Modeling showed that perfusion is underestimated for both agents if exchange is ignored, although the relationships of measured to actual perfusion were monotonic. We conclude that myocardial water exchange strongly affects first-pass enhancement but that ignoring the effects of exchange may still provide reasonable estimates of regional perfusion differences.     

Comparison of iron oxide particles (AMI 227) with a gadolinium complex (Gd-DOTA) in dynamic susceptibility contrast MR imagings (FLASH and EPI) for both phantom and rat brain at 1.5 Tesla

The goal of the study was to compare, in phantom and normally perfused rat brain tissue, a superparamagnetic iron oxide particle-based contrast agent (AMI 227) with a low-molecular-weight gadolinium chelate, gadolinium tetraazacyclododecanetetraacetic acid (Gd-DOTA), in two susceptibility contrast magnetic resonance imaging (MRI) modes [fast low-angle shot sequence (FLASH) and echoplanar imaging (EPI)]. A phantom consisting of dilution series of both contrast agents was manufactured. Dilutions were obtained with isotonic serum from the available agent solutions (0.5 mmol Gd/mL Gd-DOTA; 350 mumol Fe/mL AMI 227). Eighteen rats were studied. Contrast agent (0.1 mL) was bolus injected in each rat, and dynamic MRI was performed (first pass of the contrast agent) in rat brain. The doses of AMI 227 injected were extrapolated from the phantom experiment: 0.2 mmol/kg body weight of Gd-DOTA and 7, 14, and 28 mumol Fe/kg body weight of AMI 227 were injected. For both sequences, signal-to-noise ratios (S/N) were measured on each tube of the phantom and on rat brain from each image of the dynamic imaging. S/N was plotted versus contrast dilution (phantom) and versus time (rats). In the FLASH sequence, a well-shaped curve (S/N decrease, S/N peak decrease, S/N increase) of the first pass of the contrast agent was demonstrated for Gd-DOTA and for AMI 227 (7 mumol Fe/kg body weight). In the EPI sequence, a well-shaped curve was demonstrated for Gd-DOTA, and a plateau effect was noted for both concentrations of AMI 227. With the FLASH technique, dynamic susceptibility contrast imaging of rat brain can be performed with very low concentrations of AMI 227 compared with the Gd-DOTA concentration (0.2 mmol Gd/kg body weight) used in clinical practice. This could be of interest in perfusion imaging, because it may allow for first-pass susceptibility imaging after administration of a small volume in a narrow bolus.     

Comparison of liver perfusion parameters studied with conventional extravascular and experimental intravascular CT contrast agents

RATIONALE AND OBJECTIVES: To compare liver perfusion parameters obtained by using an extravascular contrast agent and a blood-pool agent. MATERIALS AND METHODS: Fifteen rabbits were imaged with a continuous 40-second single-slice computed tomography acquisition after a bolus injection of contrast agent (physiologic bolus duration 4-5 seconds, extravascular iohexol, n = 7; experimental nanoparticulated blood-pool agent WIN8883, n = 8). Time-density curves were generated for the aorta, portal vein, and liver. From the curves, arterial, portal, and total blood flows and hepatic perfusion index (HPI, arterial-to-total perfusion ratio) were determined by using two commonly applied fundamentally different analyzing methods: the single-compartment model and the peak gradient (PG) method. Also, the gamma variate fitting method was used. RESULTS: By using the single-compartment model, the obtained HPI and total blood flow were 0.14 +/- 0.04 and 2.29 +/- 0.40 (mL/min/mL(tissue)) for WIN8883, and 0.15 +/- 0.06 (P = .54) and 4.60 +/- 1.14 (mL/min/mL(tissue)) (P = .0002) for iohexol, respectively. With the PG, HPI and total blood flow were 0.15 +/- 0.08 and 1.27 +/- 0.24 (mL/min/mL(tissue)) for WIN8883, and 0.20 +/- 0.06 (P = .12) and 2.11 +/- 0.25 (mL/min/mL(tissue)) (P = .00002) for iohexol, respectively. With the blood pool agent, similar contrast enhancement to the conventional agent was achieved with about 36% reduced dosage of iodine per body weight (mg I/kg). CONCLUSIONS: HPI was found to be quite insensitive to different contrast agent types and analyzing methods. However, the arterial, portal and total liver blood flow values strongly depend on contrast agent type and modeling method.     

Contrast agent-enhanced, free-breathing, three-dimensional coronary magnetic resonance angiography.

