Center for Thrombosis and Hemostasis, University Medical Center Mainz, Mainz, Germany.
Sechenov Institute of Evolutionary Physiology and Biochemistry, Russian Academy of Sciences, St. Petersburg, Russia.
Thromb Haemost. 2019 Jun;119(6):916-929. doi: 10.1055/s-0039-1685139. Epub 2019 Apr 20.
The direct thrombin inhibitor (DTI) dabigatran is a non-vitamin K antagonist oral anticoagulant for the prevention of stroke and systemic embolism in patients with non-valvular atrial fibrillation. In addition to its anti-thrombotic efficacy, dabigatran has been suggested to exert some pro-thrombotic effect due to fostering the ligation of thrombin to its high affinity platelet receptor glycoprotein (GP) Ibα in patients with atrial fibrillation. On the other hand, we provided evidence that a member of another class of DTIs, lepirudin, stimulates the inhibitory cyclic guanosine monophosphate (cGMP)/soluble guanylate cyclase pathway in human platelets. Here, we investigated the effect of lepirudin and dabigatran spiked to platelets from healthy volunteers on GPIbα-mediated platelet aggregation and agglutination. Ristocetin/von Willebrand factor (vWF)-induced aggregation of platelets in the presence or absence of plasma was significantly inhibited by lepirudin, dabigatran and D-phenylalanyl-L-prolyl-L-arginine chloromethyl ketone (PPACK). However, ristocetin/vWF-mediated platelet agglutination and binding of vWF to platelets were not affected by the DTIs. The anti-aggregatory effect was confirmed by using the GPIbα-specific agonist echicetin beads for human and murine platelets. DTIs diminished echicetin beads-induced Syk Y352 phosphorylation (used here as readout for an early signal occurring during echicetin-induced platelet aggregation), but did not inhibit adenosine diphosphate- or thromboxane A2-induced platelet aggregation. Thrombin was not generated in response to ristocetin/vWF or echicetin beads and therefore did not explain the inhibitory effect of the DTIs. Therapeutic concentration of lepirudin and dabigatran did not affect significantly platelet vasodilator-stimulated phosphoprotein S239 phosphorylation or cGMP and cyclic adenosine monophosphate levels. These data suggest that the DTIs, lepirudin and dabigatran, impair platelet activation measured during platelet aggregation induced by ristocetin/vWF or echicetin beads.
直接凝血酶抑制剂（DTI）达比加群是一种非维生素 K 拮抗剂口服抗凝剂，用于预防非瓣膜性心房颤动患者的中风和全身性栓塞。除了其抗血栓形成作用外，达比加群由于在心房颤动患者中促进凝血酶与高亲和力血小板受体糖蛋白（GP）Ibα的结合，因此被认为具有一定的促血栓形成作用。另一方面，我们提供了证据表明，另一种 DTI 类别的成员 lepirudin，刺激人血小板中抑制性环鸟苷单磷酸（cGMP）/可溶性鸟苷酸环化酶途径。在这里，我们研究了 lepirudin 和 dabigatran 对来自健康志愿者血小板的 GPIbα 介导的血小板聚集和凝集的影响。在存在或不存在血浆的情况下，ristocetin/von Willebrand 因子（vWF）诱导的血小板聚集被 lepirudin、dabigatran 和 D-苯丙氨酰-L-脯氨酰-L-精氨酰氯甲基酮（PPACK）显著抑制。然而，ristocetin/vWF 介导的血小板凝集和 vWF 与血小板的结合不受 DTI 的影响。通过使用 GPIbα 特异性激动剂 echicetin 珠子用于人源和鼠源血小板，证实了抗聚集作用。DTI 降低了 echicetin 珠子诱导的 Syk Y352 磷酸化（这里用作 echicetin 诱导的血小板聚集过程中早期信号的读数），但不抑制二磷酸腺苷或血栓烷 A2 诱导的血小板聚集。对 ristocetin/vWF 或 echicetin 珠子没有生成凝血酶，因此不解释 DTI 的抑制作用。治疗浓度的 lepirudin 和 dabigatran 对血小板血管舒张刺激磷酸蛋白 S239 磷酸化或 cGMP 和环腺苷单磷酸水平的影响没有显著影响。这些数据表明，DTI lepirudin 和 dabigatran 可损害由 ristocetin/vWF 或 echicetin 珠子诱导的血小板聚集期间测量的血小板激活。
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