Ultrasound Enhancement Research Articles

04-17 Evaluation of TIPS stenosis with grey-scale ultra-sound contrast enhancement

04-17 Evaluation of TIPS stenosis with grey-scale ultra-sound contrast enhancement

Ultrasound enhancement of cationic lipid-mediated gene transfer to primary tumors following systemic administration.

The impact of a localized application of ultrasound on gene transfer to primary tumors following systemic administration of cationic lipid based transfection complexes was investigated. We have previously shown that systemic administration of DOTMA (N-[(1-(2-3-dioleyloxy) propyl)]-N-N-N-trimethylammonium chloride):cholesterol-based transfection complexes to tumor-bearing mice resulted in expression in the tumor and other tissues, primarily the lungs. Application of ultrasound to the tumor before or after the injection resulted in a significant increase in gene transfer to the tumor with no increase observed in other tissues. The magnitude of increased expression ranged from three- to 270-fold depending upon the DNA dose. The following parameters were optimized for maximal increase: duration of ultrasound application, the time interval between plasmid injection and sonoporation, and plasmid dose. A combination of plasmid quantitation and fluorescence microscopy showed that ultrasound increased tumor uptake of the plasmid and that uptake was limited to the tumor vasculature. Using an IL- 12 expression plasmid, the combination of a single plasmid dose (10 microg) and ultrasound treatment produced significantly higher levels of IL-12 in tumor. This increased expression was sufficient to inhibit tumor growth compared with the control conditions. These data demonstrate the potential application of sonoporation as an effective method for enhancing the expression of systemically administered genes in tumor endothelium for cancer gene therapy.

Ultrasound enhancement of liposome-mediated cell transfection is caused by cavitation effects

Cationic liposomes (CL) are widely used vectors for gene transfer. Recently, ultrasound (US) was reported to enhance liposome-mediated gene transfer to eucaryotic cells in culture. The present study was aimed at studying the effects of 2-MHz pulsed Doppler US on malignant brain tumor cells transfection by cationic liposome/plasmid-DNA complexes (lipoplexes). Cationic liposomes consisting of DOSPA/DOPE were complexed with a plasmid carrying the cDNA encoding green autofluorescent protein (EGFP). Rodent (9L) and canine (J3T) glioma cells were exposed to pulsed US in the presence of EGFP-lipoplexes. A diagnostic transcranial Doppler device (MultiDop L) was used for insonation for 30, 60, and 90 s at 2 MHz/0.5 W/cm 2. To eliminate US reflection and cavitation, a custom-made absorption chamber was designed, where US is applied through a water tank before interacting with the cells and is fully absorbed after passing through the cell layer. Expression of the marker gene EGFP was quantified by FACS analysis and intravital fluorescent microscopy. Cell viability was accessed by Trypan Blue staining. US treatment of tumor cells on microplates for 60 s yielded a significant increase in transfection rates without damaging the cells, but 90-s treatment killed most of the cells. In the absorption chamber, no significant effects of US on transfection were noted. Additional experiments employed US contrast agent (Levovist®, Schering) which was able to significantly increase tumor cell transfection rate by enhancing cavitation effects, and also severely damaged most cells when applied at a concentration of 200 mg/mL. In conclusion, our results support the assumption that US effects on lipoplex transfection rates in brain tumor cells in culture are mediated by cavitation effects.

Characterization of the liver tumors and evaluation of the treatment of HCCS with ultrasound contrast enhancement using wideband gray-scale harmonic imaging

Characterization of the liver tumors and evaluation of the treatment of HCCS with ultrasound contrast enhancement using wideband gray-scale harmonic imaging

Ultrasound enhancement of thrombolytic therapy: observations and mechanisms.

Fibrinolytic therapy is a proven approach for achieving reperfusion of occluded coronary arteries during myocardial infarction, resulting in reduced mortality and preservation of ventricular function. The amount of myocardial muscle loss is proportional to the duration of ischemia. Bleeding complications are not infrequent. Adjuvant therapy by ultrasound might enhance the rate of fibrinolysis and reduce the concentrations of lytic agents required to achieve an equivalent degree of clot lysis. Noninvasive ultrasound at low intensities and high frequencies, parameters that potentially could be applied and tolerated in vivo, have been proven to significantly accelerate the rate of fibrinolysis in both in vitro and in vivo models, in pure fibrin as well as whole blood clots. Such enhancement is not drug-specific. These effects were achieved by nonthermal mechanism. Ultrasound exposure did not cause mechanical fragmentation of the clot, did not alter the size of plasmatic derivates and degradation products. Ultrasound caused increased flow rate through thrombi, probably by cavitation-induced changes in fibrin ultrastructure; disaggregation of uncrosslinked fibrin fibers into smaller fibers has been shown. This resulted in increased transport of the lytic agent into the clot, alteration of binding affinity and increased maximum binding. Presence of echo-contrast agent induced further acceleration of thrombolysis by ultrasound.

