凝血因子VIIa在流动状态下对活化血小板的凝血酶原酶功能。

Factor VIIa-mediated tenase function on activated platelets under flow.

作者信息

Goel M S, Diamond S L

机构信息

Institute for Medicine and Engineering, Department of Chemical and Biomolecular Engineering, University of Pennsylvania, Philadelphia, PA, USA.

出版信息

J Thromb Haemost. 2004 Aug;2(8):1402-10. doi: 10.1111/j.1538-7836.2004.00828.x.

DOI:10.1111/j.1538-7836.2004.00828.x

PMID:15304047

Abstract

BACKGROUND

Tissue factor (TF) and/or active factor (F)VIIa may be stored inside resting platelets.

OBJECTIVES

The objective of this study was to examine if platelets, following activation of GPVI, could support tenase and prothrombinase activity without any exogenously added tissue factor.

METHODS

Thrombin (IIa) formation on gel-filtered platelets with added factors or the clotting of platelet-free plasma (PFP) or platelet-rich plasma (PRP) supplemented with corn trypsin inhibitor (CTI) (to inhibit factor XIIa) was studied in well plate assays with a fluorogenic thrombin substrate or in flow assays by fibrin visualization.

RESULTS

Pretreatment of convulxin (CVX)-stimulated, fibrinogen-adherent, gel-filtered platelets with anti-TF, anti-FVII/VIIa, or 1 nm PPACK [inhibitor of FVIIa, factor XIa and factor (F)IIa] delayed fibrin deposition on platelets perfused with PFP/CTI at 62.5 s(-1). Anti-TF or anti-FVII/VIIa also attenuated thrombin generation in plate assays using recalcified PRP/CTI treated with CVX. Anti-TF or anti-FVII/VIIa (but not inhibited factor IXa) delayed the burst in thrombin production by gel-filtered platelets suspended in prothrombin and CVX by 14 min and 40 min, respectively. Anti-FVII/VIIa completely eliminated thrombin generation on fibrinogen-adherent, gel-filtered platelets pretreated with 10 micro m PPACK and 10 micro m EGR-CK [inhibitor of factor (F)Xa], rinsed, and then supplemented with CVX, prothrombin, and FX. Addition of anionic phospholipid to PFP/CTI or to a mixture of prothrombin, FX, and recVIIa was not sufficient to generate detectable tenase activity. Lastly, isolated, unactivated neutrophils suspended in FX, FII and recVIIa supported a very low level of thrombin generation sensitive to antagonism of P-selectin, CD18, and TF.

CONCLUSIONS

Activated platelets supported tenase and prothrombinase activity by elevating the function or level of FVIIa and exposing active FVIIa or FVIIa-cofactor(s), distinct from anionic lipid, that may be, in part, TF.

摘要

背景

组织因子（TF）和/或活性因子（F）VIIa可能储存于静息血小板内。

目的

本研究旨在检测糖蛋白VI（GPVI）激活后的血小板能否在不添加任何外源性组织因子的情况下支持凝血酶原酶和凝血酶原激活物活性。

方法

在微孔板分析中，使用荧光凝血酶底物，或在流动分析中通过纤维蛋白可视化，研究添加因子的凝胶过滤血小板上凝血酶（IIa）的形成，或补充玉米胰蛋白酶抑制剂（CTI）（以抑制因子XIIa）的无血小板血浆（PFP）或富血小板血浆（PRP）的凝血情况。

结果

用抗TF、抗FVII/VIIa或1 nM PPACK[FVIIa、因子XIa和因子（F）IIa的抑制剂]预处理convulxin（CVX）刺激的、纤维蛋白原黏附的凝胶过滤血小板，可使在62.5 s(-1)灌注PFP/CTI的血小板上的纤维蛋白沉积延迟。抗TF或抗FVII/VIIa在使用CVX处理的重新钙化PRP/CTI的平板分析中也减弱了凝血酶的生成。抗TF或抗FVII/VIIa（但不是抑制因子IXa）分别使悬浮在凝血酶原和CVX中的凝胶过滤血小板的凝血酶产生爆发延迟14分钟和40分钟。抗FVII/VIIa完全消除了用10 μM PPACK和10 μM EGR-CK[因子（F）Xa的抑制剂]预处理、冲洗后再补充CVX、凝血酶原和FX的纤维蛋白原黏附的凝胶过滤血小板上的凝血酶生成。向PFP/CTI或凝血酶原、FX和重组VIIa的混合物中添加阴离子磷脂不足以产生可检测到的凝血酶原酶活性。最后，悬浮在FX、FII和重组VIIa中的分离的未激活中性粒细胞支持对P-选择素、CD18和TF拮抗敏感的非常低水平的凝血酶生成。