bluuebunny/crossref_metadata_2025_split huggingface dataset:
Additive Effect of the Combined Administration of Low Molecular Weight Heparin and Recombinant Hirudin on Thrombus Growth in a Rabbit Jugular Vein Thrombosis Model
Authors: Biemond Bart J , Levi Marcel , Nurmohanned Michael T , Büller Harry R , ten Cate Jan W 
Published: July 2018

SummaryRecombinant Hirudin (r-Hirudin) is a new anticoagulant with specific antithrombin activity independently of antithrombin III. Low molecular weight heparins (LMWH) exert predominantly anti-Xa activity. Therefore, we hypothesized that combined administration of r-Hirudin and LMWH would induce a stronger antithrombotic effect as compared to r-Hirudin administered alone or combined with unfractionated heparin. To assess the effect on thrombus growth, we determined the accretion of l25I-labeled fibrinogen onto autologous non-radioactive thrombi preformed in the jugular veins of rabbits. The rabbits received unfractionated heparin (80 anti-factor Xa U), LMWH (80 anti-factor Xa U) or r-Hirudin (0.3, 5.0 and 10.0 mg/kg) either separately or by combined infusion for a 3 h period.R-Hirudin reduced the thrombus growth in a dose dependent fashion. The combined administration of 80 anti-Xa U LMWH and r-Hirudin at a dose of 0.3 mg/kg resulted in a stronger antithrombotic effect as compared to the combined infusion of unfractionated heparin and r-Hirudin (thrombus growth: 14.3% ± 6.0 vs 28.9% ± 6.5; p = 0.001). This difference in additive antithrombotic effect of 80 anti-Xa ULMWH versus unfractionated heparin on r-Hirudin was also observed when LMWH was combined with 5.0 mg/kg and 10.0 mg/kg r-Hirudin versus unfractionated heparin combined with r-Hirudin (thrombus growth: 16.4% ± 1.6 vs 29.1% ± 3.9; p = 0.01 and 10.1% ± 1.8 vs 20.4% ± 4.5; p = 0.001, respectively).In conclusion, this study showed an additive antithrombotic effect of LMWH on the thrombus growth reducing effect of r-Hirudin. This may potentially be of clinical interest, i. e. much lower doses of r-Hirudin may be used to achieve a similar effective anticoagulant effect when combined with LMWH.
Antithrombotic Effects of Recombinant Human Soluble Thrombomodulin in a Rat Model of Vascular Shunt Thrombosis
Authors: Ono Mayumi , Nawa Katsuhiko , Marumoto Yasumasa 
Published: July 2018

SummaryThrombomodulin on endothelial cells is a cofactor for thrombin-catalyzed activation of protein C. We have investigated the anticoagulant function of recombinant human soluble thombomodulin (rsTM) in a rat model of arterio-venous (AV)-shunt thrombosis. A bolus injection of rsTM 30 s before the induction of AV-shunt thrombosis inhibited the thrombus formation in a dose-dependent manner. The dose of anticoagulant that inhibited thrombus formation by 50% was 0.4 mg/kg rsTMα, 0.15 mg/kg rsTMβ, and 13 U/kg heparin. Recently, we characterized three monoclonal antibodies (moAbs) against human TM whose epitopes are located in the TM epidermal growth factor-like domain (Nawa et al., 1994). moAb 2A2 inhibited thrombin binding to rsTM, and abolished both TM functions as a cofactor in thrombin-catalyzed activation of protein C and as an anticoagulant by modifying thrombin-induced fibrinogen clotting and platelet aggregation. moAb 1F2 preserved the latter activities as an anticoagulant, but inhibited cofactor activity. moAb 10A3 had no inhibitory effect on either activity. Analysis of the in vivo anticoagulant mechanism of rsTM was facilitated by the availability of these moAbs. After incubation at rsTM/moAb molar ratios of 1:1.25, the effect of the mixtures were examined in the AV-shunt thrombosis model. An injection of 0.8 mg/kg rsTMα or 0.4 mg/kg rsTMβ resulted in a significant reduction on thrombus formation, as expected. moAb 10A3 had no effect on rsTM activity. However, co-injection of rsTM with moAb 1F2 resulted in a significant decrease of the inhibitory activity on thrombus formation. moAb 2A2 essentially abolished the inhibitory effect of rsTM. These moAb effects were observed for both rsTMα and β. These results suggest that the protein C activation accelerated by the injection of rsTM plays an important role for inhibiting thrombus formation in the AV-shunt thrombosis model.
Polymorphonuclear Leukocyte-Platelet Interaction: Role of P-Selectin in Thromboxane B2 and Leukotriene C4 Cooperative Synthesis
Authors: Maugeri Norma , Evangelista Virgilio , Celardo Antonio , Dell’Elba Giuseppe , Martelli Nicola , Piccardoni Paola , de Gaetano Glovanni , Cerletti Chiara 
Published: July 2018