Thermal effect of ultrasound enhancer for transdermal insulin delivery

Ultrasound has been known as a good enhancer for transdermal drug delivery. However, major mechanisms in the ultrasound enhancer to improve the transdermal drug delivery were not well understood. In this study a simple experiment was designed to simulate the human skin system with rat back skin. An unfocussed 1-MHz ultrasound transducer and a heating wire loop were used as enhancers. Transmitted amounts of insulin with the ultrasound enhancer were increased ten times more than those without any enhancer. The ultrasound enhancer also improved by 25% the enhancement of insulin transportation compared with that due to thermal effect solely by heating wire loop. It was also observed that rapid temperature increase with ultrasound more efficiently enhanced transdermal transport of insulin. The thermal effect of the ultrasound enhancer is suggested as a most likely mechanism for transdermal insulin delivery.

Diagnostic value of signal-enhanced color Doppler sonography in reactively enlarged lymph nodes

This prospective study was performed to investigate characteristic sonomorphological features and vascularity of reactively enlarged lymph nodes. We examined 40 patients with enlarged lymph nodes of the neck by Doppler sonography before and after administration of an ultrasound signal-enhancing agent. Peak flow velocity, pulsatility- and resistant indices were assessed. Lymph nodes were sonomorphologically classified into three groups: homogeneous parenchyma (1), a centrally located echogenoic line (2) and an echogenoic "hilus reflex" (3). The quantitative and qualitative criteria were compared to histological findings. A total of 15 lymph nodes appeared homogeneous. A centrally located echogenoic line was found in 17 (42.5%) nodes and echogenoic "hilus reflex" in 8 (20%). These sonomorphological patterns correlate to hilus fibrosis in the histological specimen. In 32.5% (13) of the nodes the analysis of vascularity was possible only after application of the galactose-based ultrasound signal enhancer. Quantitative analysis did not result in specific new aspects. Reactive lymph nodes show typical sonomorphological features. Administration of an ultrasound enhancer allows assessment of a characteristic nodal vascualrity.

In vitro studies of a new thrombus-specific ultrasound contrast agent

Ultrasound is used as a primary diagnostic technique for the detection of deep venous thrombosis. The purpose of this study is to describe the development of a new thrombus-specific ultrasound contrast agent: The linear hexapeptide (lysine-glutamine-alanine-glycine-aspartate-valine) was synthesized and coupled to a lipid moiety. The targeted lipid was then incorporated into the lipid blend for the contrast agent Aerosomes (ImaRx, Tucson, AZ, USA). The lipid blend was used to entrap perfluorobutane microbubbles. The microbubbles were sized and studied in vitro for acoustic stability, binding to blood clot, and ultrasound enhancement in vitro of blood clot. The results showed the mean size of the specific ultrasound contrast agent (MRX-408) was about 2.0 μm. The microbubbles appeared as smooth spherical structures. Microscopy showed that the targeted bubbles bound to blood clot whereas control, nontargeted bubbles did not bind to blood clot. In vitro acoustic study showed similar stability of the microbubbles compared with control microbubbles. The targeted microbubbles enhanced blood clot in vitro whereas nontargeted microbubbles did not enhance clot. Thus this promising new thrombus-specific ultrasound contrast agent could potentially improve detection of thrombosis by ultrasound and might be useful for distinguishing between new and old thrombosis. In vivo studies are in progress.

Ultrasound enhancement of rabbit femoral artery thrombolysis

Experiments were conducted to evaluate the potential of low-intensity, externally applied ultrasound to accelerate arterial thrombolysis in an animal model and to characterize potential effects of ultrasound exposure on vessel wall morphology. The femoral arteries of 32 rabbits were exposed, a flowprobe was positioned around the vessel, and a stenosis produced with two circumferential silk sutures to reduce flow by 50%. Thrombosis was achieved by injecting thrombin through the cannulated superficial epigastric branch into a 1-cm segment of femoral artery which was isolated for 20 min. Streptokinase was administered intravenously as a 15 000 U/kg bolus followed by an infusion of 15 000 U/kg per h. Ultrasound (1 MHz, 2 W/cm 2) was delivered to the thrombosed vessel during streptokinase administration in 17 animals, and 15 control animals received sham ultrasound only. Thrombolysis occurred in nine of 17 (53%) animals receiving both streptokinase and ultrasound, and this was significantly greater than the rate in animals receiving streptokinase alone ( 2 15 , 13%; P = 0.025). Ultrasound caused a mean temperature elevation of 4°C in exposed tissues. Light and electron microscopy demonstrated increased platelet accumulation on thrombi in ultrasound-treated vessels compared with controls. Endothelial cell vacuolation was seen by electron microscopy in ultrasound-exposed vessels. The results indicate that externally applied, low-intensity ultrasound can significantly enhance thrombolysis in a rabbit arterial model. Possible adverse effects are minor and include platelet accumulation, temperature elevation and minor endothelial changes. Externally applied ultrasound has potential value as an adjunct to thrombolytic therapy. © 1997 The International Society for Cardiovascular Surgery.

