Clot Retraction Affects the Extent of Ultrasound-Enhanced Thrombolysis in an Ex Vivo Porcine Thrombosis Model

We investigated ultrasound-enhanced thrombolysis in two whole-blood clot models using a Food and Drug Administration-approved contrast agent (Definity, Lantheus Medical Imaging; Billerica, MA USA) and thrombolytic drug (recombinant tissue-type plasminogen activator [rt-PA]) (Genentech; South San Francisco, CA USA). Porcine venous blood was collected from donor hogs and coagulated in vials made of two different materials. This method produced clots with differing compositional properties, as determined by routine scanning electron microscopy and histology. Clots were deployed in an ex vivo porcine thrombosis model, and exposed to an intermittent ultrasound scheme previously developed to maximize stable cavitation while acoustic emissions were detected. Exposure to 3.15 μg/mL rt-PA promoted lysis in both clot models, compared with exposure to plasma alone. However, only unretracted clots experienced significant enhancement of thrombolysis in the presence of rt-PA, Definity, and ultrasound, compared with treatment with rt-PA. In these clots, microscopy revealed loose erythrocyte aggregates, a significantly less extensive fibrin network and a higher porosity, which may facilitate increased penetration of thrombolytics by cavitation.     

Ultrasound-enhanced rt-PA thrombolysis in an ex vivo porcine carotid artery model

Ultrasound is known to enhance recombinant tissue plasminogen activator (rt-PA) thrombolysis. In this study, occlusive porcine whole blood clots were placed in flowing plasma within living porcine carotid arteries. Ultrasonically induced stable cavitation was investigated as an adjuvant to rt-PA thrombolysis. Aged, retracted clots were exposed to plasma alone, plasma containing rt-PA (7.1 ± 3.8 μg/mL) or plasma with rt-PA and Definity^® ultrasound contrast agent (0.79 ± 0.47 μL/mL) with and without 120-kHz continuous wave ultrasound at a peak-to-peak pressure amplitude of 0.44 MPa. An insonation scheme was formulated to promote and maximize stable cavitation activity by incorporating ultrasound quiescent periods that allowed for the inflow of Definity^®-rich plasma. Cavitation was measured with a passive acoustic detector throughout thrombolytic treatment. Thrombolytic efficacy was measured by comparing clot mass before and after treatment. Average mass loss for clots exposed to rt-PA and Definity^® without ultrasound (n = 7) was 34%, and with ultrasound (n = 6) was 83%, which constituted a significant difference (p < 0.0001). Without Definity^® there was no thrombolytic enhancement by ultrasound exposure alone at this pressure amplitude (n = 5, p < 0.0001). In the low-oxygen environment of the ischemic artery, significant loss of endothelium occurred but no correlation was observed between arterial tissue damage and treatment type. Acoustic stable cavitation nucleated by an infusion of Definity^® enhances rt-PA thrombolysis without apparent treatment-related damage in this ex vivo porcine carotid artery model.     

Correlation of cavitation with ultrasound enhancement of thrombolysis

Pulsed ultrasound, when used as an adjuvant to recombinant tissue plasminogen activator (rt-PA), has been shown to enhance thrombolysis in the laboratory as well as in clinical trials for the treatment of ischemic stroke. The exact mechanism of this enhancement has not yet been elucidated. In this work, stable and inertial cavitation (SC and IC) are investigated as possible mechanisms for this enhancement. A passive cavitation detection scheme was utilized to measure cavitation thresholds at 120 kHz (80% duty cycle, 1667 Hz pulse repetition frequency) for four host fluid and sample combinations: plasma, plasma with rt-PA, plasma with clot and plasma with clot and rt-PA. Following cavitation threshold determination, clots were exposed to pulsed ultrasound for 30 min in vitro using three separate ultrasound treatment regimes: (1) no cavitation (0.15 MPa), (2) SC alone (0.24 MPa) or (3) SC + IC combined (0.36 MPa) in the presence of rt-PA. Percent clot mass loss after each treatment was used to determine thrombolysis efficacy. The highest percent mass loss was observed in the stable cavitation regime (26%), followed by the combined stable and inertial cavitation regime (20.7%). Interestingly, the percent mass loss in clots exposed to ultrasound without cavitation (13.7%) was not statistically significantly different from rt-PA alone (13%) [p > 0.05]. Significant enhancement of thrombolysis correlates with presence of cavitation and stable cavitation appears to play a more important role in the enhancement of thrombolysis. (E-mail: ).     

