Peak Negative Pressure Research Articles

Efficacy optimization of low frequency microbubble-mediated sonoporation as a drug delivery platform to cancer cells

Ultrasound insonation of microbubbles can be used to form pores in cell membranes and facilitate the local trans-membrane transport of drugs and genes. An important factor in efficient delivery is the size of the delivered target compared to the generated membrane pores. Large molecule delivery remains a challenge, and can affect the resulting therapeutic outcomes. To facilitate large molecule delivery, large pores need to be formed. While ultrasound typically uses megahertz frequencies, it was recently shown that when microbubbles are excited at a frequency of 250 kHz (an order of magnitude below the resonance frequency of these agents), their oscillations are significantly enhanced as compared to the megahertz range. Here, to promote the delivery of large molecules, we suggest using this low frequency and inducing large pore formation through the high-amplitude oscillations of microbubbles. We assessed the impact of low frequency microbubble-mediated sonoporation on breast cancer cell uptake by optimizing the delivery of 4 fluorescent molecules ranging from 1.2 to 70 kDa in size. The optimal ultrasound peak negative pressure was found to be 500 kPa. Increasing the pressure did not enhance the fraction of fluorescent cells, and in fact reduced cell viability. For the smaller molecule sizes, 1.2 kDa and 4 kDa, the groups treated with an ultrasound pressure of 500 kPa and MB resulted in a fraction of 58% and 29% of fluorescent cells respectively, whereas delivery of 20 kDa and 70 kDa molecules yielded 10% and 5%, respectively. These findings suggest that low-frequency (e.g., 250 kHz) insonation of microbubbles results in high amplitude oscillation in vitro that increase the uptake of large molecules. Successful ultrasound-mediated molecule delivery requires the careful selection of insonation parameters to maximize the therapeutic effect by increasing cell uptake.

Enhanced Sonothrombolysis Induced by High-Intensity Focused Acoustic Vortex.

High-intensity focused ultrasound (HIFU) thrombolysis provides a targeted and non-invasive therapy for thrombosis-related diseases. Rapid thrombolysis and restoration of blood flow are vital to reduce the disability and death rate. The objective of this study was to explore the feasibility of using a high-intensity focused acoustic vortex (HIFAV) to enhance sonothrombolysis. The in vitro clots were treated with HIFU with a peak negative pressure (PNP) of 2.86 MPa (HIFU A) or 3.27 MPa (HIFU B) or HIFAV with a PNP of 2.14 MPa. The results revealed that HIFAV thrombolysis could achieve a significantly higher efficiency than HIFU (HIFAV: 65.4%, HIFU A: 24.1%, HIFU B: 31.6%, p < 0.01), even at a lower intensity. The average size of the debris particles generated in HIFAV thrombolysis was similar to that in HIFU. Additionally, the cavitation activities were found to be more intense in HIFAV thrombolysis. Although the efficiency of HIFAV thrombolysis was higher when the pulse repetition frequency increased from 100 to 500 Hz (41.4% vs. 65.4%, p < 0.05), it decreased when the PRF reached 1000 Hz (29.9%). Lastly, it was found that increasing the duty cycle from 5% to 15% led to a higher efficiency in HIFAV thrombolysis (40.3% vs. 75.2%, p < 0.001). This study illustrated that HIFAV provided enhanced thrombolysis and that its efficiency could be further increased by optimizing the ultrasound parameters.

Investigation of the aerodynamic characteristics of horizontal axis wind turbine using an active flow control method via boundary layer suction

The boundary layer suction (BLS) has the merit of effectively suppressing the formation and development of separation vortices by removing low-momentum fluid occurs on the leading-edge of airfoil suction surface. The present paper numerically investigates the impacts of BLS on the aerodynamic performance of a horizontal axis wind turbine (HAWT). First, the dynamic mode decomposition method is adopted to gain an insight into the improvement mechanism of BLS control by analyzing the flow patterns of a two-dimensional S809 airfoil. Subsequently, the BLS is arranged on a NREL Phase VI reference turbine to study the power increment and flow field characteristics, and the effects of different working angles αs and momentum coefficients Cμ are investigated at different tip speed ratios (TSRs). The results indicate that the use of BLS control can increase the pressure difference between the suction surface and the pressure surface, raise the peak negative pressure at the leading edge and shift the airfoil flow separation point backwards, and thus effectively enhance the stability of the flow field. Furthermore, the BLS control can significantly improve the power extraction at low to medium TSR if this active device is appropriately implemented on the HAWT.

