Nagase H, Enjyoji K, Minamiguchi K, Kitazato K T, Kitazato K, Saito H, Kato H
Taiho Pharmaceutical Co Ltd, Tokushima, Japan.
Blood. 1995 Mar 15;85(6):1527-34.
The inhibition mechanism of a polysaccharide anticoagulant, depolymerized holothurian glycosaminoglycan (DHG), was examined by analyzing its effects on the clotting time of human plasma depleted of antithrombin III (ATIII), of heparin cofactor II (HCII), or of both heparin cofactors. The effect exerted by this agent on the activation of prothrombin and factor X in purified human components were also examined and all effects were compared with those of other glycosaminoglycans (GAGs). The capacity of DHG to prolong activated partial thromboplastin time was not reduced in ATIII-depleted, HCII-depleted, HCII-depleted, or ATIII- and HCII-depleted plasma, whereas its capacity to prolong prothrombin time and thrombin clotting time was reduced in HCII-depleted plasma. DHG inhibited the amidolytic activity of thrombin in the presence of HCII with a second order rate constant of 1.2 x 10(8) (mol/L)-1 min-1. These results indicated that DHG has two different inhibitory activities, one being an HCII-dependent thrombin inhibition and the other an ATIII- and HCII-independent inhibition of the coagulation cascade. The heparin cofactors-independent inhibitory activity of DHG was investigated in the activation of prothrombin by factor Xa and in the activation of factor X by tissue factor-factor VIIa complex or by factor IXa. DHG significantly inhibited the activation of factor X by factor IXa in the presence of factor VIIIa, but not in the absence of factor VIIIa. The interaction between DHG and factors IXa, VIIIa, and X was investigated with a DHG-cellulofine column, on which DHG had strong affinity for factors IXa and VIIIa. These findings show that the heparin cofactors-independent inhibition exhibited by DHG was caused by inhibition of the interaction of factor X with the intrinsic factor Xase complex, probably by binding to the factor IXa-factor VIIIa complex.
通过分析一种多糖抗凝剂——解聚海参糖胺聚糖（DHG）对去除抗凝血酶III（ATIII）、肝素辅因子II（HCII）或两种肝素辅因子的人血浆凝血时间的影响，研究了其抑制机制。还研究了该试剂对纯化的人成分中凝血酶原和因子X激活的作用，并将所有作用与其他糖胺聚糖（GAGs）的作用进行了比较。在去除ATIII、去除HCII、去除HCII或去除ATIII和HCII的血浆中，DHG延长活化部分凝血活酶时间的能力并未降低，而在去除HCII的血浆中，其延长凝血酶原时间和凝血酶凝血时间的能力降低。在存在HCII的情况下，DHG以1.2×10⁸（mol/L）⁻¹ min⁻¹的二级速率常数抑制凝血酶的酰胺水解活性。这些结果表明，DHG具有两种不同的抑制活性，一种是依赖HCII的凝血酶抑制，另一种是不依赖ATIII和HCII的凝血级联抑制。在因子Xa激活凝血酶原以及组织因子-因子VIIa复合物或因子IXa激活因子X的过程中，研究了DHG不依赖肝素辅因子的抑制活性。在存在因子VIIIa的情况下，DHG显著抑制因子IXa对因子X的激活，但在不存在因子VIIIa的情况下则不然。用DHG-纤维素柱研究了DHG与因子IXa、VIIIa和X之间的相互作用，在该柱上DHG对因子IXa和VIIIa具有很强的亲和力。这些发现表明，DHG表现出的不依赖肝素辅因子的抑制是由于抑制了因子X与内源性因子X酶复合物的相互作用，可能是通过与因子IXa-因子VIIIa复合物结合。
