Cell Maps for Artificial Intelligence: AI-Ready Maps of Human Cell Architecture from Disease-Relevant Cell Lines.

This article describes the Cell Maps for Artificial Intelligence (CM4AI) project and its goals, methods, standards, current datasets, software tools , status, and future directions. CM4AI is the Functional Genomics Data Generation Project in the U.S. National Institute of Health's (NIH) Bridge2AI program. Its overarching mission is to produce ethical, AI-ready datasets of cell architecture, inferred from multimodal data collected for human cell lines, to enable transformative biomedical AI research.

Development of a contrast-enhanced ultrasound guided high intensity focused ultrasound system for coagulation of liver parenchyma.

BACKGROUND: The liver is the most common organ injured in blunt abdominal trauma and makes up roughly 5% of all trauma admissions. Current treatments are invasive and resource-intensive, which may delay care. We aim to develop and validate a contrast-enhanced ultrasound (CEUS)guided noninvasive tool to treat liver lacerations at the bedside. METHODS: Two 1.8 MHz high-intensity focused ultrasound (HIFU) elements were coupled to a C1-6 diagnostic ultrasound probe and a Logiq E10 scanner (GE HealthCare) utilizing a custom enclosure for co-registered imaging and ablation. A phantom was created from polyacrylamide gel combined with thermochromic ink whose color changes above biological ablative temperatures (60 °C). The HIFU wave was focused approximately 0.5 cm below the surface utilizing a 50% duty cycle generating 11.9 MPa for 20, 30, 40, 50, and 60s. Experiments were repeated on ex vivo chicken livers in a water bath. Finally, the livers of 4 live swine underwent up to 6 CEUS-guided treatments using parameters optimized from in vitro work. RESULTS: Treatment of the phantom between 20-60s, produced ablation sizes from 0.016 to 0.4 cm 3 . The relationship between time and size was exponential (R 2 = 0.992). Ablation areas were also well visualized on with ultrasound imaging. The ex vivo liver ablation size at 20s was 0.37 cm 3 , at 30s was 0.66 cm 3 , and at 100 s was 5.0 cm 3 . For the in-vivo swine experiments, the average ablation area measured 2.0x0.75 cm with a maximum of 3.5x1.5 cm. CEUS was utilized with the contrast agent Definity (Lantheus) for identification of lacerations as well as immediate post operative evaluation of therapy. CONCLUSION: These experiments demonstrate the feasibility of CEUS guided transdermal HIFU ablation and the time-dependent size of ablation. This work warrants future investigations into using ultrasound to detect active bleeding and HIFU to coagulate grade III and IV liver laceration. STUDY TYPE: Therapeutic/care management.

Comparison of hypospadias phenotype pixel segmentation to GMS score.

BACKGROUND: Hypospadias phenotype assessment determines if the anatomy is favorable for reconstruction. Glans-Urethral Meatus-Shaft (GMS) has been adopted in an effort to standardize hypospadias classification. While extremely subjective, GMS has been widely used to classify the severity of the phenotype to predict surgical outcomes. The use of digital image analysis has proven to be feasible and prior efforts by our team have demonstrated that machine learning algorithms can emulate an expert's assessment of the phenotype. Nonetheless, the creation of these image recognition algorithms is highly subjective. In order to reduce a subjective input in the evaluation of the phenotype, we propose a novel approach to analyze the anatomy using digital image pixel analysis and to compare the results using the GMS score. Our hypothesis is that pixel cluster segmentation can discriminate between favorable and unfavorable anatomy. OBJECTIVE: To evaluate whether image segmentation and digital pixel analysis are able to analyze favorable vs unfavorable hypospadias anatomy in a less subjective manner than GMS score. METHODS: A total of 148 patients with different types of hypospadias were classified by 1 of 5 independent experts following the GMS score into "favorable" (GG), "moderately favorable" (GM) and "unfavorable" (GP) glans. From there, 592 images were generated using digital image segmentation. 584 were included for final analysis due to certain images being excluded for poor image quality or inadequate capture of target anatomy. For each image, the region of interest was segmented separately by two evaluators into "glans," "urethral plate," "foreskin" and "periurethral plate". The values obtained for each segmented region using machine-learning statistical pixel k-means cluster analysis were analyzed and compared to the GMS score given to that image using an ANOVA analysis. RESULTS: Analysis of image segmentation demonstrated that k-means pixel cluster analysis discriminated "favorable" vs "unfavorable" urethral plates. There was a significant difference between scores when comparing the GG and GM groups (p = 0.03) and GG and GP groups (p = 0.05). Pixel cluster analysis could not discriminate between "moderately favorable" and "unfavorable" urethral plates. CONCLUSIONS: Through our analysis, we found significant pairwise difference for different tissue qualities. Digital image segmentation and statistical k-means cluster analysis can discriminate anatomical features in a similar way to the GMS score. Future research can target discerning between different tissue qualities in an effort to predict surgical outcomes for hypospadias repair.

