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One of the challenges of transcutaneous high-intensity focused ultrasound (HIFU) therapies, especially ones relying heavily on shock formation, such as boiling histotripsy (BH), is the loss of focusing from aberration induced by the heterogeneities of the body wall. Here, a methodology to execute aberration correction in vivo is proposed. A custom BH system consisting of a 1.5-MHz phased array of 256 elements connected to a Verasonics V1 system is used in pulse/echo mode on a porcine model under general anesthesia. Estimation of the time shifts needed to correct for aberration in the liver and kidney is done by maximizing the value of the coherence factor on the acquired backscattered signals. As this process requires multiple pulse/echo sequences on a moving target to converge to a solution, tracking is also implemented to ensure that the same target is used between each iteration. The method was validated by comparing the acoustic power needed to generate a boiling bubble at one target with aberration correction and at another target within a 5-mm radius without aberration correction. Results show that the aberration correction effectively lowers the acoustic power required to reach boiling by up to 45%, confirming that it indeed restored formation of the nonlinear shock front at the focus.

Supplementary Materials and Methods. Description of additional methods and procedures used in the study.

Supplementary Figure Legends. Legends for Supplementary Figures S1-S2.

Supplementary Table S2. Estimated regression coefficients from equation 2.

Supplementary Figure S1. A multispectral image of an in vivo murine subcutaneous pancreatic tumor treated with pHIFU with varying levels of peak-rarefactional pressure.

Pancreatic cancer is characterized by extensive stromal desmoplasia, which decreases blood perfusion and impedes chemotherapy delivery. Breaking the stromal barrier could both increase perfusion and permeabilize the tumor, enhancing chemotherapy penetration. Mechanical disruption of the stroma can be achieved using ultrasound-induced bubble activity—cavitation. Cavitation is also known to result in microstreaming and could have the added benefit of actively enhancing diffusion into the tumors. Here, we report the ability to enhance chemotherapeutic drug doxorubicin penetration using ultrasound-induced cavitation in a genetically engineered mouse model (KPC mouse) of pancreatic ductal adenocarcinoma. To induce localized inertial cavitation in pancreatic tumors, pulsed high-intensity focused ultrasound (pHIFU) was used either during or before doxorubicin administration to elucidate the mechanisms of enhanced drug delivery (active vs. passive drug diffusion). For both types, the pHIFU exposures that were associated with high cavitation activity resulted in disruption of the highly fibrotic stromal matrix and enhanced the normalized doxorubicin concentration by up to 4.5-fold compared with controls. Furthermore, normalized doxorubicin concentration was associated with the cavitation metrics (P < 0.01), indicating that high and sustained cavitation results in increased chemotherapy penetration. No significant difference between the outcomes of the two types, that is, doxorubicin infusion during or after pHIFU treatment, was observed, suggesting that passive diffusion into previously permeabilized tissue is the major mechanism for the increase in drug concentration. Together, the data indicate that pHIFU treatment of pancreatic tumors when resulting in high and sustained cavitation can efficiently enhance chemotherapy delivery to pancreatic tumors. Cancer Res; 75(18); 3738–46. ©2015 AACR.

Ultrasound and microbubbles are useful for both diagnostic imaging and targeted drug delivery, making them ideal conduits for theranostic interventions. Recent reports have indicated the preclinical success of microbubble cavitation for enhancement of chemotherapy in abdominal tumors; however, there have been limited studies and variable efficacy in clinical implementation of this technique. This is likely because in contrast to the high pressures and long cycle lengths seen in successful preclinical work, current clinical implementation of microbubble cavitation for drug delivery generally involves low acoustic pressures and short cycle lengths to fit within clinical guidelines. To translate the preclinical parameter space to clinical adoption, a relevant safety study in a healthy large animal is required. Therefore, the purpose of this work was to evaluate the safety of ultrasound cavitation treatment (USCTx) in a healthy porcine model using a modified Philips EPIQ with S5-1 as the focused source. We performed USCTx on eight healthy pigs and monitored health over the course of 1 wk. We then performed an acute study of USCTx to evaluate immediate tissue damage. Contrast-enhanced ultrasound exams were performed before and after each treatment to investigate perfusion changes within the treated areas, and blood and urine were evaluated for liver damage biomarkers. We illustrate, through quantitative analysis of contrast-enhanced ultrasound data, blood and urine analyses and histology, that this technique and the parameter space considered are safe within the time frame evaluated. With its safety confirmed using a clinical-grade ultrasound scanner and contrast agent, USCTx could be easily translated into clinical trials for improvement of chemotherapy delivery. This represents the first safety study assessing the bio-effects of microbubble cavitation from relevant ultrasound parameters in a large animal model.

