MPa Peak Rarefactional Pressure Research Articles

Quantitative evaluation of anti-biofilm cavitation activity seeded from microbubbles or protein cavitation nuclei by passive acoustic mapping.

Bacterial biofilms represent a major challenge for effective antibiotic therapy as they confer physical and functional changes that protect bacteria from their surrounding environment. In this work, focused ultrasound in combination with cavitation nuclei was used to disrupt biofilms of Staphylococcus aureus and Pseudomonas aeruginosa, both of which are on the World Health Organization's priority list for new antimicrobial research. 
Approach: Single species biofilms were exposed to ultrasound (0.5 MHz centre frequency, 0.5-1.5 MPa peak rarefactional pressure, 200 cycle pulses, 5 Hz repetition frequency, 30 s duration), in the presence of two different types of cavitation nuclei. Quantitative passive acoustic mapping (PAM) was used to monitor cavitation emissions during treatment using a calibrated linear array. 
Main Results: It was observed that the cumulative energy of acoustic emissions during treatment was positively correlated with biofilm disruption, with differences between bacterial species attributed to differences in biofilm morphology. PCaN provided increased biofilm reduction compared to microbubbles due in large part to their persistence over the duration of ultrasound exposure. There was also good correlation between the spatial distribution of cavitation as characterized by PAM and the extent of biofilm disruption observed with microscopy. 
Significance: Collectively, the results from this work indicate the potential broad applicability of cavitation for eliminating biofilms of priority pathogens and the opportunity presented by Passive Acoustic Mapping for real-time monitoring of antimicrobial processes.

Spatio-temporal evaluation of anti-biofilm cavitation activity by passive acoustic mapping

Bacterial biofilms present a major challenge to achieving effective antibiotic therapy, as these sessile communities of microbes confer protection to bacteria by decreasing antibiotic efficacy. Focused ultrasound can mechanically disrupt biofilms, offering a new ‘drug-free’ antibiotic paradigm. The goal of this work is to validate, quantify, and optimize the role of acoustic cavitation in the biofilm disruption process through spatiotemporal monitoring of cavitation activity. A clinical isolate strain of Staphylococcus aureus from native valve endocarditis was cultured for 72 hours within a flow channel to form a biofilm. A 1.1 MHz spherically focused transducer was used to expose the biofilm from below at a 45° angle. The in situ acoustic field was characterized with a fibre-optic hydrophone. A calibrated 5–11 MHz linear array was placed 25 mm above the biofilm in order to record acoustic emissions during biofilm disruption from which passive acoustic maps of cavitation could be derived. Biofilms were exposed to 4.5 MPa peak rarefactional pressure (derated), 10,000 cycles, at a 1 Hz PRF. Qualitative reduction of biofilms was assessed by live/dead staining with Syto 9/propidium iodide, which was correlated with cavitation activity observed in the passive acoustic maps.

Micro-Particle Image Velocimetry Investigation of Flow Fields of SonoVue Microbubbles Mediated by Ultrasound and Their Relationship With Delivery

