Novel mouse hemostasis model for real-time determination of bleeding time and hemostatic plug composition
Novel mouse hemostasis model for real-time determination of bleeding time and hemostatic plug composition
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DOI：
10.1111/jth.12802
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发表时间：
2015-03-01
影响因子：
10.4
通讯作者：
Bergmeier, W.
中科院分区：
文献类型：
--
作者：
Getz, T. M.;Piatt, R.;Bergmeier, W.
前言止血是人体对血管损伤部位止血的一种快速反应。血小板和纤维蛋白对止血栓的形成都很重要。小鼠已经被用来揭示在生理条件下调节血小板激活和凝血的分子机制。然而，对小鼠止血的测量是非常不同的，并且目前的方法不能量化损伤部位的血小板黏附或纤维蛋白形成。方法我们描述了一种新的止血模型，该模型使用活体荧光显微镜来实时量化血小板黏附、纤维蛋白形成和止血塞形成的时间。用激光消融技术在小鼠的隐静脉造成直径50~100m的反复血管损伤。结果糖蛋白Ib或talin-1缺陷小鼠的止血功能严重受损，而糖蛋白Ib或talin-1是血小板黏附的重要调节因子。相反，在缺乏外源性组织因子(Tflow)或内在凝血因子IX途径的小鼠中，止血栓子形成的时间受到的影响很小，即使血小板粘附性显著降低。氯吡格雷部分降低了血小板的粘附性，导致止血栓内的不稳定性，特别是在Tflow小鼠中，尤其是在凝血功能受损的情况下。结论综上所述，我们提出了一种新的、高灵敏度的方法来定量小鼠止血栓的形成。基于其对血小板黏附缺陷的敏感性和实时成像能力，我们认为该模型是研究抗血小板药物有效性和安全性的理想工具。
IntroductionHemostasis is a rapid response by the body to stop bleeding at sites of vessel injury. Both platelets and fibrin are important for the formation of a hemostatic plug. Mice have been used to uncover the molecular mechanisms that regulate the activation of platelets and coagulation under physiologic conditions. However, measurements of hemostasis in mice are quite variable, and current methods do not quantify platelet adhesion or fibrin formation at the site of injury.MethodsWe describe a novel hemostasis model that uses intravital fluorescence microscopy to quantify platelet adhesion, fibrin formation and time to hemostatic plug formation in real time. Repeated vessel injuries of similar to 50-100m in diameter were induced with laser ablation technology in the saphenous vein of mice.ResultsHemostasis in this model was strongly impaired in mice deficient in glycoproteinIb or talin-1, which are important regulators of platelet adhesiveness. In contrast, the time to hemostatic plug formation was only minimally affected in mice deficient in the extrinsic tissue factor (TFlow) or the intrinsic factorIX coagulation pathways, even though platelet adhesion was significantly reduced. A partial reduction in platelet adhesiveness obtained with clopidogrel led to instability within the hemostatic plug, especially when combined with impaired coagulation in TFlow mice.ConclusionsIn summary, we present a novel, highly sensitive method to quantify hemostatic plug formation in mice. On the basis of its sensitivity to platelet adhesion defects and its real-time imaging capability, we propose this model as an ideal tool with which to study the efficacy and safety of antiplatelet agents.