Development of a burst wave lithotripsy system for noninvasive fragmentation of ureteroliths in pet cats.

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 min. Fourteen of 25 stones fragmented to < 1 mm within the 30 min. In a second experiment, burst wave lithotripsy was performed using a second transducer and peak ultrasound pressure of 8.0 MPa (n = 10) for up to 50 min. In the second experiment, 9 of 10 stones fragmented to < 1 mm within the 50 min. Across both experiments, an average of 73-97% of stone mass could be reduced to fragments < 1 mm. A third experiment found negligible injury with 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.

Treating Porcine Abscesses with Histotripsy: A Pilot Study.

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 washout 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.

A simulated model for fluid and tissue heating during pediatric laser lithotripsy.

BACKGROUND: Laser lithotripsy (LL) is a common modality for treatment of children and adolescents with nephrolithiasis. Recent introduction of higher-powered lasers may result in more efficacious "dusting" of urinary calculi. However, in vivo animal studies and computational simulations have demonstrated rapid and sustained rise of fluid temperatures with LL, possibly resulting in irreversible tissue damage. How fluid and tissue heating during LL vary with pediatric urinary tract development, however, is unknown. We hypothesize that kidneys of younger children will be more susceptible to changes in fluid temperature and therefore tissue damage than those of older children. METHODS: Computational simulations were developed for LL in children utilizing COMSOL Multiphysics finite-element modeling software. Simulation parameters were varied, including the child's age (3, 8, and 12 years), flow of irrigation fluid (gravity - 5 mL/min or continuous pressure flow - 40 mL/min), treatment location (renal pelvis, ureter, calyx), and power settings (5 W - 40 W). Using a simplified axisymmetric geometry to represent the collecting space, the model accounted for heat transfer via diffusion, convection, perfusion, and heat sourcing as well as tissue properties and blood flow of the urothelium and renal parenchyma. Laminar and heat-induced convection flow were simulated, assuming room-temperature ureteroscopic irrigation. Renal size was varied by age, based on normative values. The maximum fluid temperature after 60 s of simulated LL was captured. Thermal dose was calculated using the t43 equivalence of 240 min as a threshold for tissue damage, as was tissue volume at risk for irreversible cellular damage. RESULTS: Simulation with gravity flow irrigation revealed generation of thermal doses sufficient to cause tissue injury for all ages at 20 W and 40 W power settings. Higher temperatures were seen in younger ages across all power settings. Temperature increases were dampened with intermittent laser activity and continuous pressure flow irrigation. CONCLUSIONS: Smaller renal size is more susceptible to thermal changes induced by LL. However, power settings equal to or greater than 20 W can result in temperatures high enough for tissue damage at any age. Continuous pressure flow and intermittent laser activity may mitigate the potential thermal damage from high power LL.

An investigation of elastic waves producing stone fracture in burst wave lithotripsy.

Burst wave lithotripsy is a method to noninvasively fragment urinary stones by short pulses of focused ultrasound. In this study, physical mechanisms of stone fracture during burst wave lithotripsy were investigated. Photoelasticity imaging was used to visualize elastic wave propagation in model stones and compare results to numerical calculations. Epoxy and glass stone models were made into rectangular, cylindrical, or irregular geometries and exposed in a degassed water bath to focused ultrasound bursts at different frequencies. A high-speed camera was used to record images of the stone during exposure through a circular polariscope backlit by a monochromatic flash source. Imaging showed the development of periodic stresses in the stone body with a pattern dependent on frequency. These patterns were identified as guided wave modes in cylinders and plates, which formed standing waves upon reflection from the distal surfaces of the stone model, producing specific locations of stress concentration in the models. Measured phase velocities compared favorably to numerically calculated modes dependent on frequency and material. Artificial stones exposed to bursts produced cracks at positions anticipated by this mechanism. These results support guided wave generation and reflection as a mechanism of stone fracture in burst wave lithotripsy.

Defining Thermally Safe Laser Lithotripsy Power and Irrigation Parameters: In Vitro Model.

Introduction: High-power laser settings are commonly employed for stone dusting techniques. Previous in vitro and in vivo studies have demonstrated that a toxic thermal dose can result from treatment within a renal calix without adequate irrigation. Hence, both laser power and irrigation rate must be considered together to determine safe laser lithotripsy parameters. The objective of this in vitro study was to map parameter safety boundaries and create guidelines for selection of safe laser and irrigation settings. Methods: The experimental system consisted of in vitro models simulating ureter, renal calix, and renal pelvis placed in a water bath maintained at 37°C. Temperature was recorded during ureteroscopy with laser activation for 60 seconds. Trials were conducted at strategically selected power levels and irrigation rates. Thermal dose for each trial was calculated based on Sapareto and Dewey t43 methodology with thermal dose >120 equivalent minutes considered to result in thermal tissue injury. A parameter safety boundary was established by plotting the maximal safe power level for each irrigation rate. Results: The parameter safety boundary was found to be linear for each scenario with the renal pelvis able to tolerate the highest laser power and the renal calix the least power without injury. Conclusion: This study describes the methodology to determine parameter safety boundaries that can be used to guide proper selection of thermally safe laser settings and irrigation rates during ureteroscopy with laser lithotripsy. This work provides a framework to assess the effectiveness of various strategies to control and mitigate thermal dose.