The flow fields generated by the acoustic behavior of microbubbles can significantly increase cell permeability. This facilitates the cellular uptake of external molecules in a process known as ultrasound-mediated drug delivery. To promote its clinical translation, this study investigated the relationships among the ultrasound parameters, acoustic behavior of microbubbles, flow fields, and delivery results. SonoVue microbubbles were activated by 1 MHz pulsed ultrasound with 100 Hz pulse repetition frequency, 1:5 duty cycle, and 0.20/0.35/0.70 MPa peak rarefactional pressure. Micro-particle image velocimetry was used to detect the microbubble behavior and the resulting flow fields. Then HeLa human cervical cancer cells were treated with the same conditions for 2, 4, 10, 30, and 60 s, respectively. Fluorescein isothiocyanate and propidium iodide were used to quantitate the rates of sonoporated cells with a flow cytometer. The results indicate that (1) microbubbles exhibited different behavior in ultrasound fields of different peak rarefactional pressures. At peak rarefactional pressures of 0.20 and 0.35 MPa, the dispersed microbubbles clumped together into clusters, and the clusters showed no apparent movement. At a peak rarefactional pressure of 0.70 MPa, the microbubbles were partially broken, and the remainders underwent clustering and coalescence to form bubble clusters that exhibited translational oscillation. (2) The flow fields were unsteady before the unification of the microbubbles. After that, the flow fields showed a clear pattern. (3)The delivery efficiency improved with the shear stress of the flow fields increased. Before the formation of the microbubble/bubble cluster, the maximum shear stresses of the 0.20, 0.35, and 0.70 MPa groups were 56.0, 87.5 and 406.4 mPa, respectively, and the rates of the reversibly sonoporated cells were 2.4% ± 0.4%, 5.5% ± 1.3%, and 16.6% ± 0.2%. After the cluster formation, the maximum shear stresses of the three groups were 9.1, 8.7, and 71.7 mPa, respectively. The former two could not mediate sonoporation, whereas the last one could. These findings demonstrate the critical role of flow fields in ultrasound-mediated drug delivery and contribute to its clinical applications.

Standing wave assisted acoustic droplet vaporization for dual payload release from acoustically responsive scaffolds

Ultrasound standing waves have been utilized for many biomedical applications. Here, we demonstrate how standing waves can enhance drug release using acoustic droplet vaporization (ADV), which is the phase-transitioning of perfluorocarbon (PFC) emulsions via ultrasound. These experiments utilized acoustically-responsive scaffolds (ARSs), which are composite fibrin hydrogels containing payload-carrying, monodispersed PFC emulsions. Single- and bi-layer ARSs were generated with dextran-loaded emulsions (diameter: 6 mm) containing either perfluorohexane, or perfluorooctane in each layer. First, we studied the influence of standing waves on payload release from single-layer ARSs. At 4 MPa peak rarefactional pressure, elevated amplitudes due to constructive superposition in the standing wave field enhanced payload release up to 35% at 2.5 MHz in a seven-day longitudinal study. Second, the effect of standing waves was combined with the frequency-dependent ADV to enhance dual payload release from bi-layer ARSs. We demonstrated the sequential release of two dextran payloads from ARSs, which were, respectively, contained within each emulsion, using temporally staggered ADV at 3 MHz (day 0) and 8.8 MHz (day 4). These results will also be discussed in the context of practical strategies for achieving similar conditions for in vivo applicationsUltrasound standing waves have been utilized for many biomedical applications. Here, we demonstrate how standing waves can enhance drug release using acoustic droplet vaporization (ADV), which is the phase-transitioning of perfluorocarbon (PFC) emulsions via ultrasound. These experiments utilized acoustically-responsive scaffolds (ARSs), which are composite fibrin hydrogels containing payload-carrying, monodispersed PFC emulsions. Single- and bi-layer ARSs were generated with dextran-loaded emulsions (diameter: 6 mm) containing either perfluorohexane, or perfluorooctane in each layer. First, we studied the influence of standing waves on payload release from single-layer ARSs. At 4 MPa peak rarefactional pressure, elevated amplitudes due to constructive superposition in the standing wave field enhanced payload release up to 35% at 2.5 MHz in a seven-day longitudinal study. Second, the effect of standing waves was combined with the frequency-dependent ADV to enhance dual payload release from bi-layer ARSs. ...

