Department of Clinical Chemistry and Haematology, University Medical Centre Utrecht, Utrecht, The Netherlands.
Thromb Haemost. 2010 May;103(5):1033-43. doi: 10.1160/TH09-11-0751. Epub 2010 Mar 9.
Incubation at 0 degrees C is known to expose b- N -acetyl-D-glucosamine residues on glycoprotein (GP) Ibalpha inducing receptor clustering and alpha(M)beta(2)-mediated platelet destruction by macrophages. Here we show that incubation at 0/37 degrees C (4 hours at 0 degrees C, followed by 1 hour at 37 degrees C to mimic cold-storage and post-transfusion conditions) triggers a conformational change in the N -terminal flank (NTF, amino acids, aa 1-35) but not in aa 36-282 of GPIbalpha as detected by antibody binding. Addition of the sugar N -acetyl-D-glucosamine (GN) inhibits responses induced by 0/37 degrees C. Incubation at 0 degrees C shifts GPIbalpha from the membrane skeleton to the cytoskeleton. Different GPIbalpha conformations have little effect on VWF/ristocetin-induced aggregation, but arrest of NTF change by GN interferes with agglutination and spreading on a VWF-coated surface under flow. Strikingly, incubation at 0/37 degrees C initiates thromboxane A(2) formation through a von Willebrand factor (VWF)-independent and GPIbalpha-dependent mechanism, as confirmed in VWF- and GPIbalpha-deficient platelets. We conclude that the NTF change induced by 0/37 degrees C incubation reflects clustering of GPIbalpha supports VWF/ristocetin-induced agglutination and spreading and is sufficient to initiate platelet activation in the absence of VWF.
在 0°C 下孵育已知会暴露糖蛋白 (GP) Ibalpha 上的 b-N-乙酰-D-葡萄糖胺残基，从而诱导受体聚集，并通过巨噬细胞介导 α(M)β(2) 介导的血小板破坏。在这里，我们表明，在 0/37°C（0°C 下孵育 4 小时，然后在 37°C 下孵育 1 小时以模拟冷藏和输血条件）会触发 N-末端侧翼（NTF，氨基酸，aa 1-35）但不会触发 GPIbalpha 的 aa 36-282 发生构象变化，如通过抗体结合检测到的。添加糖 N-乙酰-D-葡萄糖胺 (GN) 可抑制 0/37°C 诱导的反应。在 0°C 下孵育会将 GPIbalpha 从膜骨架转移到细胞骨架。不同的 GPIbalpha 构象对 VWF/瑞斯托菌素诱导的聚集几乎没有影响，但通过 GN 阻止 NTF 变化会干扰在流动下 VWF 涂层表面上的聚集和扩展。引人注目的是，在 0/37°C 下孵育会通过一种与 von Willebrand 因子 (VWF) 无关且依赖于 GPIbalpha 的机制引发血栓烷 A(2) 的形成，这在 VWF 和 GPIbalpha 缺陷型血小板中得到了证实。我们得出结论，0/37°C 孵育诱导的 NTF 变化反映了 GPIbalpha 的聚集，支持 VWF/瑞斯托菌素诱导的聚集和扩展，并且足以在没有 VWF 的情况下引发血小板激活。