SummaryIn PMN/platelet suspensions stimulated by fMLP giant mixed aggregates are formed and TxB2 and LTC4 are synthesized as the result of the cooperation in the arachidonic acid (AA) metabolism during cell/cell contact. PMN-derived cathepsin G induced the expression of P-selectin on platelet surface. GE12, an antibody against P-selectin, significantly reduced mixed cell aggregates. GE12 did not affect platelet aggregation induced by PMN-derived supernatants, indicating that the inhibitory effect of GE12 on mixed cell aggregation depends on inhibition of PMN/platelet adhesion. GE12 significantly reduced TxB2 and LTC4 production in PMN/platelet mixed cell suspensions stimulated by fMLP. As previously reported, synthesis of 3H-TxB2 in 3H-AA-labeled PMN/unlabeled platelets indicates that platelets utilize 3H-AA from PMN. 3H-LTC4 production in unlabeled PMN/3H-AA-labeled platelets indicates that bidirectional routes are utilized in this system for LTC4 synthesis. GE12 significantly reduced 3H-TxB2 and 3H-LTC4 synthesis. These results show that cathepsin G released by activated PMN induces the expression of P-selectin on platelet membrane: this adhesive glycoprotein modulates cell-cell contact and transcellular metabolism of AA.
Participation of Soluble Fibrin Monomer Complexes and Platelet Factor 4 in the Generalized Shwartzman Reaction
Authors: Lipiński B , Worowski K , Jeljaszewicz J , Niewiarowski S , Rejniak L 
Published: July 2018

SummaryThe generalized Shwartzman reaction was produced in rabbits by the typical way of 2 consecutive injections of Salmonella typhi endotoxin. Generalized Shwartzman reaction was also induced by “preparation” of the rabbits with epsilon-aminocaproic acid or mercuric chloride intoxication, followed by a single injection of the endotoxin. Significant impairment of blood fibrinolytic activity resulting from inhibition by endotoxin of a release of the kidney plasminogen activator, was demonstrated in the classical generalized Shwartzman reaction. Inhibition of fibrinolysis led to the accumulation of soluble fibrin monomer complexes in the circulation, detectable in plasma by protamine sulfate precipitation. It is postulated that hypercoagulability, regularly occurring in the generalized Shwartzman phenomen, is due to fibrinolysis inhibition. Endotoxin causes release of platelet factor 4 from rabbit blood platelets both in vitro and in vivo. This factor is thought to be responsible for nonenzymatic intravascular precipitation of fibrin from circulating soluble fibrin monomer complexes. Bilateral cortical necrosis in kidneys is due by fibrin deposition resulting from the disturbance in the equillibrium between blood clotting and fibrinolytic systems and the interaction of accumulated soluble fibrin monomer complexes with platelet factor 4.
A Sensitive and Facile Assay for the Measurement of Activated Protein C Activity Levels in Vivo
Authors: Orthner Carolyn L , Kolen Billy , Drohan William N 
Published: July 2018