Capillary Hemorrhage Induced by Contrast-Enhanced Diagnostic Ultrasound in Rat Intestine

Contrast-enhanced diagnostic ultrasound (CEDUS) can lead to microvascular injury and petechial hemorrhage by the cavitational mechanism of ultrasonic bioeffects. Capillary hemorrhage has been noted in the heart and kidney, which are common targets of CEDUS examination. CEDUS has also become useful for monitoring intestinal inflammation. In the 1990s, the risk of intestinal microvascular hemorrhage was investigated both for incidental exposure by lithotripter shockwaves and for contrast agent microbubbles acting as cavitation nuclei with laboratory pulsed ultrasound systems. This study was initiated to update the risk assessment for intestine exposed to diagnostic imaging simulating CEDUS. The abdomens of anesthetized rats were scanned by a 1.6 MHz phased array probe during infusion of microbubble suspensions simulating Definity ultrasound contrast agent. Dual image frames were triggered intermittently, and the output power was varied to assess the exposure response. Petechiae counts in small intestine mucosa and muscle layers increased with increasing trigger interval from 2 s to 10 s, indicative of a slow refill after microbubble destruction. The counts increased with increasing output above a threshold of 1.4 MPa peak rarefactional pressure amplitude. Petechiae were also seen in Peyer's patches, and occult blood was detected in many affected segments of intestine. These results are consistent with early laboratory pulsed-ultrasound results.

Multiple ultrasound cavitation-enabled treatments for myocardial reduction

BackgroundUltrasound myocardial cavitation enabled treatment (MCET) is an image-guided method for tissue reduction. In this study, a strategy of fractionated (multiple) treatments was tested for efficacy.MethodsDahl SS rats were anesthetized and prepared for treatment with a focused ultrasound transducer in a warm water bath. Aiming at the anterior left ventricular wall was facilitated by imaging with a 10 MHz phased array (10S, GE Vivid 7, GE Vingmed Ultrasound, Horten, Norway). MCET was accomplished at 1.5 MHz by pulse bursts of 4 MPa peak rarefactional pressure amplitude, which were intermittently triggered 1:8 from the ECG during infusion of a microbubble suspension for cavitation nucleation. Test groups were sham, a 200 s treatment, three 200 s treatments a week apart, and a 600 s treatment. Treatment outcome was observed by plasma troponin after 4 h, echocardiographic monitoring and histology at 6 wk.ResultsThe impacts of the fractionated treatments summed to approximately the same as the long treatment; e. g. the troponin result was 10.5 ± 3.2 for 200 s, 22.7 ± 5.4 (p < 0.001) for the summed fractionated treatments and 29.9 ± 6.4 for 600 s (p = 0.06 relative to the summed fractionated). While wall thickness was not reduced for the fractionated treatment, tissue strain was reduced by 35% in the target area relative sham (p < 0.001).ConclusionThe ability to fractionate treatment may be advantageous for optimizing patient outcome relative to all-or nothing therapy by surgical myectomy or alcohol ablation.

Ultrasonic cavitation-enabled treatment for therapy of hypertrophic cardiomyopathy: Proof-of-principle

Ultrasound myocardial cavitation enabled treatment (MCET) creates scattered microlesions in the myocardium, which can be accumulated to produce a desired macrolesion. MCET was applied to the SS-16BN rat model of hypertrophic cardiomyopathy (HCM) for proof-of-principle as a means for myocardial reduction. A focused ultrasound transducer was targeted using 10 MHz imaging (10S, GE Vivid 7) to the left ventricular wall of anesthetized rats in a warmed water bath. Pulse bursts of 4 MPa peak rarefactional pressure amplitude were intermittently triggered 1:8 heartbeats during 10 min infusion of a microbubble suspension. Methylprednisolone was given to reduce initial inflammation and Losartan was given to improve healing. MCET significantly reduced the targeted wall thickness (n = 11) at 28 d post treatment by 16.2% (P<0.01) relative to shams (n = 8), with myocardial strain rate and endocardial border displacement reduced by 34% and 29%, respectively. This demonstrates sufficient effect for a therapeutic outcome ...

Enhanced delivery of paclitaxel liposomes using focused ultrasound with microbubbles for treating nude mice bearing intracranial glioblastoma xenografts.

