Department of Chemical and Biomolecular Engineering, Institute for Medicine and Engineering, University of Pennsylvania, Philadelphia, Pennsylvania, United States.
Thromb Haemost. 2021 Jan;121(1):46-57. doi: 10.1055/s-0040-1715648. Epub 2020 Sep 22.
As thrombosis proceeds, certain platelets in a clot expose phosphatidylserine (PS) on their outer membrane. These PS platelets subsequently sort to the perimeter of the mass via platelet contraction. It remains unclear how thrombin and fibrin may alter PS platelet sorting within a clot.
We investigated the role of fibrin in PS platelet sorting.
We used an 8-channel microfluidic assay of clotting over collagen (±tissue factor) at 100 s initial wall shear rate. Temporal PS platelet sorting was measured using a Pearson's correlation coefficient between the annexin V distribution in a clot at 9 versus 15 minutes. Spatial PS platelet sorting was measured using an autocorrelation metric of the final annexin V distribution.
By 6 minutes, PS platelets were distributed throughout the platelet deposits and became highly spatially sorted by 15 minutes when thrombin and fibrin were blocked with Phe-Pro-Arg-chloromethylketone (PPACK). Fibrin polymerization (no PPACK) attenuated temporal and spatial PS sorting and clot contraction. With Gly-Pro-Arg-Pro (GPRP) added to block fibrin polymerization, PS sorting was prominent as was clot contraction. Exogenously added tissue plasminogen activator drove fibrinolysis that in turn promoted clot contraction and PS sorting, albeit to a lesser degree than the PPACK or GPRP conditions. Clots lacking fibrin displayed 3.6 times greater contraction than clots with fibrin.
PS sorting correlated with clot contraction, as previously reported. However, fibrin inversely correlated with both percent contraction and PS sorting. Fibrin attenuated clot contraction and PS sorting relative to clots without fibrin.
随着血栓的形成，血栓中的某些血小板会在其外膜上暴露磷脂酰丝氨酸（PS）。这些 PS 血小板随后通过血小板收缩作用向血栓的周边分拣。目前尚不清楚凝血酶和纤维蛋白如何改变血栓内 PS 血小板的分拣。
我们研究了纤维蛋白在 PS 血小板分拣中的作用。
我们使用了一种在 100 秒初始壁剪切率下在胶原上（±组织因子）凝固的 8 通道微流控测定法。使用在 9 分钟和 15 分钟时血栓中 annexin V 分布之间的 Pearson 相关系数来测量 PS 血小板的时间分拣。使用 annexin V 最终分布的自相关度量来测量 PS 血小板的空间分拣。
在 6 分钟时，PS 血小板分布在血小板沉积物中，并在凝血酶和纤维蛋白被 Phe-Pro-Arg-chloromethylketone（PPACK）阻断时在 15 分钟时变得高度空间分拣。纤维蛋白聚合（无 PPACK）减弱了 PS 的时间和空间分拣以及血栓收缩。用 Gly-Pro-Arg-Pro（GPRP）添加物来阻断纤维蛋白聚合，PS 分拣明显，血栓收缩也明显。外源性添加组织纤溶酶原激活剂会导致纤维蛋白溶解，进而促进血栓收缩和 PS 分拣，但程度不及 PPACK 或 GPRP 条件。缺乏纤维蛋白的血栓比有纤维蛋白的血栓收缩程度大 3.6 倍。
PS 分拣与血栓收缩相关，如前所述。然而，纤维蛋白与收缩百分比和 PS 分拣呈负相关。纤维蛋白相对于无纤维蛋白的血栓减弱了血栓收缩和 PS 分拣。