Background High‐intensity focused ultrasound (HIFU) has been in clinical use for a variety of solid tumors and cancers. Accurate and reliable calibration is in a great need for clinical applications. An extracorporeal clinical HIFU system applied for the investigational device exemption (IDE) to the Food and Drug Administration (FDA) so that evaluation of its characteristics, performance, and safety was required. Methods The acoustic pressure and power output was characterized by a fiber optic probe and a radiation force balance, respectively, with the electrical power up to 2000 W. An in situ acoustic energy was established as the clinical protocol at the electrical power up to 500 W. Temperature elevation inside the tissue sample was measured by a thermocouple array. Generated lesion volume at different in situ acoustic energies and pathological examination of the lesions was evaluated ex vivo. Results Acoustic pressure mapping showed the insignificant presence of side/grating lobes and pre‐ or post‐focal peaks (≤−12 dB). Although distorted acoustic pressure waveform was found in the free field, the nonlinearity was reduced significantly after the beam propagating through tissue samples (i.e., the second harmonic of −11.8 dB at 500 W). Temperature elevation was

Background: Upper urinary tract stones are increasingly prevalent in pet cats and are difficult to manage. Surgical procedures to address obstructing ureteroliths have short- and long-term complications, and medical therapies (e.g. fluid diuresis and smooth muscle relaxants) are infrequently effective. Burst wave lithotripsy is a non-invasive, ultrasound-guided, handheld focused ultrasound technology to disintegrate urinary stones, which is now undergoing human clinical trials in awake unanesthetized subjects. Results: In this study, we designed and performed in vitro testing of a modified burst wave lithotripsy system to noninvasively fragment stones in cats. The design accounted for differences in anatomic scale, acoustic window, skin-to-stone depth, and stone size. Prototypes were fabricated and tested in a benchtop model using 35 natural calcium oxalate monohydrate stones from cats. In an initial experiment, burst wave lithotripsy was performed using peak ultrasound pressures of 7.3 (n = 10), 8.0 (n = 5), or 8.9 MPa (n =10) for up to 30 minutes. Fourteen of 25 stones fragmented to in vivo exposure of kidneys and ureters in a porcine animal model. Conclusions: These data support further evaluation of burst wave lithotripsy as a noninvasive intervention for obstructing ureteroliths in cats.

Peyronie’s disease affects penile mechanics, but published research lacks biomechanical characterization of affected tunica albuginea. This work aims to establish mechanical testing methodology and characterize pathological tissue mechanics of Peyronie’s disease. Tunica albuginea was obtained from patients (n = 5) undergoing reconstructive surgery for Peyronie’s disease, sectioned into test specimens (n = 12), stored frozen at −20 °C, and imaged with micro-computed tomography (µCT). A tensile testing protocol was developed based on similar soft tissues. Correlation of mechanical summary variables (force, displacement, stiffness, work, Young’s modulus, ultimate tensile stress, strain at ultimate tensile stress, and toughness) and µCT features were assessed with linear regression. Specimens empirically grouped into hard or soft stress–strain behavior were compared using a Student’s t-test. Surface strain and failure patterns were described qualitatively. Specimens displayed high inter- and intra-subject variability. Mineralization volume was not correlated with mechanical parameters. Empirically hard tissue had higher ultimate tensile stress. Failure mechanisms and strain patterns differed between mineralized and non-mineralized specimens. Size, shape, and quantity of mineralization may be more important in determining Peyronie’s disease plaque behavior than presence of mineralization alone, and single summary variables like modulus may not fully describe mechanical behavior.

Infected abscesses are walled-off collections of pus and bacteria. They are a common sequela of complications in the setting of surgery, trauma, systemic infections, and other disease states. Current treatment is typically limited to antibiotics with long-term catheter drainage, or surgical wash-out when inaccessible to percutaneous drainage or unresponsive to initial care efforts. Antibiotic resistance is also a growing concern. Although bacteria can develop drug resistance, they remain susceptible to thermal and mechanical damage. In particular, short pulses of focused ultrasound (i.e., histotripsy) generate mechanical damage through localized cavitation, representing a potential new paradigm for treating abscesses non-invasively, without the need for long-term catheterization and antibiotics. In this pilot study, boiling and cavitation histotripsy treatments were applied to subcutaneous and intramuscular abscesses developed in a novel porcine model. Ultrasound imaging was used to evaluate abscess maturity, for treatment monitoring and assessment of post-treatment outcomes. Disinfection was quantified by counting bacteria colonies from samples aspirated before and after treatment. Histopathological evaluation of the abscesses was performed to identify changes resulting from histotripsy treatment and potential collateral damage. Cavitation histotripsy was more successful in reducing the bacterial load while having a smaller treatment volume compared with boiling histotripsy. The results of this pilot study suggest focused ultrasound may lead to a technology for in situ treatment of acoustically accessible abscesses.

