In Vivo Dynamic Real-Time Monitoring and Quantification of Platelet-Thrombus Formation

Abstract

Abstract-Current methods for monitoring thrombosis and thrombus growth are invasive and provide only single-timepoint data. Animal models rely mainly on flow changes as a surrogate of thrombus formation. Our aim was to validate a unique potentially noninvasive system to detect and quantify dynamic thrombus formation in vivo by using a porcine model of carotid artery injury. Thrombus growth was monitored by deposition of autologous 111 In-labeled platelet activity over the injured artery by use of miniaturized gamma detectors and Doppler blood flow. Counts were recorded at 2-minute intervals for 2 hours. The technique was validated by comparing standard antithrombotic agents against controls. Platelet recruitment was detected before significant change in flow. Thrombus formation, calculated as the area under the curve (plateletsϫminutesϫ10 6 ), was greatest for control animals (11.7Ϯ1.28), followed by animals treated with aspirin (6.13Ϯ0.91, PϽ0.05), heparin (2.45Ϯ0.34, PϽ0.05), and hirudin (0.2Ϯ0.01, PϽ0.01 compared with heparin). The rate of platelet deposition was assessed as the slope of the curve in the first 30 minutes (plateletsϫ10 6 per minute) for the following treatment groups of animals: control, 3.53Ϯ0.34; aspirin, 1.67Ϯ0.34 (PϽ0.01); heparin, 1.55Ϯ0.3 (PϽ0.01); and hirudin, 0.25Ϯ0.03 (PϽ0.001). There was no statistical difference between heparin and aspirin treatments. Change in flow was assessed as reduction from baseline: control, Ͼ99Ϯ0.34%; aspirin, 39Ϯ9.1%; heparin, 36Ϯ12.5%; and hirudin, 17Ϯ5.4%. There was no statistical difference between the aspirin-and heparin-treated groups. Morphometric analysis revealed Ͼ99Ϯ0.63% occlusion of the luminal area with thrombus for the control group, 43Ϯ14.3% for the aspirin-treated group, 30Ϯ5.6% for the heparin-treated group, and Ͻ10Ϯ1.8% for the hirudin-treated group. Assessment of platelet-thrombus formation with this technique was more sensitive than change in flow in determining antithrombotic efficacy, and thrombus formation was detected earlier. This study validates a new quantitative, sensitive, potentially noninvasive, portable, in vivo monitoring of dynamic thrombus growth, which appears applicable to phase II studies in humans. Key Words: platelets Ⅲ thrombus Ⅲ antithrombotics Ⅲ radioisotopes P latelet-vessel wall interaction and thrombus formation are ongoing dynamic processes. The extent of thrombus formation at the site of arterial injury (whether spontaneous or induced), in addition to acute complications, has long-term consequences because it promotes neointimal proliferation and restenosis. 1,2 Current methods for the diagnosis of thrombus formation in the clinical setting include intravascular ultrasound, angiography, and angioscopy. All of these are invasive and qualitative, and the results are limited to a single time point of an ongoing dynamic process. Furthermore, their invasive nature does not permit the continuous and quantitative monitoring of thrombus growth or assess the efficacy of treatment over time. Noninvasive methods such as v...

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