There is evidence that by catalyzing thrombin inhibition, several glycosaminoglycans can inhibit the thrombinmediated amplification reactions of coagulation and thereby delay prothrombin activation. The two amplification reactions can apparently be catalysed by endogenously generated factor Xa and thrombin. This study provides evidence which suggests that on a molar basis, an agent which can only catalyse thrombin inhibition is approximately 10 times more effective than an agent which can only catalyse factor Xa inhibition in their ability to inhibit intrinsic prothrombin activation. We determined the concentrations of each of heparin, dermatan sulfate and a pentasaccharide with high affinity for antithrombin 111, to delay intrinsic prothrombin activation for at least 15s. Heparin catalyses both thrombin and factor Xa inhibition; dermatan sulfate catalyses only thrombin inhibition, while the pentasaccharide only catalyses factor Xa inhibition. Efficient prothrombin activation, which coincided with both factor X activation and factor V proteolysis, was first observed 45s after CaC1, was added to contactactivated plasma. Heparin (= 0.1 pM) prolonged by at least 30 s the time required for the activation of the three clotting factors to begin. The minimum concentrations of the pentasaccharide and dermatan sulfate to delay the activation of prothrombin, factors X and V were z 50 pM and z 5 pM, respectively. Thus, each anticoagulant could inhibit intrinsic prothrombin activation only when it inhibited activation of both factors X and V. A combination of z 5 pM pentasaccharide and z 0.05 pM dermatan sulfate similarly delayed the activation of all three clotting factors. Thus, while catalysis of thrombin inhibition is a more effective pathway than catalysis of factor Xa inhibition for delaying prothrombin activation, the simultaneous catalysis of thrombin and factor Xa inhibition can synergistically improve the ability of a sulfated polysaccharide to delay prothrombin activation.Heparin and other glycosaminoglycans can prolong the lag phase associated with prothrombin activation [l -41. The efficiency with which glycosaminoglycans delay intrinsic prothrombin activation parallels their ability to catalyze thrombin inhibition in plasma [5 -71. This observation suggests that the anticoagulant effects of glycosaminoglycans result principally from their catalytic effects of thrombin inhibition. Anticoagulant effect, in this context, refers to the ability to delay intrinsic prothrombin activation. In addition to catalyzing thrombin inhibition, heparin catalyses factor Xa inhibition [4, 5,8 -lo]. Factor Xa is the only plasma enzyme known to efficiently convert prothrombin to thrombin and prothrombin fragment 1.2 [l 1,121. For these reasons, it has been difficult to determine the relative contributions of catalysis of factor Xa, and thrombin inhibition to the ability of heparin to delay prothrombin activation in plasma. Delineation of the relative roles of the catalysis of thrombin and factor Xa inhibition for anticoagulation i...

SummaryA recent study (Fernandez et al., Thromb. Haemostas. 1987; 57: 286-93) demonstrated that when rabbits were injected with the minimum weight of a variety of glycosaminoglycans required to inhibit tissue factor-induced thrombus formation by —80%, exogenous thrombin was inactivated —twice as fast in the post-treatment plasmas as the pre-treatment plasmas. In this study, we investigated the relationship between inhibition of thrombus formation and the extent of thrombin inhibition ex vivo. We also investigated the relationship between inhibition of thrombus formation and inhibition of prothrombin activation ex vivo. Four sulfated polysaccharides (SPS) which influence coagulation in a variety of ways were used in this study. Unfractionated heparin and the fraction of heparin with high affinity to antithrombin III potentiate the antiproteinase activity of antithrombin III. Pentosan polysulfate potentiates the activity of heparin cofactor II. At less than 10 pg/ml of plasma, all three SPS also inhibit intrinsic prothrombin activation. The fourth agent, dermatan sulfate, potentiates the activity of heparin cofactor II but fails to inhibit intrinsic prothrombin activation even at concentrations which exceed 60 pg/ml of plasma. Inhibition of thrombus formation by each sulfated polysaccharides was linearly related to the extent of thrombin inhibition achieved ex vivo. These observations confirm the utility of catalysis of thrombin inhibition as an index for assessing antithrombotic potential of glycosaminoglycans and other sulfated polysaccharides in rabbits. With the exception of pentosan polysulfate, there was no clear relationship between inhibition of thrombus formation and inhibition of prothrombin activation ex vivo.