Synergistic enhancement of cell death by triple combination therapy of docetaxel, ultrasound and microbubbles, and radiotherapy on PC3 a prostate cancer cell line

The application of ultrasound and microbubbles (USMB) has been shown to enhance both chemotherapy and radiotherapy. This study investigated the potential of triple combination therapy comprised of USMB, docetaxel (Taxotere: TXT) chemotherapy and XRT to enhance treatment efficacy. Prostate cancer (PC3) cells in suspension were treated with various combinations of USMB, chemotherapy and radiotherapy. Cells were treated with ultrasound and microbubbles (500 kHz pulse center frequency, 580 kPa peak negative pressure, 10 μs pulse duration, 60 s insonation time and 2% Definity microbubbles (v/v)), XRT (2 Gy), and Taxotere (TXT) at concentrations ranging from 0.001 to 0.1 nM for 5- and 120-minutes duration. Following treatment, cell viability was assessed using a clonogenic assay. Therapeutic efficiency of the combined treatments depended on chemotherapy and microbubble exposure conditions. Under the exposure conditions of the study, the triple combination therapy synergistically enhanced clonogenic cell death compared to single and double combination therapy. Cell viability of ∼2% was achieved with the triple combination therapy corresponding to ∼29, ∼37, and ∼38 folds decrease compared to XRT (57%), USMB (74%) and TXT (76%) alone conditions, respectively. In addition, the triple combination therapy decreased cell viability by ∼29, ∼19- and ∼11 folds compared to TXT2hr + USMB (58%), TXT2hr + XRT (37%), and USMB + XRT (22%), respectively.The in vivo PC3 tumours showed that USMB significantly enhanced cell death through detection of apoptosis (TUNEL) with both TXT and TXT + XRT. The study demonstrated that the triple combination therapy can significantly enhance cell death in prostate cancer cells both in vitro and in vivo under relatively low chemotherapy and ionizing radiation doses.

Moving model test on the aerodynamic pressure of bilateral inverted-L-shaped noise barriers caused by high-speed trains

With the increasing speed of high-speed trains, the aerodynamic impact of trains on high-speed railway noise barriers has gradually become a focus of attention. Compared with traditional vertical noise barriers, the bilateral inverted-L-shaped noise barrier (BILSNB) has better noise insulation performance, but its aerodynamic impact may be more prominent. In this research, a moving train-BILSNB test system with a scale ratio of 1:16.8 is established, and the influence of train speed and the opening width on the aerodynamic pressure is analysed and discussed. The large eddy simulation is applied to investigate the influence mechanism of flow field on the aerodynamic performance of the BILSNB. The main conclusions are as follows. (1) The pressure amplitude caused by the passing of the train head is 36.67% greater than that generated by the passing of the train tail on average. (2) The maximum pressure of the BILSNB appears at the bottom area. When the height of the measuring point increases from 0.5H to 1.6H, the pressure coefficient of peak positive pressure, peak negative pressure and pressure amplitude decrease by 29.0%, 17.0% and 15.5%, respectively. (3) The aerodynamic pressure in the BILSNB has a longitudinal end effect: the pressure coefficient of positive peak pressure and pressure amplitude at the inlet section are 27.6% and 17.5% higher than the corresponding average value of all middle sections, respectively. (4) The pressure amplitude of the BILSNB is approximately proportional to 2.15 power of the train speed. The pressure coefficient of the BILSNB is approximately a power function of the opening width, and the value range of the index is about (−0.26, −0.30). (5) The mechanism of the longitudinal end effect is as follows: When the train arrives at the inlet section, the high-speed airflow in front of the train head cannot flow around in time resulting in the impact effect of the airflow on the inlet section becomes greater than that on the middle section. The influence mechanism of the opening width is as follows: The narrower the opening width is, the more intense the airflow inside the BILSNB is compressed, and the impact effect of the airflow on the noise barrier is more significant.