Acceleration of thrombolysis with ultrasound through the cranium in a flow model

Thrombolysis is an efficient therapy for hyperacute stroke within a limited time window. Neurological outcome depends on the recanalization time of the occluded vessel. Nonthermal effects of low-frequency ultrasound (US) accelerate enzymatic fibrinolysis in vitro. We examined the effects of transcranially applied US on recombinant tissue plasminogen activator (rt-PA)-mediated thrombolysis in a flow model in vitro. Pure fibrin clots were placed in a continuous-pressure flow model and treated with rt-PA during 1-MHz US exposure (0.5 W/cm(2); spatial peak, temporal peak intensity). Transcranial and direct US application in combination with rt-PA significantly (p<0.001) shortened recanalization time, increased perfusion flow and reperfusion rate in comparison with rt-PA-mediated thrombolysis alone. Recanalization rate within 30 min was 90-100% in the US-exposed clots vs. 30% in the clots treated only with rt-PA. Our results suggest that transcranial application of 1-MHz US may accelerate reperfusion and recanalization rate of occluded intracerebral vessels by enhancing rt-PA-mediated thrombolysis. Shortening of recanalization time could contribute to optimizing effects of acute thrombolytic stroke therapy.     

Ultrasound-induced thermal elevation in clotted blood and cranial bone

Ultrasound thermal effects have been hypothesized to contribute to ultrasound-assisted thrombolysis. To explore the thermal mechanism of ultrasound-enhanced thrombolysis with recombinant tissue plasminogen activator (rt-PA) for the treatment of ischemic stroke, a detailed investigation is needed of the heating produced in skull, brain and blood clots. A theoretical model is developed to provide an estimate for the worst-case scenario of the temperature increase in blood clots and on the surface of cranial bone exposed to 0.12- to 3.5-MHz ultrasound. Thermal elevation was also assessed experimentally in human temporal bone, human clots and porcine clots exposed to 0.12 to 3.5-MHz pulsed ultrasound in vitro with a peak-to-peak pressure of 0.25 MPa and 80% duty cycle. Blood clots exposed to 0.12-MHz pulsed ultrasound exhibited a small temperature increase (0.25 degrees C) and bone exposed to 1.0-MHz pulsed ultrasound exhibited the highest temperature increase (1.0 degrees C). These experimental results were compared with the predicted temperature elevations.     

In vitro thrombolysis enhanced by standing and travelling ultrasound wave fields

Success of thrombolytic therapy depends on penetration of recombinant tissue plasminogen activator (rt-PA) into clots. Ultrasound (US) of therapeutic quality accelerates thrombolysis in vitro. As yet, only the effects of travelling acoustic waves on thrombolysis have been investigated, and the impact of standing acoustic waves has been neglected. In the present study, we examined the effects of standing and travelling US wave fields applied continuously for 1 h (frequency 2 MHz, acoustic intensity 1.2 W/cm(2)) on thrombolysis enhancement by measuring clot weight reduction and concentration of fibrin degradation product D-dimer (FDP-DD) produced from clots subjected to rt-PA. The level of FDP-DD was 1.8 times greater in travelling than in standing acoustic waves. Thrombolysis enhancement was 46.0 +/- 20.8% in standing and 116.8 +/- 23.1% in travelling acoustic waves. Travelling waves enhanced thrombolysis significantly more (p < 0.0001) than did standing waves.     

Low-frequency, low-intensity ultrasound accelerates thrombolysis through the skull.

Systemic thrombolysis of acute ischemic stroke with recombinant tissue plasminogen activator (rt-PA) has been established recently. Whereas the delay to and the rate of vessel recanalization are unknown, they are likely slower and smaller than for local application of rt-PA. This may contribute to the small benefits of recovery reported and stimulate further investigations to improve clot lysis. Pilot studies indicate that continuous-wave low-frequency ultrasound (US) can accelerate rt-PA-mediated recanalization of peripheral thrombotic vessel occlusion. For the hypothesized therapeutical purpose in stroke treatment, we measured the attenuation of ultrasound through the skull at different frequencies and intensities (33.3 and 71.4 kHz; 0.5 and 3.4 W/cm2), and investigated thrombolysis in vitro (n = 125 clots). Attenuation was lowest by transtemporal insonation of 33.3 kHz, 0.1 dB (0.9). Thrombolysis (artificial fibrin-rich clots) was significantly increased after 1 h (p < 0.025) and after 3 h (p < 0.01) for US treatment in combination with rt-PA vs. rt-PA alone. Results suggest that US increases rt-PA-mediated thrombolysis through the skull and may improve benefits of thrombolytic stroke treatment in vivo.     