SummaryActivated protein C (APC) is a serine protease which plays an important role as a naturally occurring antithrombotic enzyme. APC, which is formed by thrombin-catalyzed limited proteolysis of the zymogen protein C, functions as an anticoagulant by proteolytic inactivation of the coagulation cofactors VIIIa and Va. APC is inhibited by several members of the serpin family as well a by α2-macroglobulin. APC is being developed as a therapeutic for the prevention and treatment of thrombosis. We have developed an assay to quantify circulating levels of enzymatically active APC during its administration to patients, in healthy individuals, and in various disease states. This assay utilizes an EDTA-dependent anti-protein C monoclonal antibody (Mab) 7D7B10 to capture both APC and protein C from plasma, prepared from blood collected in an anticoagulant supplemented with the reversible inhibitor p-aminobenzamidine. Mab 7D7B10-derivatized agarose beads are added to the wells of a 96-well filtration plate, equilibrated with Tris-buffered saline, and incubated for 10 min with 200 μl of plasma. After washing, APC and protein C are eluted from the immunosorbent beads with a calcium-containing buffer into the wells of a 96-well microtiter plate containing antithrombin III (ATIII) and heparin. The amidolytic activity of APC is then measured on a kinetic plate reader following the addition of L-pyroglutamyl-L-prolyl-L-arginine-p-nitroanilide (S-2366) substrate.The rate of substrate hydrolysis was proportional to APC concentration over a 200-fold concentration range (5.0 to 1,000 ng/ml) when measured continuously over a 15 to 30 min time period. The coefficient of variation was 5.9% at 35 ng/ml and 8.8% at 350 ng/ml APC. The sensitivity of the assay could be increased by measuring the amount of color produced after longer incubation times in the endpoint mode. The measured APC activity levels were little affected by varying protein C or prothrombin over the extremes of 0 to 150% of normal plasma concentrations. By constructing the standard curve in protein C-deficient plasma, the concentration of APC activity in normal pooled plasma was determined to be 2.8 ng/ml (45 pM), which represents 0.08% of the protein C concentration. The assay was approximately 50-fold more sensitive than the identical assay, but using Mab-coated microtiter wells rather than immunosorbent beads as the capture step.
Properties of Chimeric (Tissue-Type/Urokinase-Type) Plasminogen Activators Obtained by Fusion at the Plasmin Cleavage Site
Authors: Colucci M , Cavallo L G , Agnelli G , Mele A , Bürgi R , Heim J , Semeraro N 
Published: July 2018

SummaryTwo hybrid plasminogen activators (K2tu-PA and FK2tu-PA), linking the kringle 2 domain or the finger plus the kringle 2 domains of tissue-type plasminogen activator (t-PA) to the catalytic domain of single-chain urokinase-type plasminogen activator (scu-PA) were studied. At variance with similar constructs previously reported, they were obtained by fusion of the t-PA and scu-PA derived portions at their plasmin cleavage site (between Arg275 of t-PA and Ile159 of scu-PA), thus eliminating from scu-PA the two peptide bonds (Glu143-Leu144 and Arg156-Phe157) that lead to low molecular weight scu-PA and to thrombin-inactivated tcu-PA. The specific activities of K2tu-PA and FK2tu-PA, as measured by fibrin plate were 2.5 × 106 and 1.0 × 106 t-PA equivalent units/mg, respectively. Activation of plasminogen by hybrid PAs was stimulated by both CNBr-digested fibrinogen (40- and 80-fold) and Des-A-fibrin monomers (6- and 12-fold). The relatively weak stimulation of chimeric PAs by minimally degraded fibrin monomers was consistent with their reduced fibrin binding capacity. Like scu-PA, the chimeric PAs, in the single-chain form, were insensitive to inhibition, as they retained full activity after prolonged incubation in plasma and did not interact with SDS-reactivated recombinant PAI-1. The concentration producing 50% lysis of blood clots in 3 h was 0.5 μg/ml for K2tu-PA and 1 μg/ml for FK2tu-PA, as compared to 0.5 μg/ml and >2 μg/ml for t-PA and scu-PA, respectively. Plasminogen and α2-antiplasmin consumption induced by the hybrid PAs in clot-free plasma was comparable to (K2tu-PA) or lower than (FK2tu-PA) that induced by either t-PA or scu-PA. When exposed to plasmin, the hybrids were completely converted into two-chain molecules with full enzymatic activity. At variance with u-PA, however, the two-chain recombinant activators still required fibrin for full expression of activity. These data indicate that the products of such “artificial” fusion behave like true chimeras without loss of biological activity. The insensitivity to thrombin inactivation and to the proteolytic cleavage leading to low molecular weight scu-PA might confer enhanced stability to the molecules, especially at thrombus level. Moreover, if the thrombolytic activity observed in vitro is maintained in vivo, the prolonged half life of these hybrids should result in higher plasma levels of activator and thus in more extensive and rapid lysis.
Extent of Differentiated Gene Expression in the Human Endothelium-Derived EA.hy926 Cell Line
Authors: Rieber Alison J , Marr Henry S , Comer Mary B , Edgell Cora-Jean S 
Published: July 2018