971-54 A Non-invasive Method of Visually Assessing Renal Perfusion Using a Newly Developed Intravenous Ultrasound Contrast

Present methods of quantifying renal artery blood flow (RABF) in renovascular disease require either radionuclide techniques or invasive delivery of radiographic or ultrasound contrast. Perfluoropropane is a gas routinely used for intraocular injections which, when sonicated with dextrose albumin (PESDA). produces microbubbles with prolonged survival in blood. We hypothesized, therefore, that this prolonged ultrasound contrast effect could be utilized to non-invasively evaluate RABF and perfusion, Accordingly, we gave intravenous injections (IVI) of PESDA (0.06 cc/kg) to seven dogs while imaging with an external 4.5 MHz linear array transducer RABF was monitored using a Transonic Doppler probe placed around the renal artery. Contrast two-dimensional enhancement was quantified off-line. Both echo and color Doppler enhancement were also qualitatively graded as 0 = no enhancement, 1+ = mild, 2+ = marked enhancement. Following all 36 (100%) IVI of PESDA, there was 2+ contrast ultrasound enhancement of the renal cortex. A linear correlation existed between Doppler renal artery flow and peak renal cortex videointensity follOWing IV PESDA (r = 0.65, P < 0.001), Color Doppler signals were also enhanced following IV PESDA, and resulted in excellent visualization of the main renal artery as well as segmental and lobar arteries. When renal artery stenosis was induced to decrease RABF to less than 10% of baseline, the segmental and lobar arteries were not visualized with color Doppler following IV PESDA, and peak renal cortex videointensity was reduced from 26 ± 10 to 15 ± 8 units (p < 0.05). These data demonstrate that renal artery and cortical blood flow abnormalities can be detected using intravenous PESDA. This ultrasound contrast agent could be a new non-invasive method to detect renal artery stenosis or abnormal renal perfusion.

Ultrasonically enhanced transdermal drug delivery. Experimental approaches to elucidate the mechanism.

The effect of therapeutic range ultrasound on skin permeability was studied in vitro. Permeating molecule ionization state, pH, ultrasound duration, reversibility of the enhancement phenomenon, and skin structural alterations were evaluated. It was found that ultrasound affects the permeability of both ionized and unionized molecules. No irreversible structural alterations due to the ultrasound exposure were detected in the stratum corneum. Ultrasound enhancing mechanism was discussed.

Ultrasound enhancement of thrombolysis and reperfusion in vitro

Objectives. The aims of this study were 1) to develop an in vitro flow system in which reperfusion mediated by ultrasound-accelerated thrombolysis could be studied, and 2) to test whether ultrasound-accelerated thrombolysis could hasten reperfusion in this system.Background. Ultrasound has been shown to increase tissue plasminogen activator (t-PA)-induced thrombolysis in vitro as assessed by radioactive fibrinogen release from labeled clots and in an animal in vivo model.Methods. To test whether reperfusion is accelerated, we created obstructive whole blood clots in an in vitro flow system. Four control clots were exposed to ultrasound only without any thrombolytic agent (group 1). Sixteen clots were exposed to continuous infusion of recombinant tissue-type plasminogen activator rt-PA and randomized to either continuous wave ultrasound exposure at a frequency of 0.5 MHz and an intensity of 8 W/cm2(group 2) or to no ultrasound (group 3). Flow distal to the clot and the rate of release of radiolabeled fibrin products were used as an index of reperfusion and thrombolysis, respectively. Samples were obtained for measurements of lytic variables such as plasminogen, fibrinogen and rt-PA concentrations.Results. Flow was significantly higher in the rt-PA-treated clots within 10 min of exposure to ultrasound than in those without such exposure (9.4 ± 9.9% of maximal flow in group 2 vs. 0.5 ± 1.5% in group 3, p < 0.05). The maximal difference in flow between groups 2 and 3 was achieved at 25 min (61.0 ± 30.4% vs. 14.2 ± 14.7%, p = 0.03). Thrombolysis was significantly higher after 15 min of ultrasound exposure (12.8 ± 9.1% in the ultrasound-treated group 2 vs. 4.0 ± 3.9% in group 3, p < 0.05). The maximal difference between groups 2 and 3 occurred at 25 min (26.7 ± 13.1% vs. 7.24 ± 5.7%, p < 0.004). Neither flow nor clot lysis occurred in group 1. Plasminogen and fibrinogen concentrations and rt-PA antigen concentrations were consistent with those observed during fibrinolytic therapy in vivo.Conclusions. Continuous wave ultrasound at 0.5 MHz and an intensity of 8 W/cm2accelerates rt-PA-induced thrombolysis and reperfusion in vitro.