PIXUL-ChIP: integrated high-throughput sample preparation and analytical platform for epigenetic studies.

Chromatin immunoprecipitation (ChIP) is the most widely used approach for identification of genome-associated proteins and their modifications. We have previously introduced a microplate-based ChIP platform, Matrix ChIP, where the entire ChIP procedure is done on the same plate without sample transfers. Compared to conventional ChIP protocols, the Matrix ChIP assay is faster and has increased throughput. However, even with microplate ChIP assays, sample preparation and chromatin fragmentation (which is required to map genomic locations) remains a major bottleneck. We have developed a novel technology (termed 'PIXUL') utilizing an array of ultrasound transducers for simultaneous shearing of samples in standard 96-well microplates. We integrated PIXUL with Matrix ChIP ('PIXUL-ChIP'), that allows for fast, reproducible, low-cost and high-throughput sample preparation and ChIP analysis of 96 samples (cell culture or tissues) in one day. Further, we demonstrated that chromatin prepared using PIXUL can be used in an existing ChIP-seq workflow. Thus, the high-throughput capacity of PIXUL-ChIP provides the means to carry out ChIP-qPCR or ChIP-seq experiments involving dozens of samples. Given the complexity of epigenetic processes, the use of PIXUL-ChIP will advance our understanding of these processes in health and disease, as well as facilitate screening of epigenetic drugs.

Simulation of Laser Lithotripsy-Induced Heating in the Urinary Tract.

PURPOSE: Holmium laser lithotripsy is a common modality used to fragment urinary stones during ureteroscopy. Laser energy deposited during activation produces heat and potentially causes thermal bioeffects. We aimed to characterize laser-induced heating through a computational simulation. MATERIALS AND METHODS: A finite-element model was developed and used to estimate temperature in the urinary tract. Axisymmetric models of laser lithotripsy in a renal calyx, the renal pelvis, and proximal ureter were created. Heat generation by laser and heat transfer were simulated under different laser powers between 5 and 40 W. Irrigation fluid flow was introduced at rates between 0 and 40 mL/min. The model was validated by comparison with previous in vitro temperature data in a test tube, then used to calculate heating and thermal dose in the three tissue models. RESULTS: Simulated temperature rises agreed well with most in vitro experimental measurements. In tissue models, temperature rises depended strongly on laser power and irrigation rate, and to a lesser extent on location. Injurious temperatures were reached for 5-40 W laser power without irrigation, >10 W with 5 mL/min irrigation, 40 W with 15 mL/min irrigation, and were not found at 40 mL/min irrigation. Tissue injury volumes up to 2.3 cm3 were calculated from thermal dose. CONCLUSIONS: The results suggest a numerical model can accurately simulate the thermal profile of laser lithotripsy. Laser heating is strongly dependent on parameters and may cause a substantial temperature rise in the fluid in the urinary tract and surrounding tissue under clinically relevant conditions.

Ultrasound-based cell sorting with microbubbles: A feasibility study.

The isolation and sorting of cells is an important process in research and hospital labs. Most large research and commercial labs incorporate fluorescently or magnetically labeled antibodies adherent to cell surface antigens for cell identification and separation. In this paper, a process is described that merges biochemical labeling with ultrasound-based separation. Instead of lasers and fluorophore tags, or magnets and magnetic particle tags, the technique uses ultrasound and microbubble tags. Streptavidin-labeled microbubbles were mixed with a human acute lymphoblastic leukemia cell line, CCL 119, conjugated with biotinylated anti-CD7 antibodies. Tagged cells were forced under ultrasound, and their displacement and velocity quantified. Differential displacement in a flow stream was quantified against erythrocytes, which showed almost no displacement under ultrasound. A model for the acoustic radiation force on the conjugated pairs compares favorably with observations. This technology may improve on current time-consuming and costly purification procedures.

Inactivation of Planktonic Escherichia coli by Focused 1-MHz Ultrasound Pulses with Shocks: Efficacy and Kinetics Upon Volume Scale-Up.