SummaryVascular endothelial cells perform many differentiated functions in processes such as angiogenesis, hemostasis, and inflammation. The number of recognized differentiated functions has increased rapidly in recent years, but there may be many more still unrecognized. The purpose of this study is to estimate the fraction of differentially expressed mRNA in a continuous human endothelium-derived cell line, EA.hy926. Random cDNA clones representing mRNAs from this cell line were labeled and used to probe blots of RNA from EA.hy926 cells and from cells of a relatively undifferentiated line. Of 49 random cDNAs, 5 cDNAs or 10% were found to represent mRNAs that are differentially expressed in EA.hy926 and in early passage umbilical vein endothelial cells. Since more than 104 different genes are thought to be expressed in the typical mammalian cell, our data indicate that about 103 gene products contribute to the differentiated properties of endothelial cells.
Transmembrane Calcium Influx Associated with von Willebrand Factor Binding to GP Ib in the Initiation of Shear-Induced Platelet Aggregation
Authors: Ikeda Yasuo , Handa Makoto , Kamata Tetsuji , Kawano Koichi , Kawai Yohko , Watanabe Kiyoaki , Kawakami Keiko , Sakai Kiyotaka , Fukuyama Mayumi , Itagaki Ichiro , Yoshioka Akira , Ruggeri Zaverio M 
Published: July 2018

SummaryWe found that the binding of multimeric vWF to GP Ib under a shear force of 108 dynes/cm2 resulted in the transmembrane flux of Ca2+ ions with a two-to three-fold increase in their intracellular concentration ([Ca2+]i). The blockage of this event, obtained by inhibiting the vWF-GP Ib interaction, suppressed aggregation. In contrast, the blockage of vWF binding to GP IIb-IIIa, as well as the prevention of activation caused by increased intracellular cAMP levels, inhibited aggregation but had no significant effect on [Ca2+]i increase. A monomeric recombinant fragment of vWF containing the GP Ib-binding domain of the molecule (residues 445-733) prevented all effects mediated by multimeric vWF but, by itself, failed to support the increase in [Ca2+]i and aggregation. These results suggest that the binding of multimeric vWF to GP Ib initiates platelets aggregation induced by high shear stress by mediating a transmembrane flux of Ca2+ ions, perhaps through a receptor-dependent calcium channel. The increase in [Ca2+]i may act as an intracellular message and cause the activation of GP IIb-IIIa; the latter receptor then binds vWF and mediates irreversible aggregation.
Comparison Of Progressive Antithrombin Activity And The Concentration Of Three Thrombin Inhibitors In Nephrotic Syndrome
Authors: Boneu B , Bouissou F , Sié P , Caranobe C , Abbal M , Barthe P 
Published: July 2018