Cavitation-induced pressure saturation: a mechanism governing bubble nucleation density in histotripsy.

Objective.Histotripsy is a noninvasive focused ultrasound therapy that mechanically disintegrates tissue by acoustic cavitation clouds. In this study, we investigate a mechanism limiting the density of bubbles that can nucleate during a histotripsy pulse. In this mechanism, the pressure generated by the initial bubble expansion effectively negates the incident pressure in the vicinity of the bubble. From this effect, the immediately adjacent tissue is prevented from experiencing the transient tension to nucleate bubbles. Approach.A Keller-Miksis-type single-bubble model was employed to evaluate the dependency of this effect on ultrasound pressure amplitude and frequency, viscoelastic medium properties, bubble nucleus size, and transducer geometric focusing. This model was further combined with a spatial propagation model to predict the peak negative pressure field as a function of position from a cavitating bubble.Main results. The single-bubble model showed the peak negative pressure near the bubble surface is limited to the inertial cavitation threshold. The predicted bubble density increased with increasing frequency, tissue viscosity, and transducer focusing angle. The simulated results were consistent with the trends observed experimentally in prior studies, including changes in density with ultrasound frequency and transducerF-number.Significance.The efficacy of the therapy is dependent on several factors, including the density of bubbles nucleated within the cavitation cloud formed at the focus. These results provide insight into controlling the density of nucleated bubbles during histotripsy and the therapeutic efficacy.

Validation of Urostomy Parastomal Herniation Incisional Prevention Strategies.

OBJECTIVE: To evaluate simulated parastomal herniation forces in in vitro abdominal fascial models. Our group previously illustrated how incision type may play a consequential role in bowel herniation force generated across an incision using several abdominal fascia models. We sought to (1) Confirm findings in fresh human tissue, (2) Assess correlation between herniation force and incision size, and (3) Determine whether incision type impacts drainage in a simulated ex vivo ileal conduit. MATERIALS AND METHODS: Axial tension force (N) of herniation was measured using our previously published protocol, pulling a Foley catheter balloon 3.8 cm diameter affixed to a dynamometer through silicone/fascial incisions ranging 3-5.8 cm. We simulated ileal conduits using bovine small intestine with stoma matured through human fascia using 3.0 cm linear or cruciate incisions. The conduit's caudal end was catheterized and filled at 20 mL/min. Drainage was measured by pad weight change. Two-sided α < 0.05 was used to reject the null hypothesis. RESULTS: Mean (±SD) herniation forces in fresh human fascia varied significantly across linear longitudinal, linear transverse, and cruciate incisions (20.9 ± 3.7, 23.3 ± 8.8, and 8.9 ± 3.8 N, respectively [P = .011]). Fresh human fascial linear incisions 3 cm in diameter had a herniation force of 22.1 ± 6.3 vs 3.5 ± 0.7 N for 5.8 cm incisions when herniating a 3.8 cm balloon (P = .002). All observations were similar in silicone. In simulated ileal conduit, mean drainage: 70.8 ± 3.6 vs 82.1 ± 9.7 mL (linear vs cruciate) after 100 mL instilled, respectively (P = .05). CONCLUSION: This ex vivo study further suggests incision type has predictable influence on herniation force. These data support standardization of urostomy construction techniques and evaluating the clinical impact of stomal maturation techniques on parastomal hernia rates.

Effects of pulse repetition frequency on bubble cloud characteristics and ablation in single-cycle histotripsy.

