An assay to evaluate heparanase procoagulant activity

Abstract

BackgroundHeparanase that was cloned from and is abundant in the placenta is implicated in cell invasion, tumor metastasis and angiogenesis. Recently, we demonstrated that heparanase is directly involved in the regulation of the hemostatic system. Heparanase was shown to form a complex and enhance tissue factor (TF) activity, resulting in increased factor Xa production (Nadir et al. Haematologica, 2010). The present work suggests a novel assay to evaluate heparanase procoagulant activity. MethodsHeparanase procoagulant activity was studied using purified proteins of heparanase, TF, factor VIIa and factor X. The assay was verified in 55 plasma samples and compared to heparanase and tissue factor pathway inhibitor (TFPI) levels by ELISA and factor Xa, thrombin levels and antithrombin activity by chromogenic substrates. Thirty five samples were of third-trimester pregnant women (weeks 36–41) who were in labor or came for appointed elective cesarean section and 20 control samples were of non-pregnant healthy women. Resultsheparanase procoagulant activity assay was shown to differentiate heparanase procoagulant effect from TF activity, in purified proteins. Heparanase procoagulant activity was significantly higher in the plasma of pregnant women compared to non-pregnant (p<0.005). Heparanase relative contribution to the TF / heparanase complex activity was significantly higher in the plasma of pregnant women compared to non-pregnant (29% increase, p<0.0001). Differences in heparanase procoagulant activity were more prominent than changes in heparanase levels by ELISA, TF activity, factor Xa, thrombin and free TFPI levels. ConclusionsHeparanase procoagulant activity can be determined by the suggested assay. The assay revealed a significant contribution of heparanase to the procoagulant state in late third-trimester pregnancy and at delivery.