P2Y12 in Atherothrombosis Figure 3. Deficiency in mouse P2Y12 receptors diminishes thrombus formation on collagen and provokes disaggregation under flow. Blood from P2Y12 deficient (P2Y122/2) or wild type (P2Y12+/+) mice was perfused over native type I collagen at 1000 s21 for 4 min in the presence of 10 mM ADP. (A) Quantitative effect of P2Y12 deficiency on surface area coverage by platelets. (B) Average number of disaggregation events measured from a preformed aggregate during flow (n = 3). (C) Flow-cytometric evaluation of ADP-stimulated platelets with active aIIbb3 (JON/ A mAb) and of convulxin-stimulated platelets exposing P-selectin (anti-CD62 mAb) in whole blood. Experiments were performed as in Fig. 2. Data are means 6 SE (n = 5–6), *p,0.05. doi:10.1371/journal.pone.0010130.g003 vitro. Furthermore, they show that the contribution of P2Y12 to thrombus stability increases with shear rate and is maintained in the presence of coagulation. Discussion In the present paper, we used a novel in vivo mouse model of thrombus formation on acutely ruptured atherosclerotic plaques, to demonstrate a marked antithrombotic effect of two reversible blocking agents of the platelet P2Y12 receptors, i.e. ticagrelor and cangrelor. Both P2Y12 inhibitors destabilized thrombus formation by increasing the embolization of platelets, with as a result smaller sized thrombi. Our data extend earlier findings, using the same model of acute atherothrombosis, which indicated that thrombus formation is the result of a multi-factorial process involving: platelet interaction with collagen, exposure of tissue factor, generation of thrombin, and formation of fibrin fibers [26]. Recently, other authors have confirmed the importance of collagen and thrombin, using the same model of carotid plaque rupture by ultrasound [30]. Since these in vivo data pointed to collagen as a major thrombogenic component in the carotid plaque, we continued to study thrombus formation by in vitro perfusion studies with immobilized plaque material or native collagen as a surface. However, it should be noted that also other thrombogenic components have been identified in plaque PLoS ONE | www.plosone.org 7 April 2010 | Volume 5 | Issue 4 | e10130