Objective. Histotripsy is a cavitation-based ultrasound ablation method in development for multiple clinical applications. This work investigates the effects of pulse repetition frequency (PRF) on bubble cloud characteristics and ablative capabilities for histotripsy using single-cycle pulsing methods.Approach.Bubble clouds produced by a 500 kHz histotripsy system at PRFs from 0.1 to 1000 Hz were visualized using high-speed optical imaging in 1% agarose tissue phantoms at peak negative pressures,p-, of 2-36 MPa.Main results.Results showed a decrease in the cavitation cloud threshold with increasing PRF, ranging from 26.7 ± 0.5 MPa at 0.1 Hz to 15.0 ± 1.9 MPa at 1000 Hz. Bubble cloud analysis showed cavitation clouds generated at low PRFs (0.1-1 Hz) were characterized by consistently dense bubble clouds (41.7 ± 2.8 bubbles mm-2at 0.1 Hz), that closely matched regions of the focus above the histotripsy intrinsic threshold. Bubble clouds formed at higher PRFs measured lower cloud densities (23.1 ± 4.0 bubbles mm-2at 1000 Hz), with the lowest density measured for 10 Hz (8.8 ± 4.1 bubbles mm-2). Furthermore, higher PRFs showed increased pulse-to-pulse correlation, characteristic of cavitation memory effects; however, bubble clouds still filled the entire volume of the focus due to their initial density and enhanced bubble expansion from the restimulation of residual nuclei at the higher PRFs. Histotripsy ablation assessed through lesion analysis in red blood cell (RBC) phantoms showed higher PRFs generated lesions with lower adherence to the initial focal region compared to low PRF ablations; however, no trend of decreasing ablation efficiency with PRF was observed, with similar efficiencies observed for all the PRFs tested in this study.Significance.Notably, this result is different than what has previously been shown for shock-scattering histotripsy, which has shown decreased ablation efficiencies at higher PRFs. Overall, this study demonstrates the essential effects of PRF on single-cycle histotripsy procedures that should be considered to help guide future histotripsy pulsing strategies.

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.

Comparative Study of Histotripsy Pulse Parameters Used to Inactivate Escherichia coli in Suspension.

OBJECTIVE: Bacterial loads can be effectively reduced using cavitation-mediated focused ultrasound, or histotripsy. In this study, gram-negative bacteria (Escherichia coli) in suspension were used as model bacteria to evaluate the effectiveness of two regimens of histotripsy treatments: cavitation histotripsy (CH) and boiling histotripsy (BH). METHODS: Ten-milliliter volumes of Escherichia coli were treated at different negative focal pressure amplitudes and over time periods up to 40 min. Cavitation activity was characterized with coaxial passive cavitation detection (PCD) and synchronized plane wave B-mode imaging. RESULTS: CH treatments exhibited a threshold behavior that was consistent with PCD metrics of cavitation. Above the threshold, bacterial inactivation followed a monotonically increasing log-linear relationship that indicated an exponential inactivation rate. BH exhibited no threshold, but instead followed a different monotonically increasing inactivation rate. Inactivation rates were larger for BH at or below the CH threshold, and larger for CH substantially above the threshold. CH studies performed at different pulse lengths at the same duty cycle had similar inactivation rates, suggesting that at any given pressure amplitude, the "on time" was the most important variable for inactivating E. coli. The maximum inactivation was produced by CH at the highest pressure amplitudes used, leading to a log reduction >4.2 for a 40 min treatment. CONCLUSION: The results of this study suggest that both CH and BH can be used to inactivate E. coli in suspension, with the optimal regimen depending on the attainable peak negative focal pressure at the target.

Assessment of Urostomy Parastomal Herniation Forces Using Incisional Prevention Strategies with an Abdominal Fascia Model.