Sonoporation of Immune Cells: Heterogeneous Impact on Lymphocytes, Monocytes and Granulocytes.

Microbubble-mediated ultrasound (MB-US) can be used to realize sonoporation and, in turn, facilitate the transfection of leukocytes in the immune system. Nevertheless, the bio-effects that can be induced by MB-US exposure on leukocytes have not been adequately studied, particularly for different leukocyte lineage subsets with distinct cytological characteristics. Here, we describe how that same set of MB-US exposure conditions would induce heterogeneous bio-effects on the three main leukocyte subsets: lymphocytes, monocytes and granulocytes. MB-US exposure was delivered by applying 1-MHz pulsed ultrasound (0.50-MPa peak negative pressure, 10% duty cycle, 30-s exposure period) in the presence of microbubbles (1:1 cell-to-bubble ratio); sonoporated and non-viable leukocytes were respectively labeled using calcein and propidium iodide. Flow cytometry was then performed to classify leukocytes into their corresponding subsets and to analyze each subset's post-exposure viability, sonoporation rate, uptake characteristics and morphology. Results revealed that, when subjected to MB-US exposure, granulocytes experienced the highest loss of viability (64.0 ± 11.0%) and the lowest sonoporation rate (6.3 ± 2.5%), despite maintaining their size and granularity. In contrast, lymphocytes exhibited the lowest loss of viability (20.9 ± 7.0%), while monocytes had the highest sonoporation rate (24.1 ± 13.6%). For these two sonoporated leukocyte subsets, their cell size and granularity were found to be reduced. Also, they exhibited graded levels of calcein uptake, whereas sonoporated granulocytes achieved only mild calcein uptake. These heterogeneous bio-effects should be accounted for when using MB-US and sonoporation in immunomodulation applications.

Nanodroplet-Mediated Low-Energy Mechanical Ultrasound Surgery.

Mechanical ultrasound surgery methods use short, high-intensity pulses to fractionate tissues. This study reports the development of a two-step technology for low-energy mechanical ultrasound surgery of tissues using nanodroplets to reduce the pressure threshold. Step 1 consists of vaporizing the nanodroplets into gaseous microbubbles via megahertz ultrasound excitation. Then, low-frequency ultrasound is applied to the microbubbles, which turns them into therapeutic warheads that trigger potent mechanical effects in the surrounding tissue. The use of nanoscale nanodroplets coupled with low-frequency ultrasound reduces the pressure threshold required for mechanical ultrasound surgery by an order of magnitude. In addition, their average diameter of 300 nm can overcome challenges associated with the size of microbubbles. Optimization experiments were performed to determine the ultrasound parameters for nanodroplet vaporization and the subsequent microbubble implosion processes. Optimal vaporization was obtained when transmitting a 2-cycle excitation pulse at a center frequency of 5 MHz and a peak negative pressure of 4.1 MPa (mechanical index=1.8). Low-frequency insonation of the generated microbubbles at a center frequency of 850, 250 or 80 kHz caused enhanced contrast reduction at a center frequency of 80 kHz, compared with the other frequencies, while operating at the same mechanical index of 0.9. Nanodroplet-mediated insonation of ex vivo chicken liver samples generated mechanical damage. Low-frequency treatment at a mechanical index of 0.9 and a center frequency of 80 kHz induced the largest lesion area (average of 0.59 mm2) compared with 250- and 850-kHz treatments with the same mechanical index (average lesions areas of 0.29 and 0.19 mm2, respectively, p < 0.001). The two-step approach makes it possible to conduct both the vaporization and implosion stages at mechanical indices below 1.9, thus avoiding undesired mechanical damage. The findings indicate that coupled with low-frequency ultrasound, nanodroplets can be used for low-energy mechanical ultrasound surgery.

Internalization of targeted microbubbles by endothelial cells and drug delivery by pores and tunnels.