Paclitaxel liposomes (PTX-LIPO) are a clinically promising antineoplastic drug formulation for the treatment of various extracranial cancers, excluding glioblastoma. A main reason for this is the presence of the blood–brain barrier (BBB) or blood–tumor barrier (BTB), preventing liposomal drugs from crossing at a therapeutically meaningful level. Focused ultrasound (FUS) in conjunction with microbubbles (MBs) has been suggested in many studies to be an effective approach to increase the BBB or BTB permeability. In this study, we investigated the feasibility of enhancing the delivery of PTX-LIPO in intracranial glioblastoma-bearing nude mice using pulsed low-intensity FUS exposure in the presence of MBs. Our results showed that the delivery efficiency of PTX-LIPO could be effectively improved in terms of the penetration of both the BBB in vitro and BTB in vivo by pulsed FUS sonication with a 10 ms pulse length and 1 Hz pulse repetition frequency at 0.64 MPa peak-rarefactional pressure in the presence of MBs. Quantitative analysis showed that a 2-fold higher drug concentration had accumulated in the glioblastoma 3 h after FUS treatment, with 7.20±1.18 µg PTX per g glioma tissue. Longitudinal magnetic resonance imaging analysis illustrated that the intracranial glioblastoma progression in nude mice treated with PTX-LIPO delivered via FUS with MBs was suppressed consistently for 4 weeks compared to the untreated group. The medium survival time of these tumor-bearing nude mice was significantly prolonged by 20.8%, compared to the untreated nude mice. Immunohistochemical analysis further confirmed the antiproliferation effect and cell apoptosis induction. Our study demonstrated that noninvasive low-intensity FUS with MBs can be used as an effective approach to deliver PTX-LIPO in order to improve their chemotherapy efficacy toward glioblastoma.

Controlled release of basic fibroblast growth factor for angiogenesis using acoustically-responsive scaffolds.

The clinical translation of pro-angiogenic growth factors for treatment of vascular disease has remained a challenge due to safety and efficacy concerns. Various approaches have been used to design spatiotemporally-controlled delivery systems for growth factors in order to recapitulate aspects of endogenous signaling and thus assist in translation. We have developed acoustically-responsive scaffolds (ARSs), which are fibrin scaffolds doped with a payload-containing, sonosensitive emulsion. Payload release can be controlled non-invasively and in an on-demand manner using focused, megahertz-range ultrasound (US). In this study, we investigate the invitro and invivo release from ARSs containing basic fibroblast growth factor (bFGF) encapsulated in monodispersed emulsions. Emulsions were generated in a two-step process utilizing a microfluidic device with a flow focusing geometry. At 2.5MHz, controlled release of bFGF was observed for US pressures above 2.2±0.2MPa peak rarefactional pressure. Superthreshold US yielded a 12.6-fold increase in bFGF release invitro. The bioactivity of the released bFGF was also characterized. When implanted subcutaneously in mice, ARSs exposed to superthreshold US displayed up to 3.3-fold and 1.7-fold greater perfusion and blood vessel density, respectively, than ARSs without US exposure. Scaffold degradation was not impacted by US. These results highlight the utility of ARSs in both basic and applied studies of therapeutic angiogenesis.

Effect of acoustic parameters on the cavitation behavior of SonoVue microbubbles induced by pulsed ultrasound