A study was planed in 28 children affected with a nephrotic syndrome (group 1) in order to compare the total plasmatic progressive anti thrombin activity (Howie method) to the concentrations of three thrombin inhibitors, assayed by immunological methods : antithrombin III (AT III), α2 macroglobulin (α2 M), and al antitrypsin (α1 AT). The results were compared to those obtained in 21 normal children of the same age (group 2) and in 10 normal adults (age range : 25 - 50 yrs). The calibration curves were made with a pool of plasma from normal blood donors. The results (mean ± SD) and the comparison between groups 1 - 3 to group 2 are summarized.A subgroup of 15 affected children which a low AT III level (less than 80%) was isolated (mean AT III = 51% ± 18) ; the mean of the total progressive anti thrombin activity was 124% ± 16. A significant negative relationship was found between AT III and α2 M level in the group 1 (r = -0.71, P <0.01).The high plasmatic level of α2 M (1) accounts for the increased progressive anti thrombin activity in all the affected children in spite of very low levels of AT III ; (2) could explain the absence of thrombotic events in this serie ; (3) suggests that the benefit of heparin therapy might be doubtfull since it has been recently reported that heparin inhibit the binding of α2 M to thrombin.
The Effect Of Tranexamic Acid On Experimental Gastric Haemorrhage
Authors: Stenberg B , Alpsten M , Kjellström T , Risberg B 
Published: July 2018

The etiology of gastric haemorrhage is manifold. Defects in the haemostatic balance can provoke or sustain bleeding. The role of the fibrinolytic system has been debated. Some clinical studies indicate the value of antifibrinolytic treatment.It was the purpose of the present investigation to study the effectiveness of antifibrinolytic treatment in reducing gastic haemorrhage from experimentally induced ulcerations in rats.Methods. Gastric ulcerations were induced in 47 rats by pyloric ligation and intragastric instillation of 1.0 N HCI. Red blood cells were labelled with 51cr. The rats were divided in two groups: one group received tranexamic acid 60 mg per 100 g b.w. and the other group received corresponding amount normal saline. The rats were followed for 6 hours after ligation at which time the stomachs were removed and examined for the volume and radioactivity of the gastric content.Results. Six hours after pyloric ligation all rats had ulcers in the glandular part of the stomach. The volume of the gastric content in the control group was 15.1 ± 1.0 and in the tranexamic acid group 8.9 ± 2.2 ml (M±SE, p<0.01). The total amount of haemorrhage in the control group was 0.30 ± 0.02 and in the tranexamic acid group 0.21 ± 0.02 ml(M±SE, p<0.005)Conclusions. Our earlier studies demonstrated increased mucosal fibrinolytic activity after induced gastric ulcerations. Reduction of the increased muosal fibrinolysis by tranexamic acid in the present study significantly reduced the gastric haemorrhage.
Endotoxin And Blood Coagulation In Patients With Traumatic- Haemorrhagic And Bacteriotoxic Shock
Authors: Stemberger A , Straßer F , Blüimel G , Hundel shausen B v , Jelen S , Schmid O , Tempel G 
Published: July 2018

Blood coagulation and fibrinolysis was carefully monitored in patients suffering from traumatic-haemorrhagic shock. Parameters like antithrombin III (AT III),factor X (IIa), plasminogen,fibrin degradation products, antiplasmin and fibronectin were measured. It could be demonstrated that a severe haemorrhagic shock correlated with an activated blood coagulation and stabilisation of circulation was followed by normalisation of blood coagulation.However in patients developping septic complications new disorders in blood coagulation were observed. The appearance of endotoxins was measured by a recently developped limulus amoebocyte lysate (LAL). Best results were obtained by diluting heat treated heparinized plasma samples followed by an incubation step with the X a substrate S-2222. Detection of less than 0.2 ng endotoxin/ml plasma could be achieved. The available data of 30 patients with traumatic-haemorrhagic and bacteriotoxic shock showed that the appearance of endotoxin mostly correlated with a decrease of AT III and fibronectin.In conclusion, AT III, fibronectin and endotoxin may serve as sensitive markers in the early diagnosis of sepsis.
Deep Venous Thrombosis Of The Upper Extremity With Special Emphasis On Studies Of Coagulation And Fibrinolysis
Authors: Sundqvist S B , Hedner U , Bergentz S E , Kullenberg K 
Published: July 2018