Background: Approximately 10 000 patients undergo cystectomy/ileal conduit annually in the USA, of whom over 70% subsequently develop a parastomal hernia (PSH). Still, no well-established "best" practice for stoma creation to prevent a PSH exists. Objective: To measure the relationship between incision size/type/material and axial tension force (ATF) as a surrogate for herniation force, using several models to mimic abdominal fascia. Design setting and participants: Abdominal fascia models included silicone membrane, ex vivo porcine, and embalmed human cadaveric fascia. A dynamometer pulled a Foley catheter (20 mm/min) with the balloon inflated to 125% incision (linear, cruciate, and circular) diameter using a motorized positioning system. The maximum ATF before herniation was recorded. The study was repeated in unused silicone/tissue for suture reinforcement. We evaluated silicone, ex vivo porcine, and human abdominal fascia. Intervention: Incision sizes (1-3 cm) in 0.5-cm increments were evaluated in silicone. A 3-cm incision was used in porcine/human tissue. Outcome measurements and statistical analysis: ATF for herniation was recorded/compared across incision types/sizes using Mann-Whitney U and Kruskal-Wallis tests as appropriate, with α = 0.05. Results and limitations: Linear incision ATF was significantly greater than cruciate and circular incisions. A cruciate incision had significantly greater ATF than a circular incision. In cadaveric tissue, incisions were significantly greater for linear (34.5 ± 12.8 N) versus cruciate (15.3 ± 2.9 N, p = 0.004) and for cruciate versus circular (p = 0.023) incisions. Results were similar in ex vivo porcine fascia and silicone. Reinforcement with a suture significantly increased ATF in all materials/incision sizes/types. The ex vivo nature is this study's main limitation. Conclusions: This study suggests that urostomy fascial incision type may influence ATF required for herniation. Linear incisions may be preferable. Urostomy reinforcement may significantly increase ATF required for a PSH. These data may help establish best practices for PSH risk reduction. Patient summary: The results of this study illustrate that urostomy fascia incision type may influence the force required to create a parastomal hernia. Linear incisions may be preferable.

Phase holograms for the three-dimensional patterning of unconstrained microparticles.

Acoustic radiation forces can remotely manipulate particles. Forces from a standing wave field align microscale particles along the nodal or anti-nodal locations of the field to form three-dimensional (3D) patterns. These patterns can be used to form 3D microstructures for tissue engineering applications. However, standing wave generation requires more than one transducer or a reflector, which is challenging to implement in vivo. Here, a method is developed and validated to manipulate microspheres using a travelling wave from a single transducer. Diffraction theory and an iterative angular spectrum approach are employed to design phase holograms to shape the acoustic field. The field replicates a standing wave and aligns polyethylene microspheres in water, which are analogous to cells in vivo, at pressure nodes. Using Gor'kov potential to calculate the radiation forces on the microspheres, axial forces are minimized, and transverse forces are maximized to create stable particle patterns. Pressure fields from the phase holograms and resulting particle aggregation patterns match predictions with a feature similarity index > 0.92, where 1 is a perfect match. The resulting radiation forces are comparable to those produced from a standing wave, which suggests opportunities for in vivo implementation of cell patterning toward tissue engineering applications.

Comprehensive multidisciplinary phenotyping of patients with hypospadias. A pilot study.

INTRODUCTION: Hypospadias is an abnormal formation of the urethra, ventral skin, and corporal bodies. Location of the urethral meatus has historically been the phenotypic landmark that defines hypospadias. Nonetheless, classifications following location of the urethral meatus fail to consistently predict outcomes and have no correlation with the genotype. Description of the urethral plate is very subjective and difficult to reproduce. We hypothesize that the use of digital pixel cluster analysis and correlation to histological analysis can provide a novel method to describe the phenotype of patients with hypospadias. METHODS: A standardized hypospadias phenotyping protocol was developed. 1. Digital images of the anomaly, 2. Anthropometric assessment of penile dimensions (penile length, urethral plate length and width, glans width, ventral curvature), 3. Classification using the GMS score, 4. Tissue sampling (foreskin, glans, urethral plate, periurethral ventral skin) and H&E analysis by a blinded pathologist. A k-means colorimetric pixel cluster analysis was performed following the same anatomical landmark distribution as the histology samples. Analysis was performed using MATLAB v R2021b 9.11.0.1769968. RESULTS: A total of 24 patients prospectively enrolled with a standard protocol. Mean age at surgery was 16.25 months Urethral meatus was distal shaft in 7 patients, 8 coronal, 4 glanular, 3 midshaft, 2 penoscrotal. Average GMS score was 7.14 (±1.58). Average glans size was 15.71 mm (±2.33) and urethral plate width 5.57 mm (±2.06). Eleven patients underwent Thiersch-Duplay repair, 7 TIP, 5 MAGPI, and 1 a first stage preputial flap. Mean follow-up was 14.25 months ( ± 3.7 months). Two (8.3%) postoperative complications (1 urethrocutaneous fistula and 1 ventral skin wound dehiscence) were reported in the study period. Eleven (52.3%) patients with histological analysis had an abnormal pathology report. Of those, 6 (54%) had reported abnormal lymphocyte infiltration interpreted as chronic inflammation at the urethral plate. The second most common finding was hyperkeratosis visualized in the urethral plate in 4 (36.3%) and one with reported fibrosis in the urethral plate. K-means pixel analysis demonstrated a k1 mean of 64.2 for reported urethral plate inflammation vs 53.1 for non-reported urethral plate inflammation (p = 0.002) CONCLUSIONS: Current phenotyping of hypospadias using only anthropometric variables can be expanded including histological and pixel analysis correlation. Pixel clustering has a potential for a priori prediction of urethral plate quality beyond the current subjective assessment. A larger cohort will allow identification of possible predictive associations that might impact intraoperative decision-making and surgical outcomes.

