Investigating the spatial extent of acoustically activated echogenic liposomes.

The purpose of this work was to investigate the ability of bubbles entrapped within echogenic liposomes (ELIP) to serve as foci for cavitational events that would cause leakage in neighboring non-echogenic liposomes (NELIP). Previous studies have shown that entrapping bubbles into liposomes increases ultrasound-mediated leakage of hydrophilic components at ultrasound settings known to induce inertial cavitation, specifically 20kHz and 2.2W/cm2. Using tone-burst approach and pulse repetition frequency of 10Hz would bring this intensity level to the one accepted (220mW/cm2) in clinical imaging. Mixed populations of ELIP and NELIP were simultaneously exposed to ultrasound at varying ratios to examine the effect of ELIP concentration on release of a hydrophilic dye, calcein, from NELIP. Calcein release from NELIP was observed to be independent of ELIP concentration, suggesting that the release enhancement from echogenicity is strictly a localized event. Additionally, it was observed that the release mechanisms independent of echogenicity were active for the duration of experiment whereas those associated with echogenicity were active for only the initial 1-2min.

Enhanced therapeutic anti-inflammatory effect of betamethasone on topical administration with low-frequency, low-intensity (20 kHz, 100 mW/cm(2)) ultrasound exposure on carrageenan-induced arthritis in a mouse model.

The purpose of this work was to investigate whether low-frequency, low-intensity (20 kHz, <100 mW/cm(2), spatial-peak, temporal-peak intensity) ultrasound, delivered with a lightweight (<100 g), tether-free, fully wearable, battery-powered applicator, is capable of reducing inflammation in a mouse model of rheumatoid arthritis. The therapeutic, acute, anti-inflammatory effect was estimated from the relative swelling induced in mice hindlimb paws. In an independent, indirect approach, the inflammation was bio-imaged by measuring glycolytic activity with near-infrared labeled 2-deoxyglucose. The outcome of the experiments indicated that the combination of ultrasound exposure and topical application of 0.1% (w/w) betamethasone gel resulted in statistically significantly (p < 0.05) enhanced anti-inflammatory activity in comparison with drug or ultrasound treatment alone. The present study underscores the potential benefits of low-frequency, low-intensity ultrasound-assisted drug delivery. However, the proof of concept presented indicates the need for additional experiments to systematically evaluate and optimize the potential of, and the conditions for, tolerable low-frequency, low-intensity ultrasound-promoted non-invasive drug delivery.

Ultrasound-induced modulation of cardiac rhythm in neonatal rat ventricular cardiomyocytes.

Isolated neonatal rat ventricular cardiomyocytes were used to study the influence of ultrasound on the chronotropic response in a tissue culture model. The beat frequency of the cells, varying from 40 to 90 beats/min, was measured based upon the translocation of the nuclear membrane captured by a high-speed camera. Ultrasound pulses (frequency = 2.5 MHz) were delivered at 300-ms intervals [3.33 Hz pulse repetition frequency (PRF)], in turn corresponding to 200 pulses/min. The intensity of acoustic energy and pulse duration were made variable, 0.02-0.87 W/cm(2) and 1-5 ms, respectively. In 57 of 99 trials, there was a noted average increase in beat frequency of 25% with 8-s exposures to ultrasonic pulses. Applied ultrasound energy with a spatial peak time average acoustic intensity (Ispta) of 0.02 W/cm(2) and pulse duration of 1 ms effectively increased the contraction rate of cardiomyocytes (P < 0.05). Of the acoustic power tested, the lowest level of acoustic intensity and shortest pulse duration proved most effective at increasing the electrophysiological responsiveness and beat frequency of cardiomyocytes. Determining the optimal conditions for delivery of ultrasound will be essential to developing new models for understanding mechanoelectrical coupling (MEC) and understanding novel nonelectrical pacing modalities for clinical applications.

Low-frequency (<100 kHz), low-intensity (<100 mW/cm(2)) ultrasound to treat venous ulcers: a human study and in vitro experiments.

The purpose of this study was to examine whether low frequency (<100 kHz), low intensity (<100 mW/cm(2), spatial peak temporal peak) ultrasound can be an effective treatment of venous stasis ulcers, which affect 500 000 patients annually costing over $1 billion per year. Twenty subjects were treated with either 20 or 100 kHz ultrasound for between 15 and 45 min per session for a maximum of four treatments. Healing was monitored by changes in wound area. Additionally, two in vitro studies were conducted using fibroblasts exposed to 20 kHz ultrasound to confirm the ultrasound's effects on proliferation and cellular metabolism. Subjects receiving 20 kHz ultrasound for 15 min showed statistically faster (p < 0.03) rate of wound closure. All five of these subjects fully healed by the fourth treatment session. The in vitro results indicated that 20 kHz ultrasound at 100 mW/cm(2) caused an average of 32% increased metabolism (p < 0.05) and 40% increased cell proliferation (p < 0.01) after 24 h when compared to the control, non-treated cells. Although statistically limited, this work supports the notion that low-intensity, low-frequency ultrasound is beneficial for treating venous ulcers.

Influence of shell composition on the resonance frequency of microbubble contrast agents.