Boiling histotripsy (BH) is a focused ultrasound technology that uses millisecond-long pulses with shock fronts to induce mechanical tissue ablation. The pulsing scheme and mechanisms of BH differ from those of cavitation cloud histotripsy, which was previously developed for benign prostatic hyperplasia. The goal of the work described here was to evaluate the feasibility of using BH to ablate fresh ex vivo human prostate tissue as a proof of principle for developing BH for prostate applications. Fresh human prostate samples (N = 24) were obtained via rapid autopsy (24 h after death, institutional review board exempt). Samples were analyzed using shear wave elastography to ensure that mechanical properties of autopsy tissue were clinically representative. Samples were exposed to BH using 10- or 1-ms pulses with 1% duty cycle under real-time B-mode and Doppler imaging. Volumetric lesions were created by sonicating 1-4 rectangular planes spaced 1 mm apart, containing a grid of foci spaced 1-2 mm apart. Tissue then was evaluated grossly and histologically, and the lesion content was analyzed using transmission electron microscopy and scanning electron microscopy. Observed shear wave elastography characterization of ex vivo prostate tissue (37.9 ± 22.2 kPa) was within the typical range observed clinically. During BH, hyperechoic regions were visualized at the focus on B-mode, and BH-induced bubbles were also detected using power Doppler. As treatment progressed, hypoechoic regions of tissue appeared, suggesting successful tissue fractionation. BH treatment was twofold faster using shorter pulses (1 ms vs. 10 ms). Histological analysis revealed lesions containing completely homogenized cell debris, consistent with histotripsy-induced mechanical ablation. It was therefore determined that BH is feasible in fresh ex vivo human prostate tissue producing desired mechanical ablation. The study supports further work aimed at translating BH technology as a clinical option for prostate ablation.

Despite the growing incidence of urinary stones, the fundamental interventions for urinary stones have remained the same for several decades. However, technical innovations in ultrasound have enabled new strategies for stone management. Our team aims to employ these methods to establish a new paradigm to treat stones in an office or clinic rather than a surgical suite. Technologies include new imaging methods to detect and characterize stones, burst wave lithotripsy to fragment stones using focused ultrasound rather than shock waves, and ultrasonic propulsion to promote clearance of fragments, all using a single ultrasound platform. This presentation will describe the principles and development of these technologies, as well as the fruitful partnerships between teams of basic scientists, engineers, physicians, and industry that have enabled rapid development of these concepts to successful clinical trials. [Work supported by NIDDK P01 DK043881 and the Focused Ultrasound Foundation.]

Pulsed focused ultrasound (pFUS) exposures utilizing short, nonlinearly distorted pulses at low duty cycle have been shown to enhance drug and gene delivery to targeted tissue through inertial cavitation activity. Passive cavitation detection (PCD) and mapping of broadband emissions are current conventional methods to monitor and quantify cavitation but provide limited spatial resolution. Here, plane-wave Doppler imaging was used with PCD to quantify pFUS-induced cavitation in ex vivo bovine tissues and in vivo surgically exposed porcine liver, kidney, and pancreas. A 1.5 MHz FUS transducer (aperture 75 mm, F-number 0.75) was used to deliver 60 pulses (duration 1 ms, 0.1% duty cycle, focal pressure p+ = 70i–110 MPa, p− = 13–20 MPa). A coaxially mounted ATL P7-4 ultrasound imaging probe was used for PCD during the FUS pulse, and Doppler and B-mode sequences. Disrupted tissue areas were collected for histology and compared to Doppler power images. Maximum Doppler power was found to correlate to broadband noise level for each FUS pulse. The Doppler power map integrated over the exposure was observed to correlate spatially with tissue disruption area from histology, which thus represents a promising real-time metric for quantifying cavitation activity induced by pFUS exposures. [Work supported by NIH R01CA154451, R01EB025187, and R01EB23910.]

Tissue-mimicking gels provide a cost-effective medium to optimize histotripsy treatment parameters with immediate feedback. Agarose and polyacrylamide gels are often used to evaluate treatment outcomes as they mimic the acoustic properties and stiffness of a variety of soft tissues, but they do not exhibit high toughness, a characteristic of fibrous connective tissue. To mimic pathologic fibrous tissue found in benign prostate hyperplasia (BPH) and other diseases that are potentially treatable with histotripsy, an optically transparent hydrogel with high toughness was developed that is a hybrid of polyacrylamide and alginate. The stiffness was established using shear wave elastography (SWE) and indentometry techniques and was found to be representative of human BPH ex vivo prostate tissue. Different phantom compositions and excised ex vivo BPH tissue samples were treated with a 700-kHz histotripsy transducer at different pulse repetition frequencies. Post-treatment, the hybrid gels and the tissue samples exhibited differential reduction in stiffness as measured by SWE. On B-mode ultrasound, partially treated areas were present as hyperechoic zones and fully liquified areas as hypoechoic zones. Phase contrast microscopy of the gel samples revealed liquefaction in regions consistent with the target lesion dimensions and correlated to findings identified in tissue samples via histology. The dose required to achieve liquefaction in the hybrid gel was similar to what has been observed in ex vivo tissue and greater than that of agarose of comparable or higher Young's modulus by a factor10. These results indicate that the developed hydrogels closely mimic elasticities found in BPH prostate ex vivo tissue and have a similar response to histotripsy treatment, thus making them a useful cost-effective alternative for developing and evaluating different treatment protocols.