SonoVue microbubbles could serve as artificial nuclei for ultrasound-triggered stable and inertial cavitation, resulting in beneficial biological effects for future therapeutic applications. To optimize and control the use of the cavitation of SonoVue bubbles in therapy while ensuring safety, it is important to comprehensively understand the relationship between the acoustic parameters and the cavitation behavior of the SonoVue bubbles. An agarose-gel tissue phantom was fabricated to hold the SonoVue bubble suspension. 1-MHz transmitting transducer calibrated by a hydrophone was used to trigger the cavitation of SonoVue bubbles under different ultrasonic parameters (i.e., peak rarefactional pressure (PRP), pulse repetition frequency (PRF), and pulse duration (PD)). Another 7.5-MHz focused transducer was employed to passively receive acoustic signals from the exposed bubbles. The ultraharmonics and broadband intensities in the acoustic emission spectra were measured to quantify the extent of stable and inertial cavitation of SonoVue bubbles, respectively. We found that the onset of both stable and inertial cavitation exhibited a strong dependence on the PRP and PD and a relatively weak dependence on the PRF. Approximate 0.25MPa PRP with more than 20μs PD was considered to be necessary for ultraharmonics emission of SonoVue bubbles, and obvious broadband signals started to appear when the PRP exceeded 0.40MPa. Moreover, the doses of stable and inertial cavitation varied with the PRP. The stable cavitation dose initially increased with increasing PRP, and then decreased rapidly after 0.5MPa. By contrast, the inertial cavitation dose continuously increased with increasing PRP. Finally, the doses of both stable and inertial cavitation were positively correlated with PRF and PD. These results could provide instructive information for optimizing future therapeutic applications of SonoVue bubbles.

Design and Characterization of Fibrin-Based Acoustically Responsive Scaffolds for Tissue Engineering Applications

Hydrogel scaffolds are used in tissue engineering as a delivery vehicle for regenerative growth factors. Spatiotemporal patterns of growth factor signaling are critical for tissue regeneration, yet most scaffolds afford limited control of growth factor release, especially after implantation. We previously found that acoustic droplet vaporization can control growth factor release from a fibrin scaffold doped with a perfluorocarbon emulsion. This study investigates properties of the acoustically responsive scaffold (ARS) critical for further translation. At 2.5 MHz, acoustic droplet vaporization and inertial cavitation thresholds ranged from 1.5 to 3.0 MPa and from 2.0 to 7.0 MPa peak rarefactional pressure, respectively, for ARSs of varying composition. Viability of C3H/10T1/2 cells, encapsulated in the ARS, did not decrease significantly for pressures below 4 MPa. ARSs with perfluorohexane emulsions displayed higher stability versus those with perfluoropentane emulsions, while surrogate payload release was minimal without ultrasound. These results enable the selection of ARS compositions and acoustic parameters needed for optimized spatiotemporally controlled release.

Myocardial cavitation-enabled therapy modeling

A therapeutic method named Myocardial Cavitation Enabled Therapy aiming at noninvasive cardiac tissue reduction is modeled here. Sparsely distributed microlesions induced by ultrasound cavitation of contrast agents are hypothesized to cause myocardial shrinkage. The objective is to model lesion formation based on the acoustic field and plan treatments accordingly. An ultrasound field simulation was established in Field II, an acoustics toolbox. It simulates the acoustic field of a 1.5 MHz ultrasound burst of five cycles at 4.0 MPa peak rarefactional pressure amplitude (PRPA) by use of a F# 2 single element therapeutic transducer, as used in concomitant animal studies. Medium parameters, including speed of sound, density, absorption, and B/A were set to water path and heart tissue along the acoustic path. Lesions were masked as region exceeding 2 MPa PRPA, the threshold of microlesions occurrence. The lesion volume is 143 μL compared to in vivo rodent study of approximately 100 μL. To reach a larger lesion in pigs, sweeping of the ultrasound beam is needed using a phased array. Nonlinear acoustic simulation from k-Wave software will assist transitioning from single element to imaging array apertures in terms of interpreting possibly different lesion formation resulting from nonlinearities.

Frequency dependence of thresholds for lethal cardiomyocyte injury in myocardial contrast echocardiography