DNA release from plant tissue using focused ultrasound extraction (FUSE).

Premise: Sample preparation in genomics is a critical step that is often overlooked in molecular workflows and impacts the success of downstream genetic applications. This study explores the use of a recently developed focused ultrasound extraction (FUSE) technique to enable the rapid release of DNA from plant tissues for genetic analysis. Methods: FUSE generates a dense acoustic cavitation bubble cloud that pulverizes targeted tissue into acellular debris. This technique was applied to leaf samples of American chestnut (Castanea dentata), tulip poplar (Liriodendron tulipifera), red maple (Acer rubrum), and chestnut oak (Quercus montana). Results: We observed that FUSE can extract high quantities of DNA in 9-15 min, compared to the 30 min required for control DNA extraction methods. FUSE extracted DNA quantities of 24.33 ± 6.51 ng/mg and 35.32 ± 9.21 ng/mg from American chestnut and red maple, respectively, while control methods yielded 6.22 ± 0.87 ng/mg and 11.51 ± 1.95 ng/mg, respectively. The quality of the DNA released by FUSE allowed for successful amplification and next-generation sequencing. Discussion: These results indicate that FUSE can improve DNA extraction efficiency for leaf tissues. Continued development of this technology aims to adapt to field-deployable systems to increase the cataloging of genetic biodiversity, particularly in low-resource biodiversity hotspots.

Initial Assessment of Boiling Histotripsy for Mechanical Ablation of Ex Vivo Human Prostate Tissue.

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.

Quantitative Assessment of Boiling Histotripsy Progression Based on Color Doppler Measurements.

Boiling histotripsy (BH) is a mechanical tissue liquefaction method that uses sequences of millisecond-long high intensity focused ultrasound (HIFU) pulses with shock fronts. The BH treatment generates bubbles that move within the sonicated volume due to acoustic radiation force. Since the velocity of the bubbles and tissue debris is expected to depend on the lesion size and liquefaction completeness, it could provide a quantitative metric of the treatment progression. In this study, the motion of bubble remnants and tissue debris immediately following BH pulses was investigated using high-pulse repetition frequency (PRF) plane-wave color Doppler ultrasound in ex vivo myocardium tissue. A 256-element 1.5 MHz spiral HIFU array with a coaxially integrated ultrasound imaging probe (ATL P4-2) produced 10 ms BH pulses to form volumetric lesions with electronic beam steering. Prior to performing volumetric BH treatments, the motion of intact myocardium tissue and anticoagulated bovine blood following isolated BH pulses was assessed as two limiting cases. In the liquid blood the velocity of BH-induced streaming at the focus reached over 200 cm/s, whereas the intact tissue was observed to move toward the HIFU array consistent with elastic rebound of tissue. Over the course of volumetric BH treatments tissue motion at the focus locations was dependent on the axial size of the forming lesion relative to the corresponding size of the HIFU focal area. For axially small lesions, the maximum velocity after the BH pulse was directed toward the HIFU transducer and monotonically increased over time from about 20-100 cm/s as liquefaction progressed, then saturated when tissue was fully liquefied. For larger lesions obtained by merging multiple smaller lesions in the axial direction, the high-speed streaming away from the HIFU transducer was observed at the point of full liquefaction. Based on these observations, the maximum directional velocity and its location along the HIFU propagation axis were proposed and evaluated as candidate metrics of BH treatment completeness.