Liquid atomization and fountain formation by focused ultrasound was first published by Wood and Loomis [1]. Since then, the cavitation-wave hypothesis emerged to explain atomization in a fountain, which states atomization arises from a combination of surface capillary waves and the collapse of cavitation bubbles. More recently, high intensity focused ultrasound (HIFU) has been shown to fractionate tissue through either pulsed-cavitation or millisecond boiling histotripsy therapies; however it is unclear how millimeter-size boiling bubbles or cavitation bubble clouds fractionate tissue into submicron-size fragments. The objective of this work is to test the hypothesis experimentally that atomization and fountain formation occurs similarly in liquids and tissues and results in tissue erosion. A 2-MHz HIFU transducer operating at peak in situ pressures of 50 MPa and −11 MPa (linear intensity = 14,000 W/cm(2)) was focused at the interface between a liquid or tissue and air. A high-speed camera was used to monitor atomization and fountain formation in water, ethanol, glycerol, bovine liver, and porcine blood clots. The in situ linear intensity threshold for consistent atomization in one 10-ms pulse increased in the order: ethanol (180 W/cm(2)) < blood clot (250 W/cm(2)) < water (350 W/cm(2)) < liver (6200 W/cm(2)); glycerol did not atomize. Average jet velocities for the initial spray at the maximum acoustic intensity were similar for all materials and on the order of 20 m/s. The tissue erosion rate of liver approached saturation at around 300 10-ms pulses repeated at 1 Hz, which had an average erosion volume of 25.7±10.9 mm(3). While tissue atomization and fountain formation does not completely mimic what is observed in liquids, atomization provides a plausible explanation of how tissue is fractionated in millisecond boiling and possibly even cavitation cloud histotrispy therapies.

Boiling histotripsy (BH) is a High Intensity Focused Ultrasound (HIFU) method for precise mechanical disintegration of target tissue using millisecond-long pulses containing shocks. BH treatments with real-time ultrasound (US) guidance allowed by BH-generated bubbles were previously demonstrated ex vivo and in vivo in exposed porcine liver and small animals. Here, the feasibility of US-guided transabdominal and partially transcostal BH ablation of kidney and liver in an acute in vivo swine model was evaluated for 6 animals. BH parameters were: 1.5 MHz frequency, 5–30 pulses of 1–10 ms duration per focus, 1% duty cycle, peak acoustic powers 0.9–3.8 kW, sonication foci spaced 1–1.5 mm apart in a rectangular grid with 5–15 mm linear dimensions. In kidneys, well-demarcated volumetric BH lesions were generated without respiratory gating and renal medulla and collecting system were more resistant to BH than cortex. The treatment was accelerated 10-fold by using shorter BH pulses of larger peak power without affecting the quality of tissue fractionation. In liver, respiratory motion and aberrations from subcutaneous fat affected the treatment but increasing the peak power provided successful lesion generation. These data indicate BH is a promising technology for transabdominal and transcostal mechanical ablation of tumors in kidney and liver.

Introduction: Burst wave lithotripsy is an experimental technology to noninvasively fragment kidney stones with focused bursts of ultrasound (US). This study evaluated the safety and effectiveness of specific lithotripsy parameters in a porcine model of nephrolithiasis. Methods: A 6- to 7-mm human kidney stone was surgically implanted in each kidney of three pigs. A burst wave lithotripsy US transducer with an inline US imager was coupled to the flank and the lithotripter focus was aligned with the stone. Each stone was exposed to burst wave lithotripsy at 6.5 to 7 MPa focal pressure for 30 minutes under real-time image guidance. After treatment, the kidneys were removed for gross, histologic, and MRI assessment. Stone fragments were retrieved from the kidney to determine the mass comminuted to pieces

Inertial cavitation induced by pulses of nonlinearly distorted focused ultrasound (FUS) at moderate intensity can result in mild mechanical disruption of tissue, short of its complete mechanical disintegration—histotripsy. This effect can be used to enhance diffusion of subsequently systemically administered drugs or biologics. Previously, single-element passive cavitation detection (PCD) of broadband noise emissions was successfully used to quantify tissue disruption and enhancement in drug concentration. This metric, however, has limitations: minimal spatial resolution and challenges with calibration when used at large depths in acoustically variable tissue. To address this, we developed a combination of passive and active Doppler-based methods that relied on relative, rather than absolute signal metrics. Specifically, destructive cavitation behaviors were previously linked to substantial motion of the bubbles during the FUS pulse due to acoustic radiation force and shock scattering. This was reflected in the backscattered FUS harmonics as Doppler shift and measurable from PCD. The bubbles were observed to dissolve within milliseconds following each FUS pulse, thus their distribution could be visualized as an area of rapid change via fast plane wave Doppler ensemble following the FUS pulse. The Doppler power distribution was spatially correlated with the area of tissue disruption in the in vivo experiments. [Work supported by NIH R01CA154451, R01EB025187, and R01EB23910.]

