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

https://doi.org/10.1016/S0301-5629(00)00211-8Get rights and content

Abstract

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/cm2; 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.

Introduction

Ischemic cerebral stroke is a major vascular disorder with serious neurological consequences, resulting mainly from thromboembolic and atherothrombotic events (Fieschi et al. 1989). This provides the basis for the use of antithrombotic agents in acute stroke therapy Hacke et al 1995, NINDS rt-PA Stroke Study Group 1995. Disability and long-term costs of treatment and rehabilitation of stroke are enormous Jorgensen et al 1997, Wein et al 1998, demanding further therapy optimization. Initial ischemic cell damage directly after arterial occlusion is small compared with larger areas of hypoperfusion with functional loss of neurons but intact cell integrity (penumbra). The cellular integrity depends on both level and duration of hypoperfusion. Therefore, early recanalization within hours can save brain tissue Hatazawa et al 1999, Bozzao et al 1989. Although spontaneous partial or complete recanalization occurs in 15–20% of patients (Mori et al. 1992), thrombolytics may further reduce recanalization time of occluded cerebral arteries, resulting in neurological improvement.

Intravenous application of thrombolytic agents, such as recombinant tissue plasminogen activator (rt-PA), can increase the recanalization rate up to 34–47% within the first 24 h after onset of symptoms Mori et al 1992, Del Zoppo et al 1992. Studies of intraarterial local application of rt-PA have reported recanalization rates as high as 90% (Del Zoppo et al. 1988). Even after local thrombolysis, the time until recanalization occurs can take up to several hours Bendszus et al 1998, Del Zoppo et al 1998. This indicates the need for further optimization of thrombolysis. Ultrasound (US) has been proven to have thrombolytic capacity at high intensities, either by itself or to enhance the effect of thrombolytics at relatively lower intensities. Mechanical effects were mainly considered responsible for the direct thrombolytic effect of intravascular applied low-frequency US (19.5–26.5 kHz), which was used for the direct disruption of clots in occluded peripheral Trubestein et al 1976, Siegel et al 1989 and coronary vessels (Steffen et al. 1994) in vitro and in vivo.

Various recent studies described a significant acceleration of rt-PA-, streptokinase- or urokinase-induced thrombolysis by external 1-MHz US of 1–4 W/cm2 intensity Francis et al 1992, Luo et al 1993, Harpaz et al 1994. Experimental findings suggest that US accelerates enzymatic fibrinolysis by increasing transport of reactants through a cavitation-related mechanism. This results in greater penetration in the clot and a broader zone of lysis Francis et al 1995, Siddiqi et al 1995. Heating, detachment or fragmentation are not of etiological relevance Blinc et al 1993, Francis et al 1995. Experimental and clinical studies by means of US thrombolysis, as well as US-accelerated pharmacological thrombolysis, in peripheral and coronary arterial disease have already been conducted. The effects of transcranial US application, however, had been poorly investigated. Transcranial US application was thought to cause high loss of US intensity. Recent studies could show that US at low frequencies, which could be useful for enzymatically mediated thrombolysis, has an acceptable low attenuation through the skull Behrens et al 1999, Akiyama et al 1998. These studies investigated attenuation or reduction of thrombus weight in a static model. To obtain data about US-accelerated recanalization time, we measured reperfusion and recanalization in a flow model.

Section snippets

Flow model

We designed an experimental flow model utilizing a water tank (600 × 400 × 300 mm) with a temperature maintained at 37°C (± 0.1°C), in which a perfusion system, including an obstructing fibrin thrombus developed within a polyethylene tubing system, was submerged. The frame stabilizing the “fibrin clot-containing tubes” was mounted vertically and placed in the low-pressure flow system. The tube proximal to the occluding thrombus was filled with 0.05 M Tris, 0.1 M NaCl, 0.005 M EDTA buffer at a

Results

The mean flow in control group D was 0.01 ± 0.01 mL/min and the mean collected perfusion fluid weighed 0.41 ± 0.39 g. None of the clots were recanalized, even after a total observation time of 60 min.

The clots of group C underwent thrombolysis after application of 100 μg rt-PA without exposure to US. The perfusion flow during the first 15 min remained stable and became accelerated in the second half of the “protocol-defined” observation phase (Fig. 2). The mean flow was 0.45 ± 0.61 mL/min

Discussion

This study indicates that externally applied, 1-MHz, pulsed-wave US with a remarkably low peak intensity accelerates rt-PA-mediated thrombolysis, even through the human skull, in a flow model in vitro. US exposure significantly increases flow, perfusion fluid weight and reperfusion rate. Under ultrasonication, recanalization time is also significantly shortened. The source of US was a commercially available and clinically used standard Doppler probe. Our findings suggest that ultrasonication

Acknowledgements

The authors acknowledge the technical assistance of Mrs. Anja Kirchner and the help of Mrs. M.Garcia-Knapp in preparation of the manuscript.This work has been supported by the BioRegio Foundation Rhein-Neckar.

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