Low-frequency ultrasound induces nonenzymatic thrombolysis in vitro.

OBJECTIVE: To evaluate whether ultrasound, applied over a distance of several centimeters and in the absence of thrombolytic agents, may have a thrombolytic effect on blood clots. METHODS: Low-frequency (20 kHz) continuous wave ultrasound at different intensity levels (0.15-1.2 W/cm2) and exposure times (5, 10, and 20 minutes) was assessed for its potential to induce thrombolysis of fresh human blood clots. The ultrasound effect was also studied in combination with recombinant tissue-type plasminogen activator-mediated thrombolysis. Experiments were carried out in a flow model in degassed sodium phosphate buffer at 37 degrees C at a distance of 3 cm from the ultrasonic probe to the blood clots. Regardless of ultrasound exposure times, blood clots in all experimental groups and the control group were left in the flow system for 20 minutes. RESULTS: The use of ultrasound alone showed a significant thrombolytic effect compared with the control group, with a statistically significant effect at 0.15 W/cm2 and exposure of 10 minutes (P = .02). There was a clear correlation between the extent of weight loss and the chosen intensity level and exposure time. Complete disruption in 8 of 10 blood clots occurred at 1.2 W/cm2 within 10 min. Addition of ultrasound to recombinant tissue-type plasminogen activator-mediated thrombolysis significantly enhanced thrombolysis compared with application of recombinant tissue-type plasminogen activator or ultrasound alone (P = .0001), with the results pointing toward a purely additive, nonsynergistic effect of the 2 treatment modalities. Lysis was more effective in fresh thrombi. CONCLUSIONS: The use of low-frequency ultrasound alone, without addition of a thrombolytic drug, has the potential to induce thrombolysis over a distance. Combination of ultrasound with recombinant tissue-type plasminogen activator is superior to either treatment alone. Ultrasound is a promising tool for developing an alternative or additional treatment modality for acute cerebral vessel occlusion.     

Shaken and Stirred: Mechanisms of Ultrasound-Enhanced Thrombolysis

The use of ultrasound and microbubbles as an effective adjuvant to thrombolytics has been reported in vitro, ex vivo and in vivo. However, the specific mechanisms underlying ultrasound-enhanced thrombolysis have yet to be elucidated. We present visual observations illustrating two mechanisms of ultrasound-enhanced thrombolysis: acoustic cavitation and radiation force. An in vitro flow model was developed to observe human whole blood clots exposed to human fresh-frozen plasma, recombinant tissue-type plasminogen activator (0, 0.32, 1.58 or 3.15 μg/mL) and the ultrasound contrast agent Definity (2 μL/mL). Intermittent, continuous-wave ultrasound (120 kHz, 0.44 MPa peak-to-peak pressure) was used to insonify the perfusate. Ultraharmonic emissions indicative of stable cavitation were monitored with a passive cavitation detector. The clot was observed with an inverted microscope, and images were recorded with a charge-coupled device camera. The images were post-processed to determine the time-dependent clot diameter and root-mean-square velocity of the clot position. Clot lysis occurred preferentially surrounding large, resonant-sized bubbles undergoing stable oscillations. Ultraharmonic emissions from stable cavitation were found to correlate with the lytic rate. Clots were observed to translate synchronously with the initiation and cessation of the ultrasound exposure. The root-mean-square velocity of the clot correlated with the lytic rate. These data provide visual documentation of stable cavitation activity and radiation force during sub-megahertz sonothrombolysis. The observations of this study suggest that the process of clot lysis is complex, and both stable cavitation and radiation force are mechanistically responsible for this beneficial bio-effect in this in vitro model.     

Effect of Recombinant Tissue Plasminogen Activator and 120-kHz Ultrasound on Porcine Intracranial Thrombus Density

