Background-Ultrasound at frequencies of 0.5 to 1 MHz and intensities of Ն0.5 W/cm 2 accelerates enzymatic fibrinolysis in vitro and in some animal models, but unacceptable tissue heating can occur, and limited penetration would restrict application to superficial vessels. Tissue heating is less and penetration better at lower frequencies, but little information is available regarding the effect of lower-frequency ultrasound on enzymatic fibrinolysis. We therefore examined the effect of 40-kHz ultrasound on fibrinolysis, tissue penetration, and heating. Methods and Results-125 I-fibrin-radiolabeled plasma clots in thin-walled tubes were overlaid with plasma containing tissue plasminogen activator (tPA) and exposed to ultrasound. Enzymatic fibrinolysis was measured as solubilization of radiolabel. Tissue attenuation and heating were examined in samples of porcine rib cage. Fibrinolysis was increased significantly in the presence of 40-kHz ultrasound at 0.25 W/cm 2 , reaching 39Ϯ7% and 93Ϯ11% at 60 minutes and 120 minutes, compared with 13Ϯ8% and 37Ϯ4% in the absence of ultrasound (PϽ0.0001). The acceleration of fibrinolysis increased at higher intensities. Attenuation of the ultrasound field was only 1.7Ϯ0.5 dB/cm through the intercostal space and 3.4Ϯ0.9 dB/cm through rib. Temperature increments in rib were Ͻ1°C/(W/cm 2 ). Conclusions-These findings indicate that 40-kHz ultrasound significantly accelerates enzymatic fibrinolysis at intensities of Ն0.25 W/cm 2 with excellent tissue penetration and minimal heating. Externally applied 40-kHz ultrasound at low intensities is a potentially useful therapeutic adjunct to enzymatic fibrinolysis with sufficient tissue penetration for both peripheral vascular and coronary applications.

Background —We have shown previously that 40-kHz ultrasound (US) at low intensity accelerates fibrinolysis in vitro with little heating and good tissue penetration. These studies have now been extended to examine the effects of 40-kHz US on thrombolysis and tissue perfusion in a rabbit model. Methods and Results —Treatment was administered with either US alone at 0.75 W/cm 2 , streptokinase alone, or the combination of US and streptokinase. US or streptokinase resulted in minimal thrombolysis, but reperfusion was nearly complete with the combination after 120 minutes. US also reversed the ischemia in nonperfused muscle in the absence of arterial flow. Tissue perfusion decreased after thrombosis from 13.7±0.2 to 6.6±0.8 U and then declined further to 4.5±0.4 U after 240 minutes. US improved perfusion to 10.6±0.5 and 12.1±0.5 U after 30 and 60 minutes, respectively. This effect was reversible and declined to pretreatment values after US was discontinued. Similarly, tissue pH declined from normal to 7.05±0.02 after thrombosis, but US improved pH to 7.34±0.03 after 60 minutes. US-induced improvement in tissue perfusion and pH also occurred after femoral artery ligation, indicating that thrombolysis did not cause these effects. Conclusions —40-kHz US at low intensity markedly accelerates fibrinolysis and also improves tissue perfusion and reverses acidosis, effects that would be beneficial in treatment of acute thrombosis.

SummaryUltrasound accelerates enzymatic fibrinolysis in vitro and in animal models and may be used as an adjunct to thrombolytic therapy. Ultrasound can also affect vascular tone directly, and we have now investigated the effect of ultrasound on tissue perfusion in a rabbit model of acute muscle ischemia to characterize the magnitude and temporal course of vasodilation and determine its mechanism. After ligation of the femoral artery of rabbits, tissue perfusion in the gracilis muscle as determined using a laser Doppler probe declined by 53% from 13.7 ± 0.3 U to 6.4 ± 0.2 U. The tissue became acidotic as pH declined from normal to 7.05 ± 0.2. Application of 40 kHz ultrasound at intensities from 0.25 to 0.75 W/cm2 progressively improved perfusion over 60 min and reversed acidosis, but these effects were both completely blocked by pre-treatment with the nitric oxide synthase inhibitor LNAME. Nitric oxide synthase activity in muscle was measured using an assay based on the conversion of radiolabeled L-arginine to L-citrulline and demonstrated an increase of 3.6-fold following ultrasound exposure. This effect was greatest at locations close to the transducer and declined progressively away from it. Histologic examination showed greater capillary circumference in ultrasound exposed muscle compared to unexposed tissue with no other histologic changes. We conclude that the application of 40 kHz at low intensity improves perfusion and reverses acidosis in acutely ischemic muscle through a nitric oxide dependent mechanism.

Low-frequency, low-intensity ultrasound improves myocardial tissue perfusion and pH in the presence of a fixed coronary artery occlusion.

Summary. Low-intensity ultrasound (US) increases tissue perfusion in ischemic muscle through a nitric oxide (NO)-dependent mechanism. We have developed a model to expose endothelial cells to well-characterized acoustic fields in vitro and investigate the physical and biological mechanisms involved. Human umbilical vein endothelial cells (HUVEC) or bovine aortic endothelial cells (BAEC) were grown in tissue culture plates suspended in a temperaturecontrolled water bath and exposed to US. Exposure to 27 kHz continuous wave US at 0.25 W cm )2 for 10 min increased HUVEC media NO by 102 ± 19% (P < 0.05) and BAEC by 117 ± 23% (P < 0.01). Endothelial cell NO synthase activity increased by 27 ± 24% in HUVEC and by 32 ± 16% in BAEC (P < 0.05 for each). The cell response was rapid with a significant increase in NO synthesis by 10 s and a maximum increase after exposure for 1 min. By 30 min post-exposure NO synthesis declined to baseline, indicating that the response was transient.Unexpectedly, pulsing at a 10% duty cycle resulted in a 46% increase in NO synthesis over the response seen with continuous wave US, resulting in an increase of 147 ± 18%. Cells responded to very low intensity US, with a significant increase at 0.075 W cm )2 (P < 0.01) and a maximum response at 0.125 W cm )2. US caused minor reversible changes in cell morphology but did not alter proliferative capacity, indicating absence of injury. We conclude that exposure of endothelial cells to low-intensity, low-frequency US increases NO synthase activity and NO production, which could be used to induce vasodilatation experimentally or therapeutically.

No abstract

Thrombolytic therapy and mechanical interventions are frequently used in the treatment of both arterial and venous thrombotic disease. Limitations to these approaches include failure to achieve reperfusion and complications including bleeding and vessel wall damage. Increasing evidence indicates that the use of ultrasound offers potential therapeutic advantages. This review considers two distinct approaches which include the use of high intensity ultrasound to mechanically fragment clots and also the use of low intensity ultrasound to augment enzymatic fibrinolysis. High intensity ultrasound can be delivered via catheter or transcutaneously to disrupt clots in vitro or in animal models into small fragments. Initial clinical studies demonstrate potential clinical value in peripheral and coronary arterial thrombosis and occluded saphenous vein bypass grafts treated with the catheter approach. Studies in vitro indicate that low intensity ultrasound accelerates enzymatic thrombolysis through non-thermal mechanisms involving improvement in drug transport. The effect is larger at low frequencies, which also offer better tissue penetration and less heating. The ability to accelerate thrombolysis has been confirmed in animal models demonstrating markedly increased reperfusion and minimal toxicity. The use of ultrasound to mechanically disrupt occlusive thrombi or to accelerate enzymatic thrombolysis offers a new approach to treating occlusive thrombotic disease.

No abstract