Development of Tough Hydrogel Phantoms to Mimic Fibrous Tissue for Focused Ultrasound Therapies.

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 factor >10. 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.

Improving Burst Wave Lithotripsy Effectiveness for Small Stones and Fragments by Increasing Frequency: Theoretical Modeling and Ex Vivo Study.

Introduction and Objective: In clinical trial NCT03873259, a 2.6-mm lower pole stone was treated transcutaneously and ex vivo with 390-kHz burst wave lithotripsy (BWL) for 40 minutes and failed to break. The stone was subsequently fragmented with 650-kHz BWL after a 4-minute exposure. This study investigated how to fragment small stones and why varying the BWL frequency may more effectively fragment stones to dust. Methods: A linear elastic theoretical model was used to calculate the stress created inside stones from shock wave lithotripsy (SWL) and different BWL frequencies mimicking the stone's size, shape, lamellar structure, and composition. To test model predictions about the impact of BWL frequency, matched pairs of stones (1-5 mm) were treated at (1) 390 kHz, (2) 830 kHz, and (3) 390 kHz followed by 830 kHz. The mass of fragments >1 and 2 mm was measured over 10 minutes of exposure. Results: The linear elastic model predicts that the maximum principal stress inside a stone increases to more than 5.5 times the pressure applied by the ultrasound wave as frequency is increased, regardless of the composition tested. The threshold frequency for stress amplification is proportionate to the wave speed divided by the stone diameter. Thus, smaller stones may be likely to fragment at a higher frequency, but not at a lower frequency below a limit. Unlike with SWL, this amplification in BWL occurs consistently with spherical and irregularly shaped stones. In water tank experiments, stones smaller than the threshold size broke fastest at high frequency (p = 0.0003), whereas larger stones broke equally well to submillimeter dust at high, low, or mixed frequencies. Conclusions: For small stones and fragments, increasing frequency of BWL may produce amplified stress in the stone causing the stone to break. Using the strategies outlined here, stones of all sizes may be turned to dust efficiently with BWL.

Fragmentation of Stones by Burst Wave Lithotripsy in the First 19 Humans.

PURPOSE: We report stone comminution in the first 19 human subjects by burst wave lithotripsy (BWL), which is the transcutaneous application of focused, cyclic ultrasound pulses. MATERIALS AND METHODS: This was a prospective multi-institutional feasibility study recruiting subjects undergoing clinical ureteroscopy (URS) for at least 1 stone ≤12 mm as measured on computerized tomography. During the planned URS, either before or after ureteroscope insertion, BWL was administered with a handheld transducer, and any stone fragmentation and tissue injury were observed. Up to 3 stones per subject were targeted, each for a maximum of 10 minutes. The primary effectiveness outcome was the volume percent comminution of the stone into fragments ≤2 mm. The primary safety outcome was the independent, blinded visual scoring of tissue injury from the URS video. RESULTS: Overall, median stone comminution was 90% (IQR 20, 100) of stone volume with 21 of 23 (91%) stones fragmented. Complete fragmentation (all fragments ≤2 mm) within 10 minutes of BWL occurred in 9 of 23 stones (39%). Of the 6 least comminuted stones, likely causative factors for decreased effectiveness included stones that were larger than the BWL beamwidth, smaller than the BWL wavelength or the introduction of air bubbles from the ureteroscope. Mild reddening of the papilla and hematuria emanating from the papilla were observed ureteroscopically. CONCLUSIONS: The first study of BWL in human subjects resulted in a median of 90% comminution of the total stone volume into fragments ≤2 mm within 10 minutes of BWL exposure with only mild tissue injury.

Non-Invasive Monitoring of Increased Fibrotic Tissue and Hyaluronan Deposition in the Tumor Microenvironment in the Advanced Stages of Pancreatic Ductal Adenocarcinoma.

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.

Mechanical characterization of fibrotic and mineralized tissue in Peyronie's disease.

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.