This study addresses inactivation of E. coli in either 5- or 10-mL volumes, which were 50- to 100-fold greater than used in an earlier study (Brayman et al. 2017). Cells were treated with 1-MHz pulsed high-intensity focused ultrasound (10 cycles, 2-kHz repetition frequency, +65/-12.8 MPa focal pressures). The surviving fraction was assessed by coliform assay, and inactivation demonstrated curvilinear kinetics. The reduction of surviving fraction to 50% required 2.5 or 6 min in 5- or 10-mL samples, respectively. Exposure of 5 mL for 20 min reduced the surviving fraction to ∼1%; a similar exposure of 10-mL samples reduced the surviving fraction to ∼10%. Surviving cells from 5-min exposures appeared normal under light microscopy, with minimal debris; after 20 min, debris dominated. Transmission electron microscopy images of insonated samples showed some undamaged cells, a few damaged but largely intact cells and comminuted debris. Cellular damage associated with substantive but incomplete levels of inactivation can be variable, ranging from membrane holes tens of nanometers in diameter to nearly complete comminution.

Caliceal Fluid Temperature During High-Power Holmium Laser Lithotripsy in an In Vivo Porcine Model.

INTRODUCTION: With increasing use of high-power laser settings for lithotripsy, the potential exists to induce thermal tissue damage. In vitro studies have demonstrated that temperature elevation sufficient to cause thermal tissue damage can occur with certain laser and irrigation settings. The objective of this pilot study was to measure caliceal fluid temperature during high-power laser lithotripsy in an in vivo porcine model. METHODS: Four female pigs (30-35 kg) were placed under general anesthesia and positioned supine. Retrograde ureteroscopy with entry into upper or middle calices was performed. Thermocouples were placed into the calix by open exposure and puncture of the kidney or retrograde alongside the ureteroscope. A 242 µm laser fiber was positioned in the center of the calix and activated (0.5 J, 80 Hz, 40 W) for 60 seconds with high, medium, or no irrigation delivered in each trial. Finite element simulations of laser-induced heating in a renal calix were also performed. RESULTS: Peak temperatures of 84.8°C, 63.9°C, and 43.6°C were recorded for no, medium, and high irrigation, respectively. Mean time to reach threshold of thermal injury (t43 of 120 minutes) was 12.7 and 17.8 seconds for no and medium irrigation. Thermal damage thresholds were not reached in high-irrigation trials. Numerical simulations revealed similar results with peak spatial average fluid temperatures of >100°C, 58.5°C, and 37.5°C during 60 seconds of laser activation for 0.1, 15, and 40 mL/minute irrigation, respectively. CONCLUSIONS: High-power holmium laser settings (40 W) can induce potentially injurious temperatures in the porcine in vivo model, particularly with slower irrigation rates. Characterization of thermal dose across a broader range of laser parameter settings is underway to map out the thermal safety envelope.

Inactivation of Planktonic Escherichia coli by Focused 2-MHz Ultrasound.

This study was motivated by the desire to develop a non-invasive means to treat abscesses, and represents the first steps toward that goal. Non-thermal, high-intensity focused ultrasound (HIFU) was used to inactivate Escherichia coli (∼1 × 109 cells/mL) in suspension. Cells were treated in 96-well culture plate wells using 1.95-MHz ultrasound and incident focal acoustic pressures as high as 16 MPa peak positive and 9.9 MPa peak negative (free field measurements). The surviving fraction was assessed by coliform culture and the alamarBlue assay. No biologically significant heating was associated with ultrasound exposure. Bacterial inactivation kinetics were well described by a half-life model, with a half-time of 1.2 min. At the highest exposure levels, a 2log inactivation was typically achieved within 10 min. The free field-equivalent peak negative acoustic pressure threshold for inactivation was ∼7 MPa. At the highest acoustic pressures used, inactivation efficacy was insensitive to reciprocal changes in pulse length and pulse repetition frequency at constant duty factor. Although treated volumes were very small, proof of principle was provided by these experiments.

An ultrasonic caliper device for measuring acoustic nonlinearity.

In medical and industrial ultrasound, it is often necessary to measure the acoustic properties of a material. A specific medical application requires measurements of sound speed, attenuation, and nonlinearity to characterize livers being evaluated for transplantation. For this application, a transmission-mode caliper device is proposed in which both transmit and receive transducers are directly coupled to a test sample, the propagation distance is measured with an indicator gage, and receive waveforms are recorded for analysis. In this configuration, accurate measurements of nonlinearity present particular challenges: diffraction effects can be considerable while nonlinear distortions over short distances typically remain small. To enable simple estimates of the nonlinearity coefficient from a quasi-linear approximation to the lossless Burgers' equation, the calipers utilize a large transmitter and plane waves are measured at distances of 15-50 mm. Waves at 667 kHz and pressures between 0.1 and 1 MPa were generated and measured in water at different distances; the nonlinearity coefficient of water was estimated from these measurements with a variability of approximately 10%. Ongoing efforts seek to test caliper performance in other media and improve accuracy via additional transducer calibrations.

