© Schattauer 2013 Thrombosis and Haemostasis 109.1/2013 102 Platelets and Blood Cells Analysing responses to aspirin and clopidogrel by measuring platelet thrombus formation under arterial flow conditions Kazuya Hosokawa 1,3 ; Tomoko Ohnishi 1 ; Hisayo Sameshima 1 ; Naoki Miura 2 ; Takashi Ito 3 ; Takehiko Koide 3 ; Ikuro Maruyama 3 1 Research Institute, Fujimori Kogyo Co., Ltd., Yokohama, Kanagawa, Japan; 2 Department of Veterinary Medicine, Faculty of Agriculture, Kagoshima University, Kagoshima, Japan; 3 Department of System Biology in Thromboregulation, Graduate School of Medical and Dental Sciences, Kagoshima University, Kagoshima, Japan Summary High residual platelet aggregability and circulating platelet-monocyte aggregates in patients administered aspirin and clopidogrel are as- sociated with ischaemic vascular events. To determine the relevance of these factors with residual thrombogenicity, we measured platelet thrombus formation using a microchip-based flow-chamber system in cardiac patients receiving aspirin and/or clopidogrel, and evaluated its correlation with agonist-inducible platelet aggregation and platelet- monocyte aggregates. Platelet thrombus formation was analysed by measuring flow pressure changes due to the occlusion of micro-capil- laries and was quantified by calculating AUC 10 (area under the flow pressure curve). The growth and stability of platelet thrombi that formed inside microchips at shear rates of 1000, 1500, and 2000 s -1 were markedly reduced in patients receiving aspirin and/or thienopyri- dine compared to healthy controls (n=33). AUC 10 values of aspirin therapy patients (n=20) were significantly lower and higher than those of healthy controls and dual antiplatelet therapy patients (n=19), respectively, and showed relatively good correlations with col- lagen-induced platelet aggregation and platelet-monocyte aggre- gates at 1000 and 1500 s -1 (r s >0.59, p<0.01). In contrast, AUC 10 in dual antiplatelet therapy patients was significantly correlated with ADP-induced platelet aggregation at all examined shear rates (r s >0.59, p<0.01), but did not correlate with collagen-induced aggre- gation. Aspirin monotherapy patients with high residual platelet thrombogenicity also exhibited significant elevations in both collagen- induced platelet aggregation and platelet-monocyte aggregates. Our results, although preliminary, suggest that residual platelet thrombo- genicity in aspirin-treated patients is associated with either collagen- induced platelet aggregation or circulating platelet-monocyte aggre- gates, but it is predominantly dependent on ADP-induced platelet ag- gregation in patients receiving dual antiplatelet therapy. Keywords Antiplatelet therapy, aspirin, clopidogrel, platelet thrombus, blood flow Correspondence to: Ikuro Maruyama, MD, PhD Kagoshima University 8–35–1, Sakuragaoka, Kagoshima, Japan Tel.: +81 99 275 6474, Fax: +81 99 275 6463 E-mail: rinken@m3.kufm.kagoshima-u.ac.jp Received: June 27, 2012 Accepted after major revision: September 19, 2012 Prepublished online: November 22, 2012 doi:10.1160/TH12-06-0441 Thromb Haemost 2013; 109: 102–111 Introduction Aspirin and thienopyridines (clopidogrel and ticlopidine), which inhibit thromboxane A 2 (TXA 2 ) synthesis and the P2Y 12 receptor, respectively, are the most widely used antiplatelet agents for the secondary prevention of coronary heart disease. These drugs are frequently administered in combination (dual antiplatelet therapy) for long-term treatment after percutaneous coronary intervention (PCI) and significantly improve patient outcomes (1, 2). However, despite the well-demonstrated benefits in several trials, consider- able numbers of patients receiving aspirin and thienopyridines ex- perience recurrent vascular events, even with dual antiplatelet therapy. In addition, several studies have shown that low sensitiv- ity to aspirin and clopidogrel, as determined by platelet function tests, is associated with poor therapeutic outcomes (3–7). For monitoring antiplatelet treatment efficacy and identifying low responsiveness to therapy, several assay systems have been de- veloped in the clinical setting, such as the VerifyNow™ (Accumet- rics, San Diego, CA, USA) and Multiplate™ (Dynabyte Medical, Munich, Germany) systems, which measure platelet agglutination and aggregation in response to an exogenous agonist. The PFA-100 test is also capable of quantifying platelet aggregate formation on the surface of collagen coated with a specific agonist under arterial shear conditions. Although these systems are less labor intensive than light transmission aggregometry (LTA) and permit the use of whole blood, the choice of platelet agonist strongly affects the observed platelet activity and inhibitory effects of the antiplatelet agents. In addition, as the ability to identify pa- tients with low responsiveness to antiplatelet agents varies depend- ing on the test used (8–10), consensus methodologies to assess antiplatelet treatment efficacy have not yet been established. Flow cytometry (FCM) analysis is also capable of evaluating circulating activated platelets in vivo, and platelet-monocyte aggre- gate formation and platelet CD62P (P-selectin) expression are re- For personal or educational use only. No other uses without permission. All rights reserved. Downloaded from www.thrombosis-online.com on 2018-04-07 | ID: 1001066444 | IP: 54.70.40.11