Ultrasound insonification of microbubbles can locally enhance drug delivery by increasing the cell membrane permeability. To aid development of a safe and effective therapeutic microbubble, more insight into the microbubble-cell interaction is needed. In this in vitro study we aimed to investigate the initial 3D morphology of the endothelial cell membrane adjacent to individual microbubbles (n=301), determine whether this morphology was affected upon binding and by the type of ligand on the microbubble, and study its influence on microbubble oscillation and the drug delivery outcome. High-resolution 3D confocal microscopy revealed that targeted microbubbles were internalized by endothelial cells, while this was not the case for non-targeted or IgG1-κ control microbubbles. The extent of internalization was ligand-dependent, since αvβ3-targeted microbubbles were significantly more internalized than CD31-targeted microbubbles. Ultra-high-speed imaging (~17 Mfps) in combination with high-resolution confocal microscopy (n=246) showed that microbubble internalization resulted in a damped microbubble oscillation upon ultrasound insonification (2MHz, 200kPa peak negative pressure, 10cycles). Despite damped oscillation, the cell's susceptibility to sonoporation (as indicated by PI uptake) was increased for internalized microbubbles. Monitoring cell membrane integrity (n=230) showed the formation of either a pore, for intracellular delivery, or a tunnel (i.e. transcellular perforation), for transcellular delivery. Internalized microbubbles caused fewer transcellular perforations and smaller pore areas than non-internalized microbubbles. In conclusion, studying microbubble-mediated drug delivery using a state-of-the-art imaging system revealed receptor-mediated microbubble internalization and its effect on microbubble oscillation and resulting membrane perforation by pores and tunnels.

First Metabolomic Signature of Blood-Brain Barrier Opening Induced by Microbubble-Assisted Ultrasound.

Microbubble (MB)-assisted ultrasound (US) is a promising physical method to increase non-invasively, transiently, and precisely the permeability of the blood-brain barrier (BBB) to therapeutic molecules. Previous preclinical studies established the innocuity of this procedure using complementary analytical strategies including transcriptomics, histology, brain imaging, and behavioral tests. This cross-sectional study using rats aimed to investigate the metabolic processes following acoustically-mediated BBB opening in vivo using multimodal and multimatrices metabolomics approaches. After intravenous injection of MBs, the right striata were exposed to 1-MHz sinusoidal US waves at 0.6 MPa peak negative pressure with a burst length of 10 ms, for 30 s. Then, the striata, cerebrospinal fluid (CSF), blood serum, and urine were collected during sacrifice in three experimental groups at 3 h, 2 days, and 1 week after BBB opening (BBBO) and were compared to a control group where no US was applied. A well-established analytical workflow using nuclear magnetic resonance spectrometry and non-targeted and targeted high-performance liquid chromatography coupled to mass spectrometry were performed on biological tissues and fluids. In our experimental conditions, a reversible BBBO was observed in the striatum without physical damage or a change in rodent weight and behavior. Cerebral, peri-cerebral, and peripheral metabolomes displayed specific and sequential metabolic kinetics. The blood serum metabolome was more impacted in terms of the number of perturbated metabolisms than in the CSF, the striatum, and the urine. In addition, perturbations of arginine and arginine-related metabolisms were detected in all matrices after BBBO, suggesting activation of vasomotor processes and bioenergetic supply. The exploration of the tryptophan metabolism revealed a transient vascular inflammation and a perturbation of serotoninergic neurotransmission in the striatum. For the first time, we characterized the metabolic signature following the acoustically-mediated BBBO within the striatum and its surrounding biological compartments.

Abstract 2469: Image-guided focused ultrasound-mediated drug delivery for improved cancer treatment