Study of use of ultrasound technology to prepare polymer-impregnated concrete

Low intensity ultrasound waves (20 kHz, 0.19 to 0.57 W/cm 2 based on concrete area) have been applied to enhance the efficiency of impregnating a monomer into hardened concrete, without prior drying, for subsequent in situ polymerization. It was found that methyl methacrylate could be impregnated into partly air-dried Portland cement concrete, without removing the free water in the specimen, to approximately 80 percent of the depth of that into a similar, previously oven-dried specimen. A systematic study has been carried out to assess the relationship between impregnation depth and the following parameters: ultrasound energy density, impregnation time, gap distance between the ultrasound horn and the specimen surface, and the applied DC voltage gradient. The results of the experiments and a proposed mechanism for the ultrasound enhancement are presented in this paper.

Enhancement of Fibrinolysis with Ultrasound Energy

The effect of ultrasound energy on fibrinolysis of artificial thrombus in vitro was investigated. Thrombi produced by the Chandler loop method were exposed to low-energy ultrasound (5,000-6,000 Pa) in an ultrasound bath (48 kHz) for 60 seconds. Fibrinolysis with urokinase was enhanced from 40.6% +/- 1.8% to 59.2% +/- 2.6% (mean +/- standard deviation) with ultrasound exposure after a 60-minute incubation. Ultrasound alone without urokinase resulted in no fibrinolysis. In a second experiment, a newly developed miniature ultrasound-emitting ceramic element (2 x 1 x 5 mm) was attached to the tip of a catheter. Ultrasound exposure (225 kHz) from this device markedly enhanced fibrinolysis with urokinase from 8.9% +/- 1.5% to 37.3% +/- 0.8% (total ultrasound exposure 60 seconds, intensity 30 mW/cm2) after a 30-minute incubation. After a 120-minute incubation, fibrinolysis with ultrasound exposure was 61.1% +/- 1.8% versus 46.7% +/- 0.5% for the unexposed group. Ultrasound enhancement of fibrinolysis was less pronounced with longer incubation time (60 or 120 minutes). Ultrasound energy enhanced fibrinolysis with urokinase, especially in the early phase of lysis. This new device may shorten the time needed to complete fibrinolysis and reduce total drug dosage needed for treatment of thromboembolic diseases.

4948587 Ultrasound enhancement of transbuccal drug delivery: Joseph Kost, Robert S Langer, Omer, MA, Israel assigned to Massachusetts Institute of Technology

4948587 Ultrasound enhancement of transbuccal drug delivery: Joseph Kost, Robert S Langer, Omer, MA, Israel assigned to Massachusetts Institute of Technology

Ultrasound enhancement of transdermal drug delivery

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Ultrasound enhanced bioprocesses: Cholesterol oxidation by Rhodococcus erythropolis

Cholesterol oxidation to cholestenone by resting cells of Rhodococcus erythropolis ATCC25544 was investigated under a computer-controlled ultrasonic irradiation at a frequency of 20 kHz. The optimization of the ultrasound intensity and its mode of application to a stirred bioreactor was first established at a level which preserved the structural integrity of the cells and enabled their metabolic activity. A significant enhancement in the kinetic rates of the biotransformation was observed in microbial slurries of 1.0 and 2.5 g/L cholesterol when sonicated for 5 s every 10 min with a power output of 2.2 W/cm(2). In contrast, ultrasound had no effect on the enzymatic oxidation of cholesterol (2.5 g/L) by cholesterol oxidase. A high loading of cholesterol (5.0 g/L) in sonicated microbial systems had, however, an adverse effect. The ultrasound enhancement is discussed in terms of an increased dissolution rate of the sustrate crystals and more importantly, in terms of the uniquely ultrasound-induced enhancement of mass transfer inside and outside a cell.