For free-breathing, high-resolution, three-dimensional coronary magnetic resonance angiography (MRA), the use of intravascular contrast agents may be helpful for contrast enhancement between coronary blood and myocardium. In six patients, 0.1 mmol/kg of the intravascular contrast agent MS-325/AngioMARK was given intravenously followed by double-oblique, free-breathing, three-dimensional inversion-recovery coronary MRA with real-time navigator gating and motion correction. Contrast-enhanced, three-dimensional coronary MRA images were compared with images obtained with a T2 prepulse (T2Prep) without exogenous contrast. The contrast-enhanced images demonstrated a 69% improvement in the contrast-to-noise ratio (6.6 +/- 1.1 vs. 11.1 +/- 2.5; P < 0.01) compared with the T2Prep approach. By using the intravascular agent, extensive portions (> 80 mm) of the native left and right coronary system could be displayed consistently with sub-millimeter in-plane resolution. The intravascular contrast agent, MS-325/AngioMARK, leads to a considerable enhancement of the blood/muscle contrast for coronary MRA compared with T2Prep techniques. The clinical value of the agent remains to be defined in a larger patient series. J. Magn. Reson. Imaging 1999;10:790-799.     

1-molar gadobutrol as a contrast agent for computed tomography: results from a comparative porcine study

RATIONALE AND OBJECTIVES: To evaluate prospectively the efficacy of gadobutrol as contrast agent for computed tomography (CT) compared with iodinated contrast media in a porcine animal model. METHODS: In 8 domestic pigs (35 +/- 4 kg body weight [BW]), continuous spiral CTs of the chest and abdomen were performed using either 2 mmol/kg BW Gadovist 1.0 (1 mol/L gadobutrol) intravenously or Ultravist (300 mg I/mL iopromide) (slice 5 mm, table feed 7.5 mm, reconstruction increment 5 mm). One week later, the same animals were examined using the same protocol with the other contrast agent. In 2 additional animals, serial CTs were performed at the same level using gadobutrol or iopromide on day 1 and the alternate agent on day 8 inches order to determine contrast media kinetics, peak enhancement, and time enhancement-product in important vascular regions and parenchymal organs (abdominal aorta, inferior vena cava, liver, and renal parenchyma). Peak enhancement (net increase compared with nonenhanced baseline values) was measured in Hounsfield units (HU) in defined regions of interest. RESULTS: In vivo, the mean peak enhancement 5, 15, 30, and 120 seconds in the abdominal aorta after injection of 2 mL/kg BW gadobutrol and iopromide was 200 +/- 11, 224 +/- 10, 261 +/- 13, and 95 +/- 9 HU versus 232 +/- 10, 298 +/- 10, 152 +/- 11, and 123 +/- 10 HU, respectively. Differences in enhancement of vascular structures was statistically significant (P < 0.05) in carotid arteries (235 +/- 20 HU for gadobutrol and 264 +/- 19 HU for iopromide) and the aortic arch (261 +/- 14 HU for gadobutrol and 279 HU +/- 13 HU for iopromide). No statistical significance was seen in all other measured vascular structures and parenchymal organs. CONCLUSION: Contrast-enhanced CT with 1 mol/L gadobutrol in a dose of 2 mmol/kg BW resulted in an excellent vascular and parenchymal enhancement in most vascular regions and parenchymal organs similar to an equivalent volume of 300 mg/mL iodinated contrast media.     

Evidence for spleen-specific uptake of a microbubble contrast agent: a quantitative study in healthy volunteers

PURPOSE: To evaluate the pharmacokinetics of the microbubble contrast agent BR1. MATERIALS AND METHODS: Twenty healthy volunteers were injected via arm vein with a 1.2-mL bolus of BR1. Ultrasonographic images of liver and right kidney and of spleen and left kidney were obtained intermittently for 5 minutes with low-mechanical-index software (to minimize microbubble destruction) that shows stationary microbubbles in green. Percentage total uptake was calculated as the number of green pixels in the region of interest for each organ over time, divided by the total pixels. Relative uptake, the ratio of total uptake in liver to that in right kidney and of total uptake in spleen to that in left kidney, and differential uptake, the difference in total uptake between liver and right kidney and between spleen and left kidney, were calculated. Total uptake for each organ was plotted against time, and the gradient of a best-fit straight line was calculated. Wilcoxon signed rank test was used to compare mean uptake values in each subject. Mann-Whitney U test was used for comparisons in sex and age. RESULTS: Total uptake declined over 5 minutes in left and right kidney and in liver (from 88% +/- 10% [1 minute] to 67% +/- 14% [5 minutes]), but not in spleen (range, 90%-99%). Mean relative uptake +/- 1 SD for spleen increased from 2.3 +/- 0.7 (1 minute) to 3.7 +/- 2.3 (5 minutes) (P =.005) but for liver was constant: 2.1 +/- 0.9 (1 minute) and 2.3 +/- 0.4 (5 minutes) (P =.06). Mean differential uptake +/- 1 SD for spleen increased from 51.3% +/- 14.9% (1 minute) to 65.0% +/- 9.1% (5 minutes) (P =.002). Significant difference was seen over time in total uptake gradients between spleen and left kidney (P =.014) but not between liver and right kidney or right and left kidney. No difference was seen between men and women or with age. CONCLUSION: BR1 produces spleen-specific enhancement that is longer (5 minutes) than the blood pool phase.     

