Donadelli Roberta, Orje Jennifer N, Capoferri Cristina, Remuzzi Giuseppe, Ruggeri Zaverio M
Department of Molecular and Experimental Medicine, Division of Experimental Hemostasis and Thrombosis, Roon Center for Arteriosclerosis and Thrombosis, The Scripps Research Institute-MEM 175, 10550 North Torrey Pines Rd, La Jolla, CA 92037, USA.
Blood. 2006 Mar 1;107(5):1943-50. doi: 10.1182/blood-2005-07-2972. Epub 2005 Nov 17.
The metalloproteinase ADAMTS13 regulates the size of released von Willebrand factor (VWF) multimers bound to endothelial cells, but it is unknown whether it can cleave plasma VWF during thrombogenesis. To address this issue, we perfused blood over immobilized VWF and used videomicroscopy to visualize an activation-independent platelet aggregation process mediated by soluble VWF at shear rates greater than 10 000 s(-1). At normal Ca2+ concentration, platelets formed rolling as well as surface-attached clusters that grew larger during the first 5 minutes but then lost more than 70% of their mass by 10 minutes. In contrast, platelet clusters were stable in size when metal ions were chelated, anti-ADAMTS13 IgG were added, or washed blood cells were perfused with purified VWF but no plasma. In the latter case, addition of recombinant ADAMTS13 reduced platelet cluster size by more than 70%. Incubating ADAMTS13 with VWF before perfusion did not prevent the initial platelet clustering, indicating that the enzyme may act on platelet-bound VWF under shear stress. At the concentrations tested, ADAMTS13 had no effect on platelet aggregates formed upon blood perfusion over collagen fibrils. ADAMTS13, therefore, may regulate thrombus size preferentially when the cohesion between platelets depends on VWF binding induced by pathologically elevated shear stress.
金属蛋白酶ADAMTS13可调节与内皮细胞结合的血管性血友病因子（VWF）多聚体的释放大小，但在血栓形成过程中它是否能裂解血浆VWF尚不清楚。为解决这一问题，我们将血液灌注到固定化的VWF上，并使用视频显微镜观察在剪切速率大于10000 s(-1)时由可溶性VWF介导的与激活无关的血小板聚集过程。在正常Ca2+浓度下，血小板形成滚动以及表面附着的聚集体，这些聚集体在最初5分钟内会变大，但到10分钟时会损失超过70%的质量。相比之下，当金属离子被螯合、加入抗ADAMTS13 IgG或用纯化的VWF而非血浆灌注洗涤过的血细胞时，血小板聚集体的大小保持稳定。在后一种情况下，加入重组ADAMTS13可使血小板聚集体大小减少超过70%。在灌注前将ADAMTS13与VWF一起孵育并不能阻止最初的血小板聚集，这表明该酶可能在剪切应力下作用于与血小板结合的VWF。在所测试的浓度下，ADAMTS13对血液灌注到胶原纤维上时形成的血小板聚集体没有影响。因此，当血小板之间的凝聚力取决于病理性升高的剪切应力诱导的VWF结合时，ADAMTS13可能优先调节血栓大小。