Surgical intervention for the treatment of intracerebral hemorrhage (ICH) has been limited by inadequate lysis of the target thrombus. Adjuvant transcranial ultrasound exposure is hypothesized to improve thrombolysis, expedite hematoma evacuation and improve clinical outcomes. A juvenile porcine intracerebral hemorrhage model was established by direct infusion of autologous blood into the porcine white matter. Thrombi were either not treated (sham) or treated with recombinant tissue plasminogen activator alone (rt-PA only) or in combination with pulsed transcranial 120-kHz ultrasound (sonothrombolysis). After treatment, pigs were euthanized, the heads frozen and sectioned and the thrombi extracted. D-Dimer and thrombus density assays were used to assess degree of lysis. Both porcine and human D-dimer assays tested did not have sufficient sensitivity to detect porcine D-dimer. Thrombi treated with rt-PA with or without 120-kHz ultrasound had a significantly lower density compared with sham-treated thrombi. No enhancement of rt-PA-mediated thrombolysis was noted with the addition of 120-kHz ultrasound (sonothrombolysis). The thrombus density assay revealed thrombolytic efficacy caused by rt-PA in an in vivo juvenile porcine model of intracerebral hemorrhage. Transcranial sonothrombolysis did not enhance rt-PA-induced thrombolysis, likely because of the lack of exogenous cavitation nuclei.     

Cavitational mechanisms in ultrasound-accelerated thrombolysis at 1 MHz

Inertial cavitation is hypothesized to be a mechanism by which ultrasound (US) accelerates the dissolution of human blood clots when the clot is exposed to a thrombolytic agent such as tissue plasminogen activator (t-PA). To test this hypothesis, radiolabeled fibrin clots were exposed or sham-exposed in vitro to 1 MHz c.w. US in a rotating sample holder immersed in a water-filled tank at 37 degrees C. Percent clot dissolution after 60 min of US exposure was assessed by removing the samples, centrifuging, and measuring the radioactivity of the supernatant fluid relative to the pelletized material. To suppress acoustic cavitation, the exposure tank was contained within a hyperbaric chamber capable of pneumatic pressurization to 10 atmospheres (gauge). Various combinations of static pressure (0, 2, 5, and 7.5 atm gauge), US (0 or 4 W/cm(2) SATA), and t-PA (0 or 10 microg/mL) were employed, showing statistically significant reductions in thrombolytic activity as static pressure increased. To gain further insight, an active cavitation detection scheme was employed in which 1-micros duration tonebursts of 20-MHz US (< 1 kPa peak negative pressure, 1 Hz PRF) were used to interrogate clots subjected to US and static pressure. Results of this cavitation detection scheme showed that scattering from within the clot and broadband acoustic emissions that were both present during insonification were significantly reduced with application of static pressure. However, only about half of the acceleration of thrombolysis due to US could be removed by static pressure, suggesting the possibility of other mechanisms in addition to inertial cavitation.     

Duty cycle dependence of ultrasound enhanced thrombolysis in a human clot model

Combined ultrasound and tissue plasminogen activator (rt-PA) therapy, or ultrasound enhanced thrombolysis (UET), has been shown to improve recanalization in patients with acute ischemic stroke. We measured the effect of ultrasound duty cycle on the lytic efficacy of 120 kHz UET in an in vitro human clot model. The hypothesis was that an increase in duty cycle increases rt-PA lytic efficacy. Human whole blood clots were exposed to 120-kHz ultrasound and rt-PA for 30 min in human plasma. The duty cycle ranged from 0% to 80%, where 0% represents sham exposure. Clot lytic rate was measured by recording the clot width over time. The clot width after 30 min exposure to rt-PA and ultrasound decreases with increasing duty cycle. The initial lytic rate increased linearly with duty cycle.     

Effect of Clot Stiffness on Recombinant Tissue Plasminogen Activator Lytic Susceptibility in Vitro

The lytic recombinant tissue plasminogen activator (rt-PA) is the only drug approved by the Food and Drug Administration for treating ischemic stroke. Less than 40% of patients with large vessel occlusions who are treated with rt-PA have improved blood flow. However, up to 6% of all patients receiving rt-PA develop intracerebral hemorrhage. Predicting the efficacy of rt-PA treatment a priori could help guide therapeutic decision making, such that rt-PA is administered only to those individuals who would benefit from this treatment. Clot composition and structure affect the lytic efficacy of rt-PA and have an impact on elasticity. However, the relationship between clot elasticity and rt-PA lytic susceptibility has not been adequately investigated. The goal of this study was to quantify the relationship between clot elasticity and rt-PA susceptibility in vitro. Human and porcine highly retracted and mildly retracted clots were fabricated in glass pipettes. The rt-PA lytic susceptibility was evaluated in vitro using the percent clot mass loss. The Young's moduli of the clots were estimated using ultrasound-based single-track-location shear wave elasticity imaging. The percent mass loss in mildly retracted porcine and human clots (28.9 ± 6.1% and 45.2 ± 7.1%, respectively) was significantly higher (p < 0.05) than in highly retracted porcine and human clots (10.9 ± 2.1% and 25.5 ± 10.0%, respectively). Furthermore, the Young's moduli of highly retracted porcine and human clots (5.33 ± 0.92 and 3.21 ± 1.97 kPa, respectively) were significantly higher (p < 0.05) than those of mildly retracted porcine and human clots (2.66 ± 0.55 and 0.79 ± 0.21 kPa, respectively). The results revealed an inverse relationship between the percent clot mass loss and Young's modulus. These findings motivate continued investigation of ultrasound-based methods to assess clot stiffness in order to predict rt-PA thrombolytic efficacy.     

