Impaired von Willebrand factor adhesion and platelet response in thrombospondin-2 knockout mice
Impaired von Willebrand factor adhesion and platelet response in thrombospondin-2 knockout mice
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DOI：
10.1182/blood-2016-03-702845
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发表时间：
2016-09-22
期刊：
影响因子：
20.3
通讯作者：
Kyriakides, Themis R.
中科院分区：
文献类型：
--
作者：
Kristofik, Nina;Calabro, Nicole E.;Kyriakides, Themis R.
胶原性细胞外基质和血管性血友病因子（VWF）之间的相互作用对于止血和血栓形成至关重要。在本研究中，我们研究了细胞外基质（ECM）异常的血小板反应蛋白2（TSP 2）基因敲除（KO）小鼠出血素质的贡献。首先，我们进行了过继骨髓移植，并观察到野生型（WT）骨髓引入致死辐射TSP 2基因敲除小鼠没有拯救出血素质。然而，移植小鼠的血小板显示出固有的聚集缺陷，这使解释复杂化。第二，我们进行了动脉段剥脱内皮的插入。移植到WT小鼠中的裸露的TSP 2 KO动脉在体内保持通畅。相比之下，WT移植物发生血栓形成，并在24至48小时内完全闭塞。通过将血小板暴露于TSP 2 KO真皮成纤维细胞（DF）衍生的ECM，在体外证实了TSP 2 KO ECM的非血栓形成特性。为了进一步探索TSP 2缺陷的影响，通过聚合酶链反应、免疫荧光和扫描电子显微镜分析WT和TSP 2 KO DF的ECM产生和沉积，并显示出相似的模式。此外，WT和TSP 2 KO ECM的原子力显微镜（AFM）分析未显示刚度差异。相比之下，当基质在生理流动下经受血浆时，观察到TSP 2 KO ECM上的VWF积累减少。利用VWF包被的2-mm珠的AFM证实了与TSP 2 KO ECM的弱结合，为缺乏血栓形成提供了机制解释。因此，我们的研究表明ECM组装对于胶原与VWF的相互作用和随后的血栓形成反应至关重要。
Interactions between collagenous extracellular matrices and von Willebrand factor (VWF) are critical for hemostasis and thrombosis. In the present study, we investigated the contribution of an extracellular matrix (ECM) abnormality to the bleeding diathesis in thrombospondin-2 (TSP2) knockout (KO) mice. First, we performed adoptive bone marrow transplantation and observed that introduction of wild-type (WT) marrow into lethally irradiated TSP2 KO mice did not rescue the bleeding diathesis. However, platelets in transplanted mice displayed an inherent aggregation defect, which complicated interpretation. Second, we performed interposition of arterial segments denuded of endothelium. Denuded TSP2 KO arteries grafted into WT mice remained patent in vivo. In contrast, WT grafts underwent thrombosis and were completely occluded within 24 to 48 hours. The nonthrombogenic property of the TSP2 KO ECM was confirmed in vitro by exposing platelets to TSP2 KO dermal fibroblast (DF)-derived ECM. To further probe the effect of TSP2 deficiency, ECM production and deposition by WT and TSP2 KO DFs was analyzed via polymerase chain reaction, immunofluorescence, and scanning electron microscopy and showed similar patterns. In addition, atomic force microscopy (AFM) analysis of WT and TSP2 KO ECM did not reveal differences in stiffness. In contrast, reduced VWF accumulation on TSP2 KO ECM was observed when matrices were subjected to plasma under physiological flow. AFM utilizing VWF-coated 2-mm beads confirmed the weak binding to TSP2 KO ECM, providing a mechanistic explanation for the lack of thrombus formation. Therefore, our studies show that ECM assembly is critical for interaction of collagen with VWF and subsequent thrombogenic responses.