The effect of variations in microbubble shell composition on microbubble resonance frequency is revealed through experiment. These variations are achieved by altering the mole fraction and molecular weight of functionalized polyethylene glycol (PEG) in the microbubble phospholipid monolayer shell and measuring the microbubble resonance frequency. The resonance frequency is measured via a chirp pulse and identified as the frequency at which the pressure amplitude loss of the ultrasound wave is the greatest as a result of passing through a population of microbubbles. For the shell compositions used herein, we find that PEG molecular weight has little to no influence on resonance frequency at an overall PEG mole fraction (0.01) corresponding to a mushroom regime and influences the resonance frequency markedly at overall PEG mole fractions (0.050-0.100) corresponding to a brush regime. Specifically, the measured resonance frequency was found to be 8.4, 4.9, 3.3 and 1.4 MHz at PEG molecular weights of 1000, 2000, 3000 and 5000 g/mol, respectively, at an overall PEG mole fraction of 0.075. At an overall PEG mole fraction of just 0.01, on the other hand, resonance frequency exhibited no systematic variation, with values ranging from 5.7 to 4.9 MHz. Experimental results were analyzed using the Sarkar bubble dynamics model. With the dilatational viscosity held constant (10(-8) N·s/m) and the elastic modulus used as a fitting parameter, model fits to the pressure amplitude loss data resulted in elastic modulus values of 2.2, 2.4, 1.6 and 1.8 N/m for PEG molecular weights of 1000, 2000, 3000 and 5000 g/mol, respectively, at an overall PEG mole fraction of 0.010 and 4.2, 1.4, 0.5 and 0.0 N/m, respectively, at an overall PEG mole fraction of 0.075. These results are consistent with theory, which predicts that the elastic modulus is constant in the mushroom regime and decreases with PEG molecular weight to the inverse 3/5 power in the brush regime. Additionally, these results are consistent with inertial cavitation studies, which revealed that increasing PEG molecular weight has little to no effect on inethe rtial cavitation threshold in the mushroom regime, but that increasing PEG molecular weight decreases inertial cavitation markedly in the brush regime. We conclude that the design and synthesis of microbubbles with a prescribed resonance frequency is attainable by tuning PEG composition and molecular weight.

Finite element static displacement optimization of 20-100 kHz flexural transducers for fully portable ultrasound applicator.

This paper focuses on the development of a finite-element model and subsequent stationary analysis performed to optimize individual flexural piezoelectric elements for operation in the frequency range of 20-100kHz. These elements form the basic building blocks of a viable, un-tethered, and portable ultrasound applicator that can produce intensities on the order of 100mW/cm(2) spatial-peak temporal-peak (I(SPTP)) with minimum (on the order of 15V) excitation voltage. The ultrasound applicator can be constructed with different numbers of individual transducer elements and different geometries such that its footprint or active area is adjustable. The primary motivation behind this research was to develop a tether-free, battery operated, fully portable ultrasound applicator for therapeutic applications such as wound healing and non-invasive transdermal delivery of both naked and encapsulated drugs. It is shown that careful selection of the components determining applicator architecture allows the displacement amplitude to be maximized for a specific frequency of operation. The work described here used the finite-element analysis software COMSOL to identify the geometry and material properties that permit the applicator's design to be optimized. By minimizing the excitation voltage required to achieve the desired output (100mW/cm(2)I(SPTP)) the power source (rechargeable Li-Polymer batteries) size may be reduced permitting both the electronics and ultrasound applicator to fit in a wearable housing.

Optimization of un-tethered, low voltage, 20-100kHz flexural transducers for biomedical ultrasonics applications.

This paper describes optimization of un-tethered, low voltage, 20-100kHz flexural transducers for biomedical ultrasonics applications. The goal of this work was to design a fully wearable, low weight (<100g), battery operated, piezoelectric ultrasound applicator providing maximum output pressure amplitude at the minimum excitation voltage. Such implementation of ultrasound applicators that can operate at the excitation voltages on the order of only 10-25V is needed in view of the emerging evidence that spatial-peak temporal-peak ultrasound intensity (I(SPTP)) on the order of 100mW/cm(2) delivered at frequencies below 100kHz can have beneficial therapeutic effects. The beneficial therapeutic applications include wound management of chronic ulcers and non-invasive transdermal delivery of insulin and liposome encapsulated drugs. The early prototypes of the 20 and 100kHz applicators were optimized using the maximum electrical power transfer theorem, which required a punctilious analysis of the complex impedance of the piezoelectric disks mounted in appropriately shaped metal housings. In the implementation tested, the optimized ultrasound transducer applicators were driven by portable, customized electronics, which controlled the excitation voltage amplitude and facilitated operation in continuous wave (CW) or pulsed mode with adjustable (10-90%) duty cycle. The driver unit was powered by remotely located rechargeable lithium (Li) polymer batteries. This was done to further minimize the weight of the applicator unit making it wearable. With DC voltage of approximately 15V the prototypes were capable of delivering pressure amplitudes of about 55kPa or 100mW/cm(2) (I(SPTP)). This level of acoustic output was chosen as it is considered safe and side effects free, even at prolonged exposure.