From School of Physiology & Pharmacology, University of Bristol, United Kingdom (E.O.A., M.T.J.v.d.B., E.B., C.M.W., I.H., A.W.P.; Department of Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), University of Maastricht, The Netherlands (N.J.A.M., J.M.E.M.C., J.W.M.H.); and Welsh Blood Service and Arthur Bloom Haemophilia Centre, School of Medicine, Cardiff University, United Kingdom (P.W.C.).
Circulation. 2015 Oct 13;132(15):1414-24. doi: 10.1161/CIRCULATIONAHA.114.015036. Epub 2015 Sep 1.
Platelets are central to the process of hemostasis, rapidly aggregating at sites of blood vessel injury and acting as coagulation nidus sites. On interaction with the subendothelial matrix, platelets are transformed into balloonlike structures as part of the hemostatic response. It remains unclear, however, how and why platelets generate these structures. We set out to determine the physiological relevance and cellular and molecular mechanisms underlying platelet membrane ballooning.
Using 4-dimensional live-cell imaging and electron microscopy, we show that human platelets adherent to collagen are transformed into phosphatidylserine-exposing balloonlike structures with expansive macro/microvesiculate contact surfaces, by a process that we termed procoagulant spreading. We reveal that ballooning is mechanistically and structurally distinct from membrane blebbing and involves disruption to the platelet microtubule cytoskeleton and inflation through fluid entry. Unlike blebbing, procoagulant ballooning is irreversible and a consequence of Na(+), Cl(-), and water entry. Furthermore, membrane ballooning correlated with microparticle generation. Inhibition of Na(+), Cl(-), or water entry impaired ballooning, procoagulant spreading, and microparticle generation, and it also diminished local thrombin generation. Human Scott syndrome platelets, which lack expression of Ano-6, also showed a marked reduction in membrane ballooning, consistent with a role for chloride entry in the process. Finally, the blockade of water entry by acetazolamide attenuated ballooning in vitro and markedly suppressed thrombus formation in vivo in a mouse model of thrombosis.
Ballooning and procoagulant spreading of platelets are driven by fluid entry into the cells, and are important for the amplification of localized coagulation in thrombosis.
血小板是止血过程的核心，在血管损伤部位迅速聚集，并作为凝血核心部位发挥作用。血小板与血管内皮基质相互作用后，会转化为气球样结构，这是止血反应的一部分。然而，血小板如何以及为何会产生这些结构仍不清楚。我们着手确定血小板膜气球样形成的生理相关性以及细胞和分子机制。
通过 4 维活细胞成像和电子显微镜，我们显示人血小板黏附于胶原蛋白后，通过我们称之为促凝扩散的过程，转化为暴露磷脂酰丝氨酸的气球样结构，具有扩张的巨/小微泡接触表面。我们揭示了气球样形成在机制和结构上与膜起泡不同，涉及血小板微管细胞骨架的破坏和通过流体进入而膨胀。与起泡不同，促凝扩散是不可逆的，是 Na(+)、Cl(-)和水进入的结果。此外，膜气球样形成与微粒生成相关。抑制 Na(+)、Cl(-)或水进入会损害气球样形成、促凝扩散和微粒生成，并减少局部凝血酶生成。缺乏 Ano-6 表达的人类 Scott 综合征血小板也表现出明显的膜气球样形成减少，表明氯离子内流在该过程中起作用。最后，通过乙酰唑胺阻断水进入，可在体外抑制气球样形成，并在体内血栓形成的小鼠模型中显著抑制血栓形成。
血小板的气球样形成和促凝扩散是由细胞内流体进入驱动的，对血栓形成中局部凝血的放大很重要。