Abstract Introduction: The first option for cancer treatment is often chemotherapy. However, studies have shown that only 1% of the injected dose reaches the target cancer as the tumor microenvironment presents a physical barrier for optimal drug delivery. Focused ultrasound (FUS) in combination with microbubble (MB) contrast agents is an emerging therapy to improve drug delivery by temporarily increasing microvascular permeability. This research details the development and testing of a novel ultrasound (US) image-guided FUS system and method for enhancing drug delivery to tumor tissue and volume space with comparison to a 2-dimensional (2-D) US therapeutic technology. Methods: Real-time US therapy was implemented on a programmable US system (Vantage 256, Verasonics Inc) equipped with a dual US transducer configuration for interleaved anatomical imaging and volumetric treatment delivery (HIFU-Plex, Sonic Concepts Inc). Both US imaging and therapeutic transducers are co-registered 128 element arrays with center frequencies of 3.5 and 2.0 MHz, respectively. The latter is a concentric array that enables beam steering in 3-dimensional (3-D) space. US treatment was performed at a peak negative pressure of 0.7 MPa (mechanical index, MI of 0.45), pulse repetition frequency of 10 Hz, and duty cycle of 10%. BALB/c mice (N = 22, Charles River Laboratory) were implanted with 2.0 × 105 breast cancer cells (4T1, ATCC). Once tumors reached 0.6 cm in size, mice were randomly divided into a 3-D or 2-D US therapy group or sham control. US therapy was performed following an intravascular injection of microbubbles (Definity, Lantheus Medical Imaging) and IR-780 dye. Note microbubbles function as a therapeutic mediator whereas the fluorescent dye represents a surrogate small molecule drug. Live animal fluorescent imaging was performed at baseline before US therapy (0 h) and again at 1, 24, and 48 h. Following the 48 h timepoint, animals were euthanized and tumors surgically excised for ex vivo analysis. Results: 3-D US-mediated therapy improved molecular delivery to tumor tissue by 150 and 180% at 24 and 48 h, respectively, when compared to our previously established 2-D US therapeutic approach (p = 0.22) or sham therapy (p = 0.07). A similar trend was observed during ex vivo imaging of excised tumor samples treated with 3-D US therapy as compared to the 2-D US therapeutic approach (p = 0.41) or sham therapy (p = 0.04). Dye extraction further confirmed these observations (p &amp;gt; 0.12). Conclusions: 3-D US therapy improved molecular delivery to the tumor volume compared to our previously established 2-D US-based method as confirmed by a series of optical imaging studies. Citation Format: Ryan Margolis, Junjie Li, Lokesh Basavarajappa, Kenneth Hoyt. Image-guided focused ultrasound-mediated drug delivery for improved cancer treatment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2469.

DDEL-06. Drug Delivery to the Pons Using Short-Pulse Focused Ultrasound and Microbubble Exposure for the Treatment of Diffuse Midline Glioma

Despite advances in understanding diffuse midline glioma (DMG-H3K27), including DIPG, there are still no effective treatments available, and the dismal clinical prognosis remains. This is partly because of tumour spread behind an intact blood brain barrier (BBB), preventing drug delivery and the reason for many drugs failing in the clinic. The use of focused ultrasound and intravenous microbubbles enables temporary increases in BBB permeability, allowing drugs to enter the targeted brain region. Building on recent research demonstrating that short pulses (<5 µs) of ultrasound can deliver drugs safely and uniformly to the hippocampus, we evaluated whether a similar result was achievable in the pons of mice. Mice were exposed to ultrasound (peak-negative pressure: 0.4 MPa, pulse length: 5 cycles, centre frequency 1 MHz) emitted in bursts of 38 pulses. During exposure mice received an intravenous injection of SonoVue(R) microbubbles and a fluorescently-tagged tracer (dextran, 3 kDa), acting as a drug mimic. Dextran was successfully delivered to the pons of non-tumour-bearing mice assessed by fluorescence microscopy immediately post-treatment. Dextran delivery was repeatable and confined to the targeted pons region with a homogenous distribution, typical of short pulse ultrasound, and important for treating DMG to ensure all tumour cells receive an equal drug dose. No damage to the brain was observed after H&E staining. Panobinostat has shown promise in vitro but tolerated doses have not shown therapeutic benefit in vivo as it does not cross the BBB. The in vitro toxicity of panobinostat was confirmed in a Nestin-Tv-a/p53fl/fl, RCAS-ACVR1R206H + RCAS-H3.1K27M murine cell line, with a GI50 of 15.56 nM. The ability of focused ultrasound to deliver panobinostat across the BBB to these tumours grown orthotopically will be assessed. Overall, we hope to develop a drug delivery system, that enables therapeutics to cross the BBB, expanding treatment options for DMG.