Contrast enhanced diagnostic ultrasound employs microbubble activation for microvascular imaging; however, the on-screen Mechanical Index is a poor parameter for safety guidance. More research is needed on microvascular bioeffects, particularly their variation with frequency. A GE Vivid 7 with an S3 probe operated at 1.6 MHz, and an S5 probe operated at 2.5 and 3.5 MHz was used for myocardial contrast echocardiography of rats mounted in a water bath. Power settings were varied in 2 dB steps for determination of the thresholds for cardiomyocyte injury. The contrast agent was made to duplicate the properties of the clinical agent Definity. The scans were intermittently triggered each 4 heartbeats from the ECG signal. The cardiomyocyte death was assessed using Evans blue vital staining. Thresholds were defined as the mean of the lowest exposure with a statistically significant cardiomyocyte death and the next lower exposure level. Thresholds were 1.2 MPa, 1.7 MPa, and 2.7 MPa peak rarefactional pressure amplitude (derated for 1 dB/cm/MHz attenuation) for 1.6, 2.5, and 3.5 MHz, respectively. Linear regression showed that the thresholds were essentially proportional to frequency (0.72 f1.02, r2 = 0.97). Theoretical analysis is under development to explain this dependence and develop a contrast-specific safety parameter.

Pulmonary Capillary Hemorrhage Induced by Fixed-Beam Pulsed Ultrasound

The induction of pulmonary capillary hemorrhage (PCH) by pulsed ultrasound was discovered 25 y ago, but early research used fixed-beam systems rather than actual diagnostic ultrasound machines. In this study, results of exposure of rats to fixed-beam focused ultrasound for 5 min at 1.5 and 7.5 MHz were compared with recent research on diagnostic ultrasound. One exposure condition at each frequency used 10-μs pulses delivered at 25-ms intervals. Three conditions involved Gaussian modulation of the pulse amplitudes at 25-ms intervals to simulate diagnostic scanning: 7.5 MHz with 0.3- and 1.5-μs pulses at 100- and 500-μs pulse repetition periods, respectively, and 1.5 MHz with 1.7-μs pulses at 500-μs repetition periods. Four groups were tested for each condition to assess PCH areas at different exposure levels and to determine occurrence thresholds. The conditions with identical pulse timing resulted in smaller PCH areas for the smaller 7.5-MHz beam, but both had thresholds of 0.69–0.75 MPa in situ peak rarefactional pressure amplitude. The Gaussian modulation conditions for both 7.5 MHz with 0.3-μs pulses and 1.5 MHz with 1.7-μs pulses had thresholds of 1.12–1.20 MPa peak rarefactional pressure amplitude, although the relatively long 1.5-μs pulses at 7.5 MHz yielded a threshold of 0.75 MPa. The fixed-beam pulsed ultrasound exposures produced lower thresholds than diagnostic ultrasound. There was no clear tendency for thresholds to increase with increasing ultrasonic frequency when pulse timing conditions were similar.

Cavitation-enhanced delivery of insulin in agar and porcine models of human skin

Ultrasound-assisted transdermal insulin delivery offers a less painful and less invasive alternative to subcutaneous insulin injections. However, ultrasound-based drug delivery, otherwise known as sonophoresis, is a highly variable phenomenon, in part dependent on cavitation. The aim of the current work is to investigate the role of cavitation in transdermal insulin delivery. Fluorescently stained, soluble Actrapid insulin was placed on the surface of human skin-mimicking materials subjected to 265 kHz, 10% duty cycle focused ultrasound. A confocally and coaxially aligned 5 MHz broadband ultrasound transducer was used to detect cavitation. Two different skin models were used. The first model, 3% agar hydrogel, was insonated with a range of pressures (0.25–1.40 MPa peak rarefactional focal pressure—PRFP), with and without cavitation nuclei embedded within the agar at a concentration of 0.05% w/v. The second, porcine skin was insonated at 1.00 and 1.40 MPa PRFP. In both models, fluorescence measurements were used to determine penetration depth and concentration of delivered insulin. Results show that in agar gel, both insulin penetration depth and concentration only increased significantly in the presence of inertial cavitation, with up to a 40% enhancement. In porcine skin the amount of fluorescent insulin was higher in the epidermis of those samples that were exposed to ultrasound compared to the control samples, but there was no significant increase in penetration distance. The results underline the importance of instigating and monitoring inertial cavitation during transdermal insulin delivery.

Ultrasound-induced heart rate decrease: role of the vagus nerve.