In Vitro Examination of the Thrombolytic Efficacy of Desmoteplase and Therapeutic Ultrasound Compared with rt-PA

The aim of the study described here was to evaluate the thrombolytic efficacy of combined treatment with the fibrin-selective plasminogen activator desmoteplase (DSPA) and therapeutic ultrasound (sonothrombolysis [STL]) compared with conventional rt-PA (recombinant tissue plasminogen activator) treatment in vitro. Lysis rates were determined by the weight loss of platelet-rich plasma (PRP) clots treated with rt-PA (60 kU/mL) or DSPA (2 μg/mL) combined with pulsed wave ultrasound (2 MHz, 0.179 W/cm²). To reveal the individual effects of medication and ultrasound, lysis rates were also determined for DSPA monotherapy and for combined treatment with rt-PA and ultrasound. Clots solely placed in plasma served as the control group. Lysis increased significantly with rt-PA (26.5 ± 7.8%) and DSPA (30.5 ± 6%) compared with the control group (18.2 ± 5.9%) (each p < 0.001). DSPA lysis was more effective than rt-PA lysis (without STL: p = 0.015, with STL: p = 0.01). Combined treatment with DSPA and 2-MHz STL significantly exceeded rt-PA lysis (32.8% vs. 26.5%, p < 0.001).     

Cavitational mechanisms in ultrasound-accelerated fibrinolysis

The role of both inertial and stable cavitation was investigated during in vitro ultrasound-accelerated fibrinolysis by recombinant tissue plasminogen activator (rt-PA) in the presence and absence of Optison. A unique treatment configuration applied ultrasound, rt-PA and Optison to the interior of a plasma clot. Lysis efficacy was measured as clot weight reduction. Cavitational mechanisms were investigated by monitoring subharmonic and broadband noise. In the absence of Optison, 1.7 MHz pulsed ultrasound with 1.5 MPa peak-negative pressure applied for 30 min resulted in 45 +/- 19% lysis enhancement relative to rt-PA alone. Cavitation was not detected, indicating a role of noncavitational effects of ultrasound. The addition of Optison increased lysis enhancement to 88 +/- 25%. Inertial cavitation was present only at the start of the exposure, while low-amplitude subharmonic emissions persisted throughout. Additional protocols suggested a possible correlation between the increased lysis in the presence of Optison and the subharmonic emission, indicating a potentially important role of stable rather than inertial cavitation in microbubble-enhanced ultrasound-accelerated rt-PA-mediated thrombolysis.     

Ultrasound and Intra-Clot Microbubbles Enhanced Catheter-Directed Thrombolysis in Vitro and in Vivo

Insufficient penetration of microbubbles (MBs) into the vessel-obstructing thrombi significantly reduces the effectiveness of ultrasound thrombolysis (UT). The widely performed catheter-directed therapy (CDT) makes it possible to increase the local concentration of MBs in the clot. In an occluded vessel with a bypass, treatment of fresh human whole blood clots with CDT-based UT (intra-clot injection of MBs and urokinase, with ultrasound exposure) resulted in a significantly higher percentage of weight loss (35.32 ± 15.42%), compared with CDT alone (19.64 ± 4.71%), non-CDT-based UT (systemic administration of urokinase and MBs, with ultrasound exposure, 8.79 ± 3.02%) and systemic thrombolysis (7.90 ± 2.14). Ultrasound and intra-clot MB enhancement of CDT was further confirmed by a rabbit IVC thrombolysis study, where CDT-based UT resulted in significantly more effective thrombolysis compared with CDT alone. In summary, combining CDT with intra-clot MB-induced acoustic cavitation can improve thrombolysis.     

