Chemical and Biological Engineering, State University of New York, Buffalo, NY 14260, USA.
Blood. 2010 Nov 11;116(19):3990-8. doi: 10.1182/blood-2010-02-269266. Epub 2010 Aug 9.
The function of the mechanosensitive, multimeric blood protein von Willebrand factor (VWF) is dependent on its size. We tested the hypothesis that VWF may self-associate on the platelet glycoprotein Ibα (GpIbα) receptor under hydrodynamic shear. Consistent with this proposition, whereas Alexa-488-conjugated VWF (VWF-488) bound platelets at modest levels, addition of unlabeled VWF enhanced the extent of VWF-488 binding. Recombinant VWF lacking the A1-domain was conjugated with Alexa-488 to produce ΔA1-488. Although ΔA1-488 alone did not bind platelets under shear, this protein bound GpIbα on addition of either purified plasma VWF or recombinant full-length VWF. The extent of self-association increased with applied shear stress more than ∼ 60 to 70 dyne/cm(2). ΔA1-488 bound platelets in the milieu of plasma. On application of fluid shear to whole blood, half of the activated platelets had ΔA1-488 bound, suggesting that VWF self-association may be necessary for cell activation. Shearing platelets with 6-μm beads bearing either immobilized VWF or anti-GpIbα mAb resulted in cell activation at shear stress down to 2 to 5 dyne/cm(2). Taken together, the data suggest that fluid shear in circulation can increase the effective size of VWF bound to platelet GpIbα via protein self-association. This can trigger mechanotransduction and cell activation by enhancing the drag force applied on the cell-surface receptor.
机械敏感的多聚体血液蛋白 von Willebrand 因子（VWF）的功能依赖于其大小。我们检验了以下假说：VWF 可能在切变力下在血小板糖蛋白 Ibα（GpIbα）受体上自组装。这一假说与以下事实一致：虽然 Alexa-488 偶联的 VWF（VWF-488）以适度的水平结合血小板，但添加未标记的 VWF 增强了 VWF-488 的结合程度。缺乏 A1 结构域的重组 VWF 与 Alexa-488 偶联以产生 ΔA1-488。虽然 ΔA1-488 单独在切变下不结合血小板，但在添加纯化的血浆 VWF 或重组全长 VWF 时，该蛋白结合 GpIbα。自组装的程度随施加的切变应力增加而增加超过约 60 至 70 达因/平方厘米。ΔA1-488 在血浆环境中结合血小板。在向全血施加流体切变时，一半的激活血小板结合有 ΔA1-488，表明 VWF 自组装可能是细胞激活所必需的。用 6-μm 载玻片珠剪切血小板，这些载玻片珠上固定有 VWF 或抗 GpIbα mAb，导致细胞在低至 2 至 5 达因/平方厘米的切变应力下激活。总之，数据表明循环中的流体切变可以通过蛋白自组装增加与血小板 GpIbα 结合的 VWF 的有效大小。这可以通过增加作用于细胞表面受体的阻力来触发机械转导和细胞激活。