Deep venous thrombosis (DVT) in the upper extremity is rare as compared to DVT in the leg. It has been claimed to be caused often by trauma to the axillary vein or unaccustomed heavy work (“effort”) with the arms. Our series of 53 patients shows the same peak incidence with regard to age (mean age 32.4+11.7) as earlier reported. However, a preceding trauma or “effort” was only found in 12 % in this particular series of patients. Other predisposing factors such as cancer (2/53), thoracic outlet syndrome (3/53) etc were rare. The coagulation studies included the following analyses: VIII:C, VIIIR:Ag, AT III and inhibitors of the plasminogen activators (PA inhibitor). The vessel wall fibrinolysis was studied in two different ways: 1) the capacity to release fibrinolytic activators from the vessel wall (venous occlusion test according to Robertson et al (1972), and 2) the fibrinolytic activity within the vessel wall (histochemical technique according to Pandolfi 1972). An impaired vessel wall fibrinolysis was found in 49 % of our patients which is a slightly lower figure than the 70 % incidence of this abnormality found in patients with spontaneous DVT in the lower extremity. The higher incidence of females as compared to earlier reports could not be explained by the use of contraceptive pills since the frequency of women on the pill seemed to be about the same as in the Swedish population as a whole.All patients had normal values of factor VIII, AT III and PA inhibitors.It is concluded that an impaired vessel wall fibrinolysis may be an important contributing factor also for DVT in the arm. Fibrinolytic studies may therefore be important in evaluating these patients.
Consumption of Hageman Factor Activity in the Generalized Shwartzman Reaction Induced by Liquoid
Authors: Müller-Berghaus G , Lasch H. G 
Published: July 2018

SummaryThe role of Hageman factor in triggering intravascular coagulation has been studied in rabbits injected intravenously with Liquoid. Besides changes of coagulation parameters characteristic of consumption coagulopathy (e.g. decrease in platelet counts, fibrinogen levels, factor V activity), a pronounced drop in Hageman factor activity was observed after injection of Liquoid. Likewise, the partial thromboplastin time became prolonged.The activation of Hageman factor in vivo could be prevented by intravenous infusion of lysozyme. Twenty min after starting the lysozyme infusion, the partial thromboplastin time became prolonged from a mean of 29 sec to 108 sec. Animals infused with lysozyme and injected with a lethal dose of Liquoid did not develop a consumption coagulopathy. In the same manner, none of 10 animals treated with lysozyme developed the generalized Shwartzman reaction, whereas in the control group 19 out of 20 animals showed fibrin thrombi in the glomerular capillaries.From the present study it may be concluded that the intravascular coagulation process after intravenous injection of Liquoid is triggered by Hageman factor activation.
Antihemophilic Factor: Its Presence in Fibrinogen, Assay Problems and the Stability of Lyophilized AHF-Fibrinogen Preparations
Authors: Baumgarten Werner , Sanders Benjamin E. , Belkin Benita D , Pagenkemper Frank E , Albers William G , Ciminera Joseph L 
Published: July 2018

SummaryThe thromboplastin generation test of Biggs et al. and of Pitney, and adopted by Surgenor for the assay of antihemophilic factor activity in lyophilized preparations of human fibrinogen, has been modified to yield a selfcontained statistically valid assay. This procedure was designed to serve primarily as a control procedure. The AHF activity of production lots of human fibrinogen was assayed, and the stability of AHF under different conditions of storage was determined.The procedure described in this paper is a satisfactory control procedure for AHF potency testing. In the absence of a generally recognized unit of AHF potency, it would seem most helpful to the clinician to have the potency related to the average activity in fresh normal human plasma.