Tissue plasminogen activator concentration dependence of 120 kHz ultrasound-enhanced thrombolysis

It has been known for some time that the application of ultrasound can enhance the efficacy of thrombolytic medications such as recombinant tissue plasminogen activator (rt-PA). Potential clinical applications of this ultrasound-enhanced thrombolysis (UET) include the treatment of myocardial infarction, acute ischemic stroke, deep venous thrombosis and other thrombotic disorders. It may be possible to reduce the dose of rt-PA while maintaining lytic efficacy; however there is little data on the rt-PA concentration dependence of UET. In this work, the rt-PA concentration dependence of clot lysis resulting from 120 kHz UET exposure was measured in an in vitro human clot model. Clots were exposed to rt-PA for 30 min, with (UET treated) or without 120 kHz ultrasound (rt-PA treated) at 37 degrees C, and the clot width measured as a function of time. The rt-PA concentration ranged from 0-10 microg/mL. The initial lytic rate for the UET-treated group was greater than that of the rt-PA group at almost all rt-PA concentrations, and exhibited a maximum over concentration values of 1-3 microg/mL.     

Enhanced Clot Dissolution In Vitro by 1.8-MHz Pulsed Ultrasound

Clinical studies of acute stroke patients have shown that the use of high frequency, low energy transcranial “diagnostic” ultrasound (US) enhances thrombolysis (sonothrombolysis). In contrast, a previous in vitro study using a clot preparation with coagulation induced by recalcification failed to reproduce an effect of “diagnostic” transcranial US on thrombolysis. We sought to evaluate this contradiction in an in vitro model with modified clot preparation. The efficacy of 1.8-MHz pulsed-wave (PW) ultrasound (US) emitted by a commercial probe on thrombolysis was tested. Whole blood clots from 0.5-mL venous blood samples were insonated for 1 h through a human temporal bone, using 1.8-MHz PW US emitted by a diagnostic device. The experiment was performed with or without recombinant tissue-type plasminogen activator (rt-PA) at a concentration of 10 μg/mL. Thrombolysis was measured by means of clot weight loss after 1 h of insonation. A reduction in thrombus weight occurred when US was used in combination with rt-PA, compared with rt-PA alone (78.7% ± 2.1% versus 70.8% ± 4.1%, p ≤ 0.0001). Repetition of the experiment produced identical results (76.9% ± 2.5%, compared with control, p = 0.001). Even without rt-PA, US was effective (41.0% ± 1.7% versus 36.7% ± 3.7%, p = 0.04). The results of this in vitro study support the clinical observation that diagnostic transcranial US, with or without rt-PA, enhances thrombolysis. (E-mail: juergeneggers@gmx.net)     

Accelerating effects of ultrasonic thrombolysis with bubble liposomes

Purpose The accelerating effect on thrombolysis by combined use of 500-kHz low-frequency ultrasound (US), recombinant tissue plasminogen activator (rt-PA), and bubble liposomes (BLs) was verified in vitro. Methods A fibrin clot was formed by adding thrombin to bovine plasma. It was enclosed in a pressurized container, the pressure and temperature of which were maintained at 150 mmHg and 37°C, respectively. Ultrasonic conditions were set at a continuous wave, a frequency of 500 kHz, an intensity of 0.7 W/cm², and a sonication time of 60 s. We derived the rate of reduction in clot weight from the decreased clot weight and the weight before sonication. We compared the rate of reduction in groups combining physiological saline, rt-PA, BLs, and US. Results Only the rt-PA+US+BL group showed a significantly accelerated thrombolytic effect compared with any other group or with any combination of two factors in the 60-s period (0.001 < P < 0.027). Conclusion BLs have great potential to accelerate the thrombolytic effect of rt-PA with low-frequency, 500-kHz, continuous-wave ultrasound.     

In Vitro Thrombolytic Efficacy of Single- and Five-Cycle Histotripsy Pulses and rt-PA

Although primarily known as an ablative modality, histotripsy can increase the efficacy of lytic therapy in a retracted venous clot model. Bubble cloud oscillations are the primary mechanism of action for histotripsy, and the type of bubble activity is dependent on the pulse duration. A retracted human venous clot model was perfused with and without the thrombolytic recombinant tissue plasminogen activator (rt-PA). The clot was exposed to histotripsy pulses of single- or five-cycle duration and peak negative pressures of 0–30 MPa. Bubble activity within the clot was monitored via passive cavitation imaging. The combination of histotripsy and rt-PA was more efficacious than rt-PA alone for single- and five-cycle pulses with peak negative pressures of 25 and 20 MPa, respectively. For both excitation schemes, the detected acoustic emissions correlated with the degree of thrombolytic efficacy. These results indicate that rt-PA and single- or multicycle histotripsy pulses enhance thrombolytic therapy.