Acoustic Modulation Enables Proton Detection With Nanodroplets at Body Temperature.

Superheated nanodroplet (ND) vaporization by proton radiation was recently demonstrated, opening the door to ultrasound-based in vivo proton range verification. However, at body temperature and physiological pressures, perfluorobutane nanodroplets (PFB-NDs), which offer a good compromise between stability and radiation sensitivity, are not directly sensitive to primary protons. Instead, they are vaporized by infrequent secondary particles, which limits the precision for range verification. The radiation-induced vaporization threshold (i.e., sensitization threshold) can be reduced by lowering the pressure in the droplet such that ND vaporization by primary protons can occur. Here, we propose to use an acoustic field to modulate the pressure, intermittently lowering the proton sensitization threshold of PFB-NDs during the rarefactional phase of the ultrasound wave. Simultaneous proton irradiation and sonication with a 1.1 MHz focused transducer, using increasing peak negative pressures (PNPs), were applied on a dilution of PFB-NDs flowing in a tube, while vaporization was acoustically monitored with a linear array. Sensitization to primary protons was achieved at temperatures between [Formula: see text] and 40 °C using acoustic PNPs of relatively low amplitude (from 800 to 200 kPa, respectively), while sonication alone did not lead to ND vaporization at those PNPs. Sensitization was also measured at the clinically relevant body temperature (i.e., 37 °C) using a PNP of 400 kPa. These findings confirm that acoustic modulation lowers the sensitization threshold of superheated NDs, enabling a direct proton response at body temperature.

Field study on train-induced aerodynamic pressure near the entrance of fully enclosed sound barriers

A systematic field test was conducted for aerodynamic pressure near the entrance of the double-track fully enclosed sound barriers (FESBs). The generating mechanism of pressure was revealed by referring to the slipstream distribution around the moving train and considering the influence of marshalling length, speed and streamline of trains. Results show that near the entrance, the compression wave caused by the head car is predominant and determined by the maximum, minimum and peak-to-peak pressure, while near the exit, the negative and positive peak pressure are, respectively, triggered by the head car and tail car. Uneven pressure distribution along the circumference of the FESB wall and the greater pressure is on the wall near the running train. The head car of double-connection electric multiple units (EMUs) generates greater pressure near the entrance than that of common EMUs, and the tail car of double-connection EMUs causes weaker pressure transient, while near the exit, the head car of double-connection EMU induces weaker pressure. Even if the train speed increases in a small range, the pressure near the entrance/exit of the FESB increases, and the blunter the train is, the greater and the more evenly distributed pressure on the FESB will be.

Spatiotemporal Dynamics and Mechanisms of Actin Cytoskeletal Re-modeling in Cells Perforated by Ultrasound-Driven Microbubbles.

To develop new strategies for improving the efficacy and biosafety of sonoporation-based macromolecule delivery, it is essential to understand the mechanisms underlying plasma membrane re-sealing and function recovery of the cells perforated by ultrasound-driven microbubbles. However, we lack a clear understanding of the spatiotemporal dynamics of the disrupted actin cytoskeleton and its role in the re-sealing of sonoporated cells. Here we used a customized experimental setup for single-pulse ultrasound (133.33-µs duration and 0.70-MPa peak negative pressure) exposure to microbubbles and for real-time recording of single-cell (human umbilical vein endothelial cell) responses by laser confocal microscopic imaging. We found that in reversibly sonoporated cells, the locally disrupted actin cytoskeleton, which was spatially correlated with the perforated plasma membrane, underwent three successive phases (expansion; contraction and re-sealing; and recovery) to re-model and that each phase of the disrupted actin cytoskeleton was approximately synchronized with that of the perforated plasma membrane. Moreover, compared with the closing time of the perforated plasma membrane, the same time was used for the re-sealing of the actin cytoskeleton in mildly sonoporated cells and a longer time was required in moderately sonoporated cells. Further, the generation, directional migration, accumulation and re-polymerization of globular actin polymers during the three phases drove the re-modeling of the actin cytoskeleton. However, in irreversibly sonoporated cells, the actin cytoskeleton, which underwent expansion and no contraction, was progressively de-polymerized and could not be re-sealed. Finally, we found that intracellular calcium transients were essential for the recruitment of globular actin and the re-modeling of the actin cytoskeleton. These results provide new insight into the role of actin cytoskeleton dynamics in the re-sealing of sonoporated cells and serve to guide the design of new strategies for sonoporation-based delivery.