Abscesses are walled-off collections of infected fluids that often develop as complications in the setting of surgery and trauma. Treatment is usually limited to percutaneous catheterization with a course of antibiotics. As an alternative to current treatment strategies, a histotripsy approach was developed and tested in a novel porcine animal model. The goal of this article is to use advanced ultrasound imaging modes to extract sonographic features associated with the progression of abscess development in a porcine model. Intramuscular or subcutaneous injections of a bi-microbial bacteria mixture plus dextran particles as an irritant led to identifiable abscesses over a 2 to 3 wk period. Selected abscesses were imaged at least weekly with B-mode, 3-D B-mode, shear-wave elastography and plane-wave Doppler imaging. Mature abscesses were characterized by a well-defined core of varying echogenicity surrounded by a hypoechoic capsule that was highly vascularized on Doppler imaging. 3-D imaging demonstrated the natural history of abscess morphology, with the abscess becoming less complex in shape and increasing in volume. Furthermore, shear-wave elastography demonstrated variations in stiffness as phlegmon becomes abscess and then liquefies, over time. These ultrasound features potentially provide biomarkers to aid in selection of treatment strategies for abscesses.

Large volume soft tissue hematomas are a serious clinical problem, which, if untreated, can have severe consequences. Current treatments are associated with significant pain and discomfort. It has been demonstrated in an in vitro bovine hematoma model that pulsed high intensity focused ultrasound (HIFU) ablation termed histotripsy can be used to rapidly and non-invasively liquefy the hematoma through localized bubble activity, enabling fine-needle aspiration. The goals of this study were to evaluate the efficiency and speed of volumetric histotripsy liquefaction using a large in vitro human hematoma model. Large human hematoma phantoms (85 cc) were formed by recalcifying blood anticoagulated with citrate phosphate dextrose (CPD) - saline-adenine-glucose-mannitol solution (SAGM). Typical boiling histotripsy (BH) pulses (10-ms or 2-ms) or hybrid histotripsy pulses using higher amplitude and shorter pulses (0.4-ms) were delivered at 1% duty cycle while continuously translating the HIFU focus location. Histotripsy exposures were performed under ultrasound guidance with a 1.5 MHz transducer (8 cm aperture, F# = 0.75). The volume of liquefied lesions was determined by ultrasound imaging and gross inspection. Untreated hematoma samples and samples of the liquefied lesions aspirated using a fine needle were analyzed cytologically and ultrastructurally with scanning electron microscopy (SEM). All exposures resulted in uniform liquid-filled voids with sharp edges; liquefaction speed was higher for exposures with shorter pulses and higher shock amplitudes at the focus (up to 0.32, 0.68, and 2.62 mL/min for 10, 2, and 0.4 ms pulses, respectively). Cytological and ultrastructural observations revealed completely homogenized blood cells and fibrin fragments in the lysate. Most of the fibrin fragments were less than 20 μm in length, but a number of fragments were up to 150 μm. The lysate with residual debris of that size would potentially be amenable to fine-needle aspiration without risk for needle clogging in clinical implementation.

Burst wave lithotripsy (BWL) is a technology under clinical investigation for noninvasive fragmentation of urinary stones. Under certain ranges of ultrasound exposure parameters, this technology can cause cavitation in tissue leading to renal injury. This study sought to measure the focal pressure amplitude needed to cause cavitation in vivo and determine its consistency in native tissue, in an implanted stone model, and under different exposure parameters. The kidneys of eight pigs were exposed to transcutaneous BWL ultrasound pulses. In each kidney, two locations were targeted: the renal sinus and the kidney parenchyma. Each was exposed for 5 minutes at a set pressure level and parameters, and cavitation was detected using an active cavitation imaging method based on power Doppler ultrasound. The threshold was determined by incrementing the pressure amplitude up or down after each 5-minute interval until cavitation occurred/subsided. The pressure thresholds were remeasured post-surgery targeting an implanted stone or collecting space (in sham). The presence of a stone or sham surgery did not significantly impact the threshold for tissue cavitation. Targeting parenchyma instead of kidney collecting space and lowering the ultrasound pulse repetition frequency both resulted in an increased pressure threshold for cavitation.

