Background: Intravenous tissue plasminogen activator (IV-tPA) remains the only drug for arterial recanalization in acute ischemic stroke (AIS). Clot composition differs according to the stroke etiology- clot from heart being fresh and rich in fibrin while the clot from an ulcerated plaque is rich in cholesterol. These characteristics might influence the therapeutic response to tPA as well as ultrasound-enhanced thrombolysis. We evaluated the impact of cholesterol content and age of the clot on its dissolution with tPA and ultrasound in an in-vitro model. Methods: Clot was prepared by mixing horse blood with Calcium-chloride solution. Cholesterin was added to increase cholesterol level up to 100 mg/dL. Periodic serum replacement was used to ‘age’ the clot. The clots were exposed to continuous 2-MHZ diagnostic transcranial Doppler (TCD) ultrasound (acoustic intensity 340mW/cm2) during thromolysis with tPA (355 IU/ml). Efficiency of thrombolysis was calculated as the percentage of clot weight-loss after 60 minutes. Clot characteristics after lysis were evaluated with surface emission microscope (SEM). Results: Significant increase in the weight of clot occurred with its increasing age (from 2 to 10 hours, the clot-weight increased from 37.28±2.87mg to 51.56±5.34mg; p<0.005). Exposure to ultrasound and tPA resulted in significant reduction in the clot-weight at 1 hour (from 35.50±3.23% to 24.71±2.11%; p<0.005). Although, no significant differences were noted between clot-cholesterol levels and lysis with TCD or tPA alone, combination of these modalities induced significant lysis in clots with higher cholesterol levels (clot-weight reduced by 41.68±2.3%) as compared to clots with normal cholesterol (30.60±4.10%; p<0.005). SEM demonstrated thicker and denser fibrin network in aged clots while higher number of activated platelets were noted on the surface of cholesterol-rich clots. Conclusion: Ultrasound-assisted thrombolysis with tPA work better for fresh thrombi with higher cholesterol levels. Our findings provide insights into the differential recanalization rates in acute ischemic stroke of various etiologies

Abstract. Tissue plasminogen activator (tPA) remains the only drug for recanalization in acute ischemic stroke, and the dose is determined by the patient's body-weight. Properties of the blood clot as well as ultrasound exposure might affect the thrombolysis outcome. In this study, clot was prepared by mixing horse blood with CaCl 2 solution and cholesterin up to 1.0 mg/ml. To simulate the aging effect serum was replaced by fresh blood periodically. 225 IU/ml of tPA was used to initiate lysis. Clot was exposed to continuous 2 MHz transcranial Doppler ultrasound at acoustic intensity of 340 mW/cm 2 . The weight of the blood clot increased with its age (from 37.28r2.87 mg at 2 hrs to 51.56r5.34 mg at 10 hrs, p < 0.05). Although no difference between clot-cholesterol levels and thrombolysis with ultrasound or tPA alone was found, combination of these modalities induced significant lysis in the clots with cholesterol levels of more than 0.5 mg/ml (clot-weight reduced by 41.68r2.3%) as compared to clots with normal cholesterol (30.60r4.10%; p < 0.05). Altogether, sonothrombolysis seems to work better in fresh thrombi with highcholesterol levels.