Ultrasound stylet for non-image-guided ventricular catheterization.

OBJECT Urgent ventriculostomy placement can be a lifesaving procedure in the setting of hydrocephalus or elevated intracranial pressure. While external ventricular drain (EVD) insertion is common, there remains a high rate of suboptimal drain placement. Here, the authors seek to demonstrate the feasibility of an ultrasound-based guidance system that can be inserted into an existing EVD catheter to provide a linear ultrasound trace that guides the user toward the ventricle. METHODS The ultrasound stylet was constructed as a thin metal tube, with dimensions equivalent to standard catheter stylets, bearing a single-element, ceramic ultrasound transducer at the tip. Ultrasound backscatter signals from the porcine ventricle were processed by custom electronics to offer real-time information about ventricular location relative to the catheter. Data collected from the prototype device were compared with reference measurements obtained using standard clinical ultrasound imaging. RESULTS A study of porcine ventricular catheterization using the experimental device yielded a high rate of successful catheter placement after a single pass (10 of 12 trials), despite the small size of pig ventricles and the lack of prior instruction on porcine ventricular architecture. A characteristic double-peak signal was identified, which originated from ultrasound reflections off of the near and far ventricular walls. Ventricular dimensions, as obtained from the width between peaks, were in agreement with standard ultrasound reference measurements (p < 0.05). Furthermore, linear ultrasound backscatter data permitted in situ measurement of the stylet distance to the ventricular wall (p < 0.05), which assisted in catheter guidance. CONCLUSIONS The authors have demonstrated the ability of the prototype ultrasound stylet to guide ventricular access in the porcine brain. The alternative design of the device makes it potentially easy to integrate into the standard workflow for bedside EVD placement. The availability of a fast, easy-to-use, inexpensive guidance system can play a role in reducing the complication rate for EVD placement.

Simple circumcision device: proof of concept for a single-visit, adjustable device to facilitate safe adult male circumcision.

OBJECTIVE: To conduct a proof-of-concept study to determine the potential utility of a novel, adjustable single-visit, disposable device to facilitate rapid adult circumcision. DESIGN: Prospective pilot trial of a novel surgical device. SETTING: Tertiary care Veterans Administration medical center. PATIENT(S): Five adult males. INTERVENTION(S): Circumcisions performed by junior trainees using an adjustable, single-size surgical-assist device constructed by the University of Washington Applied Physics Laboratory. MAIN OUTCOME MEASURE(S): The attending surgeon and trainees completed standardized forms after each procedure to assess technical problems and ease of use. Follow-up visits were scheduled to evaluate adverse events, postoperative pain, cosmetic outcomes, and participant satisfaction at 3, 8, 30, and 90 days postoperatively. RESULT(S): The average operative time was 16.4 minutes. All cases were performed with local anesthesia, and no case required electrocautery or conversion to standard surgery. At the postoperative day 3 visit, all subjects were happy with their results and would recommend the procedure to another patient. One participant had a minor wound separation noted at the 30-day visit that resolved during follow-up. There were no wound infections, hematomas, or other adverse events. CONCLUSION(S): This proof-of-study suggests that the Simple Circumcision Device may facilitate delivery of safe adult male circumcision services.

A mechanistic analysis of stone fracture in lithotripsy.

In vitro experiments and an elastic wave model were used to analyze how stress is induced in kidney stones by lithotripsy and to test the roles of individual mechanisms-spallation, squeezing, and cavitation. Cylindrical U30 cement stones were treated in an HM-3-style lithotripter. Baffles were used to block specific waves responsible for spallation or squeezing. Stones with and without surface cracks added to simulate cavitation damage were tested in glycerol (a cavitation suppressive medium). Each case was simulated using the elasticity equations for an isotropic medium. The calculated location of maximum stress compared well with the experimental observations of where stones fractured in two pieces. Higher calculated maximum tensile stress correlated with fewer shock waves required for fracture. The highest calculated tensile stresses resulted from shear waves initiated at the proximal corners and strengthened along the side surfaces of the stone by the liquid-borne lithotripter shock wave. Peak tensile stress was in the distal end of the stone where fracture occurred. Reflection of the longitudinal wave from the distal face of the stone--spallation-produced lower stresses. Surface cracks accelerated fragmentation when created near the location where the maximum stress was predicted.