Abscesses are walled-off collections of infected fluids that often develop as complications in the setting of surgery and trauma. Abscess care depends on size, location, composition and complexity, among other patient factors. The goal of this work is to describe, using the latest ultrasound imaging technologies, the progression of abscess development in a porcine animal model. Intramuscular or subcutaneous injections of bacteria plus dextran particles as an irritant led to identifiable abscesses over a 2- to 3-week period. The abscesses were imaged at least weekly with B-mode, 3D B-mode, shear-wave elastography (SWE) and color flow imaging. Mature abscesses were characterized by a well-defined core of varying echogenicity surrounded by a hypoechoic capsule that was highly vascularized on Doppler imaging. Size and shape changes during development were quantified with 3D imaging. With SWE, the lesion stiffness varied interiorly and generally decreased over time. These ultrasound features potentially provide biomarkers to facilitate improved selection of treatment strategies for abscesses.

Infected abscesses are walled-off collections of pus and bacteria. They are a common sequela of complications in the setting of surgery, trauma, systemic infections, and other disease states. Current treatment is typically limited to antibiotics with long-term catheter drainage, or surgical wash-out when inaccessible to percutaneous drainage or unresponsive to initial care efforts. Antibiotic resistance is also a growing concern. Although bacteria can develop drug resistance, they remain susceptible to thermal and mechanical damage. In particular, short pulses of focused ultrasound (i.e., histotripsy) generate mechanical damage through localized cavitation, representing a potential new paradigm for treating abscesses non-invasively, without the need for long-term catheterization and antibiotics. In this pilot study, boiling and cavitation histotripsy treatments were applied to subcutaneous and intramuscular abscesses developed in a novel porcine model. Ultrasound imaging was used to evaluate abscess maturity, for treatment monitoring and assessment of post-treatment outcomes. Disinfection was quantified by counting bacteria colonies from samples aspirated before and after treatment. Histopathological evaluation of the abscesses was performed to identify changes resulting from histotripsy treatment and potential collateral damage. Cavitation histotripsy was more successful in reducing the bacterial load while having a smaller treatment volume compared with boiling histotripsy. The results of this pilot study suggest focused ultrasound may lead to a technology for in situ treatment of acoustically accessible abscesses.

Abscesses are walled-off collections of infected fluids most often treated with percutaneous drains placed under CT guidance. Complications such as clogged drains or secondary infections require rehospitalization and wound management. Histotripsy treatment has the potential to eliminate the need for long term catheterization and antibiotics. The progression of abscess development has yet to be fully described. The objective of this study was to use the latest advances in non-contrast ultrasound technologies to characterize abscess development in a porcine animal model. Intramuscular or subcutaneous injections of bacteria plus dextran particles as an irritant led to identifiable abscesses over a 2- to 3-week period. Ultrasound imaging was performed at least weekly, in some cases with a 3D tracking device that provided quantifiable size and shape measurements. Abscess progression was also measured with a plane-wave Doppler mode providing increased sensitivity to low-velocity flows, while abscess stiffness was quantified using shear wave elastography. Most of the mature abscesses were characterized by a rounded core of varying echogenicity surrounded by a hypoechoic capsule that was highly vascularized on Doppler imaging. A treatable abscess was defined by its hypervascular rim and avascular core. Stiffness varied within the abscess but generally decreased over time. Abscess echogenicity, shape, stiffness and vascularity potentially provide features to identify lesions suitable for treatment.

Abscesses are walled-off collections of infected fluids containing pus and bacteria. They are often treated with percutaneous drainage in which a drainage catheter may be sutured in place for up to several weeks. Complications such as clogged drains or secondary infections require rehospitalization and wound management. Bacteria are susceptible to mechanical damage, and thus we hypothesize that histotripsy may be a potential new paradigm for treating abscesses noninvasively, without the need for long term catheterization and antibiotics. We developed a porcine animal model that recapitulates some of the features of human abscesses (including size and loculations). Boiling and cavitation histotripsy treatments were applied to subcutaneous and intramuscular abscesses in this porcine model. Ultrasound imaging was used to evaluate abscess maturity, for treatment monitoring and assessment of post-treatment outcomes. Disinfection was quantified by counting bacteria colonies from samples aspirated before and after treatment. Histopathological evaluation of the abscesses was performed to identify changes resulting from histotripsy treatment and potential collateral damage. The results of this pilot study suggest focused ultrasound may lead to a technology for in situ treatment of acoustically accessible abscesses.