Difference of optimal dose of contrast agent between gray-scale and power Doppler imaging in assessing graded coronary stenosis by myocardial contrast echocardiography

RATIONALE AND OBJECTIVES: In myocardial contrast echocardiography (MCE), power Doppler imaging is more sensitive to contrast agent (microbubble) than gray-scale B-mode imaging; however, no data exist regarding the optimal contrast dose in power Doppler imaging. This study examined the optimal dose of contrast agent for power Doppler in assessing coronary stenosis. METHODS: Three grades of coronary stenosis were produced in 6 open-chest dogs. MCE was performed with gray-scale and power Doppler during continuous infusion of 0.2 mL/min FS-069. Thereafter, MCE was repeated with power Doppler during continuous infusion of 0.1 mL/min FS-069. RESULTS: Although the videointensity in the stenosed bed with power Doppler (214 +/- 14) was greater than gray scale (35 +/- 17) during 0.2 mL/min FS-069 infusion (P < 0.0001), power Doppler failed to identify milder coronary stenoses because videointensity in stenosed bed was quickly saturated with contrast agent. The videointensity in the stenosed bed with power Doppler (127 +/- 49) during 0.1 mL/min FS-069 infusion was greater than gray scale (35 +/- 17) during 0.2 mL/min FS-069 infusion (P < 0.0001), and all levels of stenosis were identified with power Doppler, even though the dose of contrast agent was half of that of gray scale imaging. The correlation between videointensity and myocardial blood flow was better in the case of power Doppler at 0.1 mL/min FS-069 infusion (r = 0.77, P < 0.0001) than in the case of gray scale imaging at 0.2 mL/min FS-069 infusion (r = 0.66, P < 0.01). CONCLUSIONS: These data support the need for a lower dose of contrast agent for power Doppler than for gray scale to detect milder coronary stenosis and avoid saturation of imaging fields.     

Gene transfer with microbubble ultrasound and plasmid DNA into skeletal muscle of mice: comparison between commercially available microbubble contrast agents

PURPOSE: To compare three commercial microbubble contrast agents (Optison, SonoVue, and Levovist) for their effect on gene delivery in skeletal muscle in conjunction with the use of therapeutic ultrasound. MATERIALS AND METHODS: The study was approved by the Animal Care and Use Committee. Plasmid DNA (10 microg) encoding green fluorescent protein (GFP) was mixed with microbubbles (or saline control) and injected into the tibialis anterior muscle of mice with and without adjunct ultrasound (1 MHz, 2 W/cm2, 30 seconds, 20% duty cycle). The efficiencies of GFP transgene expression were determined with four experimental conditions: (a) plasmid and saline as control (six mice), (b) plasmid and Optison (six mice), (c) plasmid and SonoVue (four mice), and (d) plasmid and Levovist (air based, four mice). The right legs were exposed to ultrasound, while the left legs were unexposed. Transfection efficiency was assessed by counting the number of GFP-positive fibers. Tissue damage was assessed by measuring the maximal-damage area on serial sections. RESULTS: When ultrasound was applied, both SonoVue and Optison significantly improved (P < .05) gene transfection efficiency. Optison was also effective (P < .05) even when no ultrasound was applied, which is consistent with previous studies. Levovist without ultrasound decreased the level of transfection (P < .05), with increased tissue damage. CONCLUSION: Both non-air-based agents show promise in gene delivery in skeletal muscle with undetectable tissue damage. Enhanced gene transfer with additional ultrasound was achieved only with SonoVue.     

Molecular imaging with contrast ultrasound and targeted microbubbles

There is growing interest in the development of methods for imaging cellular and molecular mediators of cardiovascular diseases. Techniques for imaging molecular and cellular alterations have been explored for essentially all noninvasive cardiac imaging modalities. Molecular imaging with contrast-enhanced ultrasound relies on the detection of novel site-targeted microbubble contrast agents. These microbubbles are retained within regions of a specific disease process, thereby allowing phenotypic characterization of tissue. As microbubbles are pure intravascular tracers, the disease processes assessed must be characterized by antigens that are expressed within the vascular compartment. Accordingly, the pathologic states that have been targeted include inflammation, neoplasms, angiogenesis, and thrombus formation, all of which are mediated in part by molecular events within the vascular space. This review describes (1) different strategies that have been used to target microbubbles to regions of disease, (2) the unique challenges for imaging targeted ultrasound contrast agents, and (3) some of the early experience imaging molecular events in animal models of disease.