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Human semen contains very potent blood clotting activity; for example, seminal serum diluted up to 10,000-fold significantly decreased the recalcification clotting time of blood plasma. This seminal coagulant activity was dependent on factor X and calcium ions, suggesting the presence of a facto X activator. Immunoblotting analysis and immunoadsorption studies confirmed the presence of tissue factor antigen (45 kDa) in semen. Centrifugation studies suggested that tissue factor was membrane associated, and fractionation of seminal serum by gel filtration followed by immunoelectron microscopy revealed that tissue factor antigen was on the prostasome vesicle surface. Tissue factor originated from prostatic fluid and not from seminal vesicle secretions. Tissue factor antigen averaged 21 ng/ml in seminal serum. Hypothetical roles for very high levels of tissue factor in semen include several possibilities. In the event of abrasion and bleeding during intercourse, rapid blood clotting at lesion sites would prevent sperm and seminal components, including infectious agents such as human immunodeficiency virus, from entering the blood stream, generating antibodies, or promoting infectious disease. This could imply that development of infection from semen-borne agents or development of antisperm antibodies in some patients could result from impairment or absence of seminal tissue factor.

We studied the interaction of factor X activation peptide (XAP) with factor IXa and factor Xa and the effect of XAP on factor IXa-catalyzed activation of factor X. XAP associated with factor Xa in the presence of 5 mM Ca2+ was dissociated from factor Xa by gel chromatography using Ultrogel AcA54 in 5 mM EDTA, or in 8 M urea-0.1% SDS. An exogenous isolated XAP inhibited the factor IXa-catalyzed factor X activation both in the presence and absence of factor VIIIa. 4-Amidinophenylmethylsulfonyl (aPMS)-factor Xa independent of XAP also inhibited the factor X activation more effectively than XAP alone in the presence of factor VIIIa. However, aPMS-factor Xa independent of XAP hardly inhibited the factor X activation in the absence of factor VIIIa. The binding of 125I-labeled factor X to the aPMS-factor IXa fixed to a microwell plate was inhibited by unlabeled factor X or XAP, but not by aPMS-factor Xa with or without XAP. Factor IXa directly bound to XAP and aPMS-factor Xa with XAP, but did not bind to aPMS-factor Xa without XAP. These findings suggest that the region of XAP in factor X directly interacts with factor IXa, and factor Xa region other than XAP interacts with factor VIIIa. Desialation or deletion of N-linked carbohydrates of XAP reduced the inhibitory activity of XAP for the factor X activation by factor IXa to approximately 50% of that of the intact XAP. This suggests that the sialic acids in the carbohydrate chains of the XAP region partly contribute to the interaction with factor IXa during its activation.

An estrogen-responsive procoagulant activity is present in the plasma membrane fraction of immature rat uterus. This procoagulant has many of the properties of tissue factor, a widely occurring, integral membrane protein which initiates the intrinsic pathway of coagulation. Procoagulant activity was demonstrated to activate prothrombin in rat uterus, to activate human coagulation factor X, and to cause clot formation by human plasma. Procoagulant activity could be solubilized from the plasma membrane by the detergent octyl glucoside and had an apparent mol wt of 20,000-40,000 by gel filtration. Procoagulant activity was increased 4-fold within 3 h after immature rats were injected with estradiol. The increase was tissue- and hormone specific and was not affected by a warfarin-induced vitamin K deficiency. Coagulation factor VII was required for clot formation by the procoagulant. These properties are consistent with identification of the procoagulant as tissue factor. mRNA for tissue factor was increased in the uterus 3 h after estrogen stimulation. In the preceding paper we showed that prothrombin is increased in the immature uterus within 3 h of estrogen stimulation. The presence of increased amounts of a tissue factor-like procoagulant in the same time period suggests a functional relationship between these two proteins and a possible role for both in uterine development. Thrombin is a growth factor in fibroblasts and endothelial cells. We propose that after estrogen stimulation, prothrombin enters the uterus with the influx of plasma proteins and is activated by the procoagulant to thrombin. We suggest that thrombin might act as a paracrine factor early in the estrogen-stimulated development of the uterus.

Antithrombin III is one of the main inhibitors in the blood coagulation mechanisms. Thrombin and factor Xa are slowly inactivated by it, as well as other serine proteinases of the coagulation mechanisms. Heparin tremendously accelerates the inhibitory function of antithrombin III. In the process antithrombin III activity is also reduced. Heparin retards the thrombin-fibrinogen reaction, but otherwise the effectiveness of heparin as an anticoagulant depends on antithrombin III in laboratory experiments, as well as in therapeutics. The activation of prothrombin is inhibited, and any thrombin or other vulnerable protease that might generate becomes inactivated. The measurement of antithrombin III concentration in blood is now achieved by research methods, as well as by methods that are practical for routine use. The tests require either thrombin or factor Xa as substrate, and could be specific for antithrombin III. There are congenital as well as acquired deficiencies of antithrombin III. The inhibitor is also found in tissues.