Burst wave lithotripsy (BWL) is a new non-invasive method for stone comminution using bursts of sub-megahertz ultrasound. A porcine model of urolithiasis and techniques to implement BWL treatment has been developed to evaluate its effectiveness and acute safety. Six human calcium oxalate monohydrate stones (6–7 mm) were hydrated, weighed, and surgically implanted into the kidneys of three pigs. Transcutaneous stone treatments were performed with a BWL transducer coupled to the skin via an external water bath. Stone targeting and treatment monitoring were performed with a co-aligned ultrasound imaging probe. Treatment exposures were applied in three 10-minute intervals for each stone. If sustained cavitation in the parenchyma was observed by ultrasound imaging feedback, treatment was paused and the pressure amplitude was decreased for the remaining time. Peak negative focal pressures between 6.5 and 7 MPa were applied for all treatments. After treatment, stone fragments were removed from the kidneys. At least 50% of each stone was reduced to

Peyronie's disease affects penile mechanics, but published research lacks biomechanical characterization of affected tunica albuginea. This work aims to establish mechanical testing methodology and characterize pathological tissue mechanics of Peyronie's disease. Tunica albuginea was obtained from patients (n = 5) undergoing reconstructive surgery for Peyronie's disease, sectioned into test specimens (n = 12), stored frozen at -20 °C, and imaged with micro-computed tomography (µCT). A tensile testing protocol was developed based on similar soft tissues. Correlation of mechanical summary variables (force, displacement, stiffness, work, Young's modulus, ultimate tensile stress, strain at ultimate tensile stress, and toughness) and µCT features were assessed with linear regression. Specimens empirically grouped into hard or soft stress-strain behavior were compared using a Student's t-test. Surface strain and failure patterns were described qualitatively. Specimens displayed high inter- and intra-subject variability. Mineralization volume was not correlated with mechanical parameters. Empirically hard tissue had higher ultimate tensile stress. Failure mechanisms and strain patterns differed between mineralized and non-mineralized specimens. Size, shape, and quantity of mineralization may be more important in determining Peyronie's disease plaque behavior than presence of mineralization alone, and single summary variables like modulus may not fully describe mechanical behavior.

The accuracy of current burn triage techniques has remained between 50-70%. Accordingly, there is a significant clinical need for the quantitative and accurate assessment of partial-thickness burn injuries. Porcine skin represents the closest animal model to human skin, and is often used in surgical skin grafting procedures. In this study, we used a standardized in vivo porcine burn model to obtain terahertz (THz) point-spectroscopy measurements from burns with various severities. We then extracted two reflection hyperspectral parameters, namely spectral area under the curve between approximately 0.1 and 0.9 THz (−10 dB bandwidth in each spectrum), and spectral slope, to characterize each burn. Using a linear combination of these two parameters, we accurately classified deep partial- and superficial partial-thickness burns (p = 0.0159), compared to vimentin immunohistochemistry as the gold standard for burn depth determination.

Boiling histotripsy (BH) is an experimental focused ultrasound technique that produces non-thermal mechanical ablation. We evaluated the feasibility, short-term histologic effects, and the resulting acute inflammatory response to BH ablation of renal cell carcinoma (RCC) in the Eker rat. Genotyped Eker rats were monitored for de novo RCCs with serial ultrasound (US) imaging. When tumors were ≥8 mm, rats underwent ultrasound-guided extracorporeal ablation of the tumor with BH, a pulsed focused US technique that produces non-thermal mechanical ablation of targeted tissues, or a sham US procedure. Treatments targeted approximately 50% of the largest RCC with a margin of normal kidney. BH treated rats were euthanized at 1 (n=4) or 48 (n=4) hours, and sham subjects (n=4) at 48 hours. Circulating plasma cytokine levels were assessed with multiplex assays prior to and at 0.25, 1, 4, 24 and 48 hours following treatment. Kidneys were collected and processed for histologic assessment, immunohistochemistry and intrarenal cytokine concentration measurements. For statistical analysis Student’s t-test was used. US-guided BH treatment was successful in all animals, producing hypoechoic regions within the targeted volume consistent with BH treatment effect. Grossly, regions of homogenized tissue were apparent with evidence of focal intra-parenchymal hemorrhage. Histologically, BH produced a sharply demarcated region of homogenized tumor and non-tumor tissue containing acellular debris. BH treatment was associated with significantly increased relative concentration of plasma TNF vs. sham treatment (p

BackgroundAbscess formation is a host defense mechanism to contain the spread of infection. Abscesses can affect any part of the body and are commonsequelaeto complications of trauma, surgery, systemic infections and other disease states. Most models for abscesses are in small animals. Pursuant to the goal of developing more advanced treatments for abscesses, we sought to develop a large animal model which would reasonably mimic a fluid-filled human abscess.MethodsDomestic swine were inoculated with a bimicrobial mixture ofBacteroides fragilis(B. fragilis) andEscherichia coli(E. coli) supplemented with an irritant (dextran). Inoculations were performed under ultrasound guidance in the muscle, subcutaneously or intradermally within the same animal. Fourteen days after inoculation, lesions were imaged with ultrasound, resected and prepared for histological evaluation.ResultsInjection of bimicrobial (aerobic and anaerobic) bacterial mixtures at multiple sites in a pig produced multiple lesions with histological features similar to encapsulated and multiloculated/multichamber abscesses often observed clinically in humans. Important salient features include the formation of a connective tissue capsule surrounding histologically nearly amorphous pus.ConclusionsThis paper provides the first description of a pig model for multiloculated abscesses. This animal model could potentially enable the evaluation of new technologies to replace or augment the current standard of care (image-guided percutaneous abscess drainage with antibiotics).

