De Marco L, Mazzucato M, De Roia D, Casonato A, Federici A B, Girolami A, Ruggeri Z M
Centro Trasfusionale e Chimica Clinica, Centro di Riferimento Oncologico Aviano, Pordenone, Italy.
J Clin Invest. 1990 Sep;86(3):785-92. doi: 10.1172/JCI114775.
We have studied the interaction of the congenitally abnormal type IIA and IIB von Willebrand factor (vWF) molecules, both lacking the larger multimeric forms, with the two vWF binding sites on platelets, the glycoprotein (GP) Ib-IX and GP IIb-IIIa complexes. Variant as well as normal (N) vWF were purified from plasma. Estimates for binding of subunit molecules per platelet at saturation (Bmax) and dissociation constant in moles/liter (Kd), respectively, were obtained from binding isotherms of 125I-labeled vWF, with the following results. In the presence of ristocetin (binding to GP Ib-IX): N, 25,693 and 0.5 x 10(-8); IIA, both parameters not measurable; IIB, 17,708 and 0.87 x 10(-8). After thrombin stimulation (binding to GP IIb-IIIa): N, 17,059 and 1.12 x 10(-8); IIA, 23,751 and 4.87 x 10(-8); IIB, 19,890 and 2.52 x 10(-8). Distinct experiments based on measuring the ability of the variant species (from the same patients and one additional IIB patient) to inhibit the binding of normal 125I-vWF to platelets gave results in agreement with those reported above. Other studies showed that only IIB vWF bound to platelets in the absence of any mediating substance (Kd = 5.21 x 10(-8) mol/liter and Bmax = 9,599 subunits per platelet) and induced aggregation at a concentration of 10 micrograms/ml (3.6 x 10(-8) M). Thus, IIB vWF binds to GP Ib-IX with high affinity and induces platelet aggregation, whether with or without ristocetin, in spite of the absence of larger multimers. In contrast, the binding of IIA vWF to GP Ib-IX occurs with very decreased affinity, and this defective function may result from specific structural abnormalities rather than just being a reflection of the absence of larger multimeric forms. Both IIA and IIB vWF exhibit decreased affinity for GP IIb-IIIa. In this case, the extent of the defect correlates with the absence of larger multimers.
我们研究了先天性异常的IIA型和IIB型血管性血友病因子（vWF）分子（均缺乏较大的多聚体形式）与血小板上两个vWF结合位点，即糖蛋白（GP）Ib-IX和GP IIb-IIIa复合物之间的相互作用。从血浆中纯化出变异型以及正常（N）vWF。通过125I标记的vWF的结合等温线，分别获得了每个血小板在饱和状态下亚基分子的结合量估计值（Bmax）和以摩尔/升为单位的解离常数（Kd），结果如下。在存在瑞斯托霉素的情况下（与GP Ib-IX结合）：N型，25,693和0.5×10^(-8)；IIA型，两个参数均无法测量；IIB型，17,708和0.87×10^(-8)。在凝血酶刺激后（与GP IIb-IIIa结合）：N型，17,059和1.12×10^(-8)；IIA型，23,751和4.87×10^(-8)；IIB型，19,890和2.52×10^(-8)。基于测量变异型（来自相同患者以及另外一名IIB型患者）抑制正常125I-vWF与血小板结合能力的不同实验，所得结果与上述结果一致。其他研究表明，只有IIB型vWF在不存在任何介导物质的情况下与血小板结合（Kd = 5.21×10^(-8)摩尔/升，每个血小板的Bmax = 9,599个亚基），并在浓度为10微克/毫升（3.6×10^(-8) M）时诱导聚集。因此，尽管缺乏较大的多聚体，IIB型vWF无论有无瑞斯托霉素，都能以高亲和力与GP Ib-IX结合并诱导血小板聚集。相比之下，IIA型vWF与GP Ib-IX的结合亲和力显著降低，这种功能缺陷可能源于特定的结构异常，而不仅仅是缺乏较大多聚体形式的反映。IIA型和IIB型vWF对GP IIb-IIIa的亲和力均降低。在这种情况下，缺陷程度与缺乏较大多聚体相关。
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