Differential inhibitory action of apixaban on platelet and fibrin components of forming thrombi: Studies with circulating blood and in a platelet-based model of thrombin generation

IntroductionMechanisms of action of direct oral anticoagulants (DOAC) suggest a potential therapeutic use in the prevention of thrombotic complications in arterial territories. However, effects of DOACs on platelet activation and aggregation have not been explored in detail. We have investigated the effects of apixaban on platelet and fibrin components of thrombus formation under static and flow conditions.MethodsWe assessed the effects of apixaban (10, 40 and 160 ng/mL) on: 1) platelet deposition and fibrin formation onto a thrombogenic surface, with blood circulating at arterial shear-rates; 2) viscoelastic properties of forming clots, and 3) thrombin generation in a cell-model of coagulation primed by platelets.ResultsIn studies with flowing blood, only the highest concentration of apixaban, equivalent to the therapeutic Cmax, was capable to significantly reduce thrombus formation, fibrin association and platelet-aggregate formation. Apixaban significantly prolonged thromboelastometry parameters, but did not affect clot firmness. Interestingly, results in a platelet-based model of thrombin generation under more static conditions, revealed a dose dependent persistent inhibitory action by apixaban, with concentrations 4 to 16 times below the therapeutic Cmax significantly prolonging kinetic parameters and reducing the total amount of thrombin generated.ConclusionsOur studies demonstrate the critical impact of rheological conditions on the antithrombotic effects of apixaban. Studies under flow conditions combined with modified thrombin generation assays could help discriminating concentrations of apixaban that prevent excessive platelet accumulation, from those that deeply impair fibrin formation and may unnecessarily compromise hemostasis.引言 直接口服抗凝剂（direct oral anticoagulants, DOAC）的作用机制提示其在动脉区域血栓并发症预防中具备潜在治疗用途。然而，直接口服抗凝剂对血小板活化与聚集的影响尚未得到详细探究。本研究探究了阿哌沙班（apixaban）在静态及血流条件下对血栓形成过程中血小板与纤维蛋白成分的影响。 方法 本研究评估了浓度分别为10、40及160 ng/mL的阿哌沙班对以下三方面的影响：1）动脉剪切速率下血液流动时，血栓源性表面上的血小板沉积与纤维蛋白形成情况；2）正在形成的血凝块的粘弹性特性；3）由血小板激活凝血的细胞模型中的凝血酶生成水平。 结果 在流动血液相关研究中，仅最高浓度的阿哌沙班（相当于治疗峰浓度Cmax）能够显著抑制血栓形成、纤维蛋白结合及血小板聚集体形成。阿哌沙班可显著延长血栓弹力图（thromboelastometry）参数，但并未影响血凝块硬度。值得注意的是，在更接近静态条件下的基于血小板的凝血酶生成模型中，结果显示阿哌沙班具备剂量依赖性的持续抑制作用：当浓度为治疗峰浓度Cmax的1/4至1/16时，即可显著延长动力学参数并降低凝血酶总生成量。 结论 本研究证实了流变条件对阿哌沙班抗血栓作用的关键影响。结合改良凝血酶生成试验的血流条件下研究，或可用于区分两类阿哌沙班浓度：一类可阻止过度血小板积聚，另一类则会严重损害纤维蛋白形成，且可能不必要地损害止血功能。