Abscesses are infected walled-off collections of pus and bacteria. They can affect any part of the body. Current treatment is typically limited to antibiotics, catheter drainage and hospitalization, or surgical wash-out. Although bacteria can develop drug resistance, they remain susceptible to thermal and mechanical damage. Histotripsy generates localized cavitation without heating and represents a potential new noninvasive treatment modality. Successful histotripsy treatments were demonstrated with gram-negative E-coli in both in vitro and in vivo studies [Brayman et al., UMB 44, 1996–2008 (2018); Matula et al., UMB 47, 603–619 (2020)]. Interestingly, preliminary data showed that boiling histotripsy was less effective than cavitation histotripsy in killing bacteria. In this talk, we will describe more in-depth studies that compare cavitation histotripsy (low number of cycles, high-pulse repetition rate) with boiling histotripsy (high number of cycles, low-pulse repetition rate) on inactivation rates of microbes. [Work funded in part by NIH NIBIB No. R01EB019365.]

Pancreatic ductal adenocarcinomas are characterized by a complex and robust tumor microenvironment (TME) consisting of fibrotic tissue, excessive levels of hyaluronan (HA), and immune cells. We utilized quantitative multi-parametric magnetic resonance imaging (mp-MRI) methods at 14 Tesla in a genetically engineered KPC (KrasLSL-G12D/+, Trp53LSL-R172H/+, Cre) mouse model to assess the complex TME in advanced stages of tumor development. The whole tumor, excluding cystic areas, was selected as the region of interest for data analysis and subsequent statistical analysis. Pearson correlation was used for statistical inference. There was a significant correlation between tumor volume and T2 (r = −0.66), magnetization transfer ratio (MTR) (r = 0.60), apparent diffusion coefficient (ADC) (r = 0.48), and Glycosaminoglycan-chemical exchange saturation transfer (GagCEST) (r = 0.51). A subset of mice was randomly selected for histological analysis. There were positive correlations between tumor volume and fibrosis (0.92), and HA (r = 0.76); GagCEST and HA (r = 0.81); and MTR and CD31 (r = 0.48). We found a negative correlation between ADC low-b (perfusion) and Ki67 (r = −0.82). Strong correlations between mp-MRI and histology results suggest that mp-MRI can be used as a non-invasive tool to monitor the tumor microenvironment.

Diabetes is associated with lower limb co-morbidities, including ulceration and subsequent amputation. As a systemic disease, diabetes affects the microstructure of soft tissues, and material microstructural changes are known to affect the macroscale mechanics. However, the associations between diabetes-related disruptions to essential microstructural components and mechanical changes in plantar skin with diabetes has not been thoroughly characterized. Plantar skin specimens were collected from four diabetic and eight non-diabetic donors at six plantar locations (hallux; first, third, and fifth metatarsals; lateral midfoot; calcaneus) from matched pairs. Mechanical testing was performed on fresh frozen specimens from one foot, and histomorphological measurement and biochemical quantification were performed on specimens from the other foot. Mechanical (compressive and shear moduli and viscoelastic slopes) and biochemical/histological (total quantity of collagen and elastin; dermal and epidermal thickness) parameters were correlated using linear mixed effects regression. There were no significant differences by disease state. Skin thicknesses were positively correlated with initial compression modulus and all three shear moduli. The final compressive modulus was significantly lower at the third metatarsal than the fifth metatarsal, lateral midfoot, and calcaneus, while the final shear modulus was significantly higher at the calcaneus than at the hallux, first, and third metatarsals. Epidermal thickness was significantly higher at the calcaneus compared to all other locations. While differences were not significant by disease state, the strong differences by locations and significant but weak correlations between skin thickness and mechanics can inform future research to understand the mechanism of ulcer formation in the diabetic foot.

Acoustic radiation forces can trap and manipulate objects in three-dimensional space (3D). The state of the art in acoustic trapping has demonstrated the ability to move small or lightweight objects using single or multiple transducers. For certain medical applications, acoustic manipulation can be utilized to control foreign objects in the body, such as kidney stones. However, difficulties from these applications include the ability to control large solid objects, transmitting robust acoustic beams through the skin, and the use of a single source due to limited acoustic window in the body. The goal of this work was to use a 1.5 MHz, focused, multi-element array to synthesize specific acoustic beams to trap, levitate, and steer kidney stone models (glass spheres) in a water bath and urinary bladders of live pigs. Stable acoustic traps were investigated numerically and experimentally in water bath to design a protocol for the 3D manipulation strategies used invivo . Three programmed path were used to levitate and manipulate glass spheres in live pigs. The motion was recorded using a camera and a synchronized ultrasound imaging probe. Deviation from the intended paths was on average