A Fiber Bragg Grating-Based Sensor for Passive Cavitation Detection at MHz Frequencies.

Fiber Bragg gratings (FBGs) are a potential alternative to piezoelectric ultrasound sensors for applications that demand high sensitivity and immunity to electromagnetic interference (EMI). However, limited data exist on the quantitative performance characterization of FBG sensors in the MHz frequency range relevant to biomedical ultrasound. In this work, we evaluated an FBG to detect MHz-frequency ultrasound and tested the feasibility of measuring passive cavitation signals nucleated using a commercial contrast agent (SonoVue). The sensitivity, repeatability, and linearity of the measurements were assessed for ultrasound measurements at 1, 5, and 10 MHz. The bandwidth of the FBG sensor was measured and compared to that of a calibrated needle hydrophone. The FBG showed a sensitivity of 0.99, 0.769, and 0.818 V/MPa for 1, 5, and 10 MHz ultrasound, respectively. The sensor also exhibited linear response ( 0.975 ≤ R -Squared ≤ 0.996) and good repeatability with a coefficient of variation (CV) less than 5.5%. A 2-MHz focused transducer was used to insonify SonoVue microbubbles at a peak negative pressure of 175 kPa and passive cavitation emissions were measured, in which subharmonic and ultraharmonic spectral peaks were observed. These results demonstrate the potential of FBGs for MHz-range ultrasound applications, including passive cavitation detection (PCD).

Slow-Flow Ultrasound Localization Microscopy Using Recondensation of Perfluoropentane Nanodroplets.

Ultrasound localization microscopy (ULM) is an emerging, super-resolution imaging technique for detailed mapping of the microvascular structure and flow velocity via subwavelength localization and tracking of microbubbles. Because microbubbles rely on blood flow for movement throughout the vascular space, acquisition times can be long in the smallest, low-flow microvessels. In addition, detection of microbubbles in low-flow regions can be difficult because of minimal separation of microbubble signal from tissue. Nanoscale, phase-change contrast agents (PCCAs) have emerged as a switchable, intermittent or persisting contrast agent for ULM via acoustic droplet vaporization (ADV). Here, the focus is on characterizing the spatiotemporal contrast properties of less volatile perfluoropentane (PFP) PCCAs. The results indicate that at physiological temperature, nanoscale PFP PCCAs with diameters less than 100 nm disappear within microseconds after ADV with high-frequency ultrasound (16 MHz, 5- to 6-MPa peak negative pressure) and that nanoscale PFP PCCAs have an inherent deactivation mechanism via immediate recondensation after ADV. This "blinking" on-and-off contrast signal allowed separation of flow in an in vitro flow phantom, regardless of flow conditions, although with a need for some replenishment at very low flow conditions to maintain count rate. This blinking behavior allows for rapid spatial mapping in areas of low or no flow with ULM, but limits velocity tracking because there is no stable bubble formation with nanoscale PFP PCCAs.

Nanobubbles are Non-Echogenic for Fundamental-Mode Contrast-Enhanced Ultrasound Imaging.

Microbubbles (1-10 μm diameter) have been used as conventional ultrasound contrast agents (UCAs) for applications in contrast-enhanced ultrasound (CEUS) imaging. Nanobubbles (<1 μm diameter) have recently been proposed as potential extravascular UCAs that can extravasate from the leaky vasculature of tumors or sites of inflammation. However, the echogenicity of nanobubbles for CEUS remains controversial owing to prior studies that have shown very low ultrasound backscatter. We hypothesize that microbubble contamination in nanobubble formulations may explain the discrepancy. To test our hypothesis, we examined the size distributions of lipid-coated nanobubble and microbubble suspensions using multiple sizing techniques, examined their echogenicity in an agar phantom with fundamental-mode CEUS at 7 MHz and 330 kPa peak negative pressure, and interpreted our results with simulations of the modified Rayleigh-Plesset model. We found that nanobubble formulations contained a small contamination of microbubbles. Once the contribution from these microbubbles is removed from the acoustic backscatter, the acoustic contrast of the nanobubbles was shown to be near noise levels. This result indicates that nanobubbles have limited utility as UCAs for CEUS.