The goal of this study is to investigate the role of the vagus nerve (VN) in the ultrasound (US)-induced negative chronotropic effect (deceased heart rate). One of the functions of the VN is to mediate lowering of the heart rate. A previous study showed a decrease of ~20% in the heart rate but the mechanism of the effect was not investigated. Sprague Dawley rats (n = 20) were exposed transthoracically to ultrasonic pulses at an approximate duty factor of 1% with sequentially 2.0, 2.5, and 3.0 MPa peak rarefactional pressure amplitudes (PRPAs). The ultrasonic exposure parameters herein were chosen to match those of the previous study to have confidence that an ultrasound-induced negative chronotropic effect would occur. For each of the three PRPA sequences, the pulse repetition frequency (PRF) started slightly greater than the rat's heart rate and then was decreased sequentially in 1-Hz steps every 10 s (i.e., 6, 5, and 4 Hz for a total duration of 30 s). The experiments were organized in a standard (2 × 2) factorial design with VN (cut versus intact) as one factor and US (on versus off) as another factor. VN (intact/cut) and US (on/off) groups were divided into four groups each consisting of 5 animals: 1) VN intact-US off, 2) VN intact-US on, 3) VN cut-US off, and 4) VN cut-US on. Two-way analysis of variance for repeated measures was used to compare heart rate, cardiac output, systolic volume, ejection fraction, end-diastolic volume, end-systolic volume, respiratory rate, and arterial pressure before and after ultrasound stimulation. In this study, the heart rate decreased ~4% for the non-vagotomy and vagotomy groups. The ultrasound effect was significant for heart rate (p = 0.02) and cardiac output (p = 0.005) at 3 min post US exposure; the vagotomy effect was not significant. For heart rate, the Bonferroni test showed no differences between the four groups. The vagotomy group showed similar ultrasound-induced cardiac effects compared with the non-vagotomy group, suggesting that the vagus nerve is not influenced by the ultrasound exposure procedures. The US application caused a negative chronotropic effect of the rat heart without affecting the hemodynamic conditions. The results at this point are suggestive for an alternative cardiac pacing capability.

Lesion generation through ribs using histotripsy therapy without aberration correction

This study investigates the feasibility of using high-intensity pulsed therapeutic ultrasound, or histotripsy, to non-invasively generate lesions through the ribs. Histotripsy therapy mechanically ablates tissue through the generation of a cavitation bubble cloud, which occurs when the focal pressure exceeds a certain threshold. We hypothesize that histotripsy can generate precise lesions through the ribs without aberration correction if the main lobe retains its shape and exceeds the cavitation initiation threshold and the secondary lobes remain below the threshold. To test this hypothesis, a 750-kHz focused transducer was used to generate lesions in tissue-mimicking phantoms with and without the presence of rib aberrators. In all cases, 8000 pulses with 16 to 18 MPa peak rarefactional pressure at a repetition frequency of 100 Hz were applied without aberration correction. Despite the high secondary lobes introduced by the aberrators, high-speed imaging showed that bubble clouds were generated exclusively at the focus, resulting in well-confined lesions with comparable dimensions. Collateral damage from secondary lobes was negligible, caused by single bubbles that failed to form a cloud. These results support our hypothesis, suggesting that histotripsy has a high tolerance for aberrated fields and can generate confined focal lesions through rib obstacles without aberration correction.

