Neutralisation of the anti-coagulant effects of heparin by histones in blood plasma and purified systems
Neutralisation of the anti-coagulant effects of heparin by histones in blood plasma and purified systems
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DOI：
10.1160/th15-03-0214
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发表时间：
2016-03-01
影响因子：
6.7
通讯作者：
Kolev, Krasimir
中科院分区：
文献类型：
--
作者：
Longstaff, Colin;Hogwood, John;Kolev, Krasimir
主要由DNA和组蛋白组成的神经元细胞外陷阱（NETs）是感染、炎症和凝血之间的联系。NET促进凝血，并且已经探索了使NET不稳定的方法以减少血栓形成和治疗脓毒症。类肝素结合组蛋白，我们报告定量研究血浆和纯化系统，以更好地了解生理后果。采用活化部分凝血活酶时间（APTT）测定普通肝素（UFH），并与低分子量肝素（LMWH）、凝血酶或凝血因子Xa（FXa）和抗凝血酶（AT）混合，测定UFH或LMWH对组蛋白的敏感性。开发了一种方法来评估DNA和非抗凝类肝素作为抗组蛋白的有效性。组蛋白能有效地中和UFH，在APTT中，0.2IU/ml UFH中和组蛋白的IC_（50）为1.8 μ g/ml，在纯化系统中，0.6IU/ml UFH中和组蛋白的IC_（50）为4.6 μ g/ml。组蛋白也抑制凝血酶（IC 50 6.1和11.0 μ g/ml组蛋白，对于不同的LMWH）或FXa（IC 50 7.8和7.0 μ g/ml组蛋白）的LMWH的活动。通过表面等离子体共振探索了UFH和LMWH与DNA和组蛋白的直接相互作用，而流变学研究显示了组蛋白、UFH和LMWH对凝块弹性的复杂影响。这些研究的结论是，即使在DNA存在的情况下，UFH和LMWH的抗凝作用也会因与循环组蛋白的高亲和力结合而受损。要全面了解组蛋白、DNA和肝素对止血系统的影响，必须了解其对纤维蛋白和凝块结构的直接影响。
Neutrophil extracellular traps (NETs) composed primarily of DNA and histones are a link between infection, inflammation and coagulation. NETs promote coagulation and approaches to destabilise NETs have been explored to reduce thrombosis and treat sepsis. Heparinoids bind histones and we report quantitative studies in plasma and purified systems to better understand physiological consequences. Unfractionated heparin (UFH) was investigated by activated partial thromboplastin time (APTT) and alongside low-molecular-weight heparins (LMWH) in purified systems with thrombin or factor Xa (FXa) and antithrombin (AT) to measure the sensitivity of UFH or LMWH to histones. A method was developed to assess the effectiveness of DNA and non-anticoagulant heparinoids as anti-histones. Histones effectively neutralised UFH, the IC50 value for neutralisation of 0.2 IU/ml UFH was 1.8 mu g/ml histones in APTT and 4.6 mu g/ml against 0.6 IU/ml UFH in a purified sys-tem. Histones also inhibited the activities of LMWHs with thrombin (IC50 6.1 and 11.0 mu g/ml histones, for different LMWHs) or FXa (IC50 7.8 and 7.0 mu g/ml histones). Direct interactions of UFH and LMWH with DNA and histones were explored by surface plasmon resonance, while rheology studies showed complex effects of histones, UFH and LMWH on clot resilience. A conclusion from these studies is that anti-coagulation by UFH and LMWH will be compromised by high affinity binding to circulating histones even in the presence of DNA. A complete understanding of the effects of histones, DNA and heparins on the haemostatic system must include an appreciation of direct effects on fibrin and clot structure.