Ultrasound-Mediated Blood-Brain Barrier Disruption for Drug Delivery: A Systematic Review of Protocols, Efficacy, and Safety Outcomes from Preclinical and Clinical Studies.

Ultrasound-mediated blood–brain barrier (BBB) disruption has garnered focus as a method of delivering normally impenetrable drugs into the brain. Numerous studies have investigated this approach, and a diverse set of ultrasound parameters appear to influence the efficacy and safety of this approach. An understanding of these findings is essential for safe and reproducible BBB disruption, as well as in identifying the limitations and gaps for further advancement of this drug delivery approach. We aimed to collate and summarise protocols and parameters for achieving ultrasound-mediated BBB disruption in animal and clinical studies, as well as the efficacy and safety methods and outcomes associated with each. A systematic search of electronic databases helped in identifying relevant, included studies. Reference lists of included studies were further screened to identify supplemental studies for inclusion. In total, 107 articles were included in this review, and the following parameters were identified as influencing efficacy and safety outcomes: microbubbles, transducer frequency, peak-negative pressure, pulse characteristics, and the dosing of ultrasound applications. Current protocols and parameters achieving ultrasound-mediated BBB disruption, as well as their associated efficacy and safety outcomes, are identified and summarised. Greater standardisation of protocols and parameters in future preclinical and clinical studies is required to inform robust clinical translation.

Quantifying the role of microstreaming in sonoporation

Cavitation microstreaming has been identified as an important mechanism underpinning sonoporation. The reported evidence includes experimental observation of the permeabilization of non-adherent cells or vesicles in microstreaming and theoretical and experimental estimates of the fluid shear stress required to induce pore formation in cell membranes and/or the activation of membrane components such as ion channels. The aim of this study was to assess the relative contribution of microstreaming to sonoporation observed in vitro with adherent cells, as commonly used in drug delivery studies. Streak velocimetry measurements were performed within a microfluidic cell of the microstreaming around a contrast agent microbubble driven at 0.5 and 1MHz and peak negative pressures up to 1MPa. The results suggest that, for the cell densities and microbubble concentrations typically reported in sonoporation studies, only a very small proportion of cells within a typical culture device would be exposed to sufficiently high shear stresses to be permeabilised. This finding is inconsistent with reported results for drug delivery efficiency and suggests that microstreaming may not be the dominant mechanism of sonoporation in this type of experiment.

Low frequency (&amp;lt;1 MHz) transducers for intravenous sonothrombolysis

Venous embolism is a great challenge in the treatment of a wide range of diseases, including cardiovascular diseases and cancers. Ultrasound-mediated thrombolysis (sonothrombolysis) presents a promising treatment of venous embolism. In this paper, miniature ultrasound transducers with frequencies ranging from 250 kHz to 800 kHz were developed for microbubble/nanodroplet-mediated intravenous sonothrombolysis tests. Relative low frequency (e.g., &amp;lt;1 MHz) is preferred because of the findings that cavitation plays an important role in sonothrombolysis, and microbubble-associated cavitation events can be enhanced at low frequencies. Small aperture (e.g. &amp;lt;2 mm × 2 mm) low-frequency transducers are usually known with high electrical impedance (e.g., &amp;gt;1000 Ω), which is not preferred for efficient electromechanical transduction. To address this challenge, multilayer acoustic stacks were designed, fabricated, and characterized. The measured electrical impedance of these stacks is mostly closer to 50 Ω, and the peak negative pressure is higher than 1MPa. These small aperture transducers were then used in in vitro- thrombolysis tests with unretracted and retracted blood clots, showing promising lysis rates. Other techniques such as laser ultrasound transducers, vortex wave generation, and magnetic microbubbles were also investigated for thrombolysis tests.