Histological Observation of Islet Hemorrhage Induced by Diagnostic Ultrasound with Contrast Agent in Rat Pancreas

Contrast enhanced diagnostic ultrasound CEDUS has been shown to induce capillary hemorrhage in heart and kidney. This study characterized the capillary hemorrhage induced in rat pancreas. The pancreata of anesthetized hairless rats were accessed by laparotomy. A 1.5 MHz diagnostic ultrasound probe with 2.3 MPa peak rarefactional pressure amplitude and 1 s intermittent trigger was used to scan the pancreas, located at the focus (3.8 cm), through saline coupling. The probe was swept to expose the entire organ in 5 min during infusion of Definity® contrast agent at 10 µL/kg/min, and this was repeated in a reverse sweep. The entire pancreas was removed, spread flat for fixation and histological slides were prepared from the mid-plane. Slides were scored blind for islet hemorrhage over the entire area of the organ. Intra-islet microlesions were evident and hemorrhage surrounded many islets. The hemorrhage often impacted nearby acini, and expanded into inter-lobular septa. In CEDUS pancreata removed soon after scanning, 76.2±11.8% (n = 6) of islets had evidence of hemorrhage and/or islet microlesions compared to 1.1±2.5% (n = 5) for sham CEDUS (P<0.001). In pancreata removed after 4 hr, fibrin formation was detected by immunohistology in the hemorrhage and intra-islet microlesions. Diagnostic ultrasound with contrast agent induced substantial capillary hemorrhage in rat pancreas, concentrated particularly in the islets.

In vivo transcranial cavitation threshold detection during ultrasound-induced blood–brain barrier opening in mice

The in vivo cavitation response associated with blood–brain barrier (BBB) opening as induced by transcranial focused ultrasound (FUS) in conjunction with microbubbles was studied in order to better identify the underlying mechanism in its noninvasive application. A cylindrically focused hydrophone, confocal with the FUS transducer, was used as a passive cavitation detector (PCD) to identify the threshold of inertial cavitation (IC) in the presence of Definity® microbubbles (mean diameter range: 1.1–3.3 µm, Lantheus Medical Imaging, MA, USA). A vessel phantom was first used to determine the reliability of the PCD prior to in vivo use. A cerebral blood vessel was simulated by generating a cylindrical channel of 610 µm in diameter inside a polyacrylamide gel and by saturating its volume with microbubbles. The microbubbles were sonicated through an excised mouse skull. Second, the same PCD setup was employed for in vivo noninvasive (i.e. transdermal and transcranial) cavitation detection during BBB opening. After the intravenous administration of Definity® microbubbles, pulsed FUS was applied (frequency: 1.525 or 1.5 MHz, peak-rarefactional pressure: 0.15–0.60 MPa, duty cycle: 20%, PRF: 10 Hz, duration: 1 min with a 30 s interval) to the right hippocampus of twenty-six (n = 26) mice in vivo through intact scalp and skull. T1 and T2-weighted MR images were used to verify the BBB opening. A spectrogram was generated at each pressure in order to detect the IC onset and duration. The threshold of BBB opening was found to be at a 0.30 MPa peak-rarefactional pressure in vivo. Both the phantom and in vivo studies indicated that the IC pressure threshold had a peak-rarefactional amplitude of 0.45 MPa. This indicated that BBB opening may not require IC at or near the threshold. Histological analysis showed that BBB opening could be induced without any cellular damage at 0.30 and 0.45 MPa. In conclusion, the cavitation response could be detected without craniotomy in mice and IC may not be required for BBB opening at relatively low pressures.

Acoustic droplet vaporization for therapeutic and diagnostic applications

A phase shift droplet emulsion is introduced as an aid to unusual ultrasound (US) applications. The transpulmonary droplet emulsion (90% < 6 μm diameter) is made by mixing saline, bovine albumin and dodecafluoropentane. It has been observed that an acoustic pressure threshold exists, above which the droplets vaporize into bubbles approximately 25 times the original diameter. For frequencies between 1.5 and 8 MHz, the threshold decreases from 4.5 to 0.75 MPa peak rarefactional pressure. This paper presents preliminary results for droplet preparation and their evaporation as a function of applied acoustic pressure and frequency, as well as simulations of the lifetime of these gas bubbles based on gas diffusion. In vivo experiments were simulated by the evaporation of droplets in blood flowing under attenuating material. We propose that this agent might be useful for tissue occlusion in cancer treatment, as well as for phase aberration corrections in acoustic imaging.