bluuebunny/crossref_metadata_2025 huggingface dataset:
Qualitätsmanagement in der Stroke Unit
Authors: Nabavi Darius , Ossenbrink Martin , Busse Otto
Published: July 2018

Qualitätsmanagement (QM) ist ein gesetzlich vorgeschriebenes strukturelles Element für Krankenhäuser in Deutschland. Zwar ist das Bestreben nach Qualitätsverbesserungen fester Bestandteil der Medizin, der formale QM-Ansatz wird in der Ärzteschaft dennoch bis heute kontrovers betrachtet. Aufgrund zeitkritischer Aspekte, risikobehafteter Therapiemaßnahmen und der interdisziplinären Herangehensweise besteht zweifelsfrei eine QM-Rationale für die Schlaganfallmedizin. Durch das in Deutschland bereits seit Mitte der 90er Jahre etablierte Zertifizierungsverfahren von Stroke Units (SU) werden qualitative Mindeststandards regelmäßig überprüft und QM-Maßnahmen verbindlich eingefordert. Dies umfasst die gesamte Versorgungskette vom prähospitalen Initialmanagement bis hin zum poststationären Konzept. Allerdings darf die Erlangung eines Zertifikates allein nicht als Endpunkt eines erfolgreichen QM betrachtet werden. Das fortlaufende Bemühen um Qualitätsverbesserungen setzt eine Haltung voraus, die von der Führungsebene glaubhaft vermittelt und in der gesamten Abteilung fest verankert sein muss. Insgesamt stellt die Entwicklung der Schlaganfallmedizin in Deutschland ein überzeugendes Beispiel für gelungenes QM dar.
Mn(OAc)3-Mediated Regioselective Radical Alkoxycarbonylation of Indoles, Pyrimidinones, and Pyridinones
Authors: Zou Jian-Ping , Zhang Wei , Li Cheng-Kun , Zhang Dong-Liang , Olamiji Ogundipe , Zhang Pei-Zhi , Shoberu Adedamola
Published: July 2018

Mn(OAc)3-mediated alkoxycarbonylation of indoles, pyrimidinones, and pyridinones with alkyl carbazates is reported. The reactions proceed through a radical process to afford regioselectively 3-carboxylated indoles, 5-carboxylated pyrimidinones, and 3-carboxylated pyridin­ones in moderate to good yields under mild reaction conditions.
Localization of the Br Polymorphism on a 144 bp Exon of the GPIa Gene and Its Application in Platelet DNA Typing
Authors: Kalb Rainer , Santoso Sentot , Unkelbach Katja , Kiefel Volker , Mueller-Eckhardt Christian
Published: July 2018

Alloimmunization against the human platelet alloantigen system Br (HPA-5) is the second most common cause of neonatal alloimmune thrombocytopenia (NAIT) in Caucasian populations. We have recently shown that a single base polymorphism at position 1648 on platelet mRNA coding for GPIa results in an aminoacid substitution at position 505 on the mature GPIa which is associated with the two serological defined Br phenotypes.

Since DNA-typing of platelet alloantigens offers possibilities for useful clinical applications, we designed genomic DNA-based restriction fragment length polymorphism (RFLP) typing for Br alloantigens. To establish this technique we analyzed the genomic organization of GPIa adjacent to the polymorphic base. Using the polymerase chain reaction (PCR) of blood cell DNA we have identified two introns (approximately 1.7 and 1.9 kb) flanking a 144 bp coding sequence of the GPIa gene encompassing the polymorphic base 1648. Based on the in- tron sequence, a PCR primer was constructed to amplify a 274 bp fragment which was used for allele-specific RFLP to determine the Br genotypes. The results of RFLP analysis using Mnll endonuclease obtained from 15 donors (2 Br37*, 2 Br^ and 11 Brb/b) correlate perfectly with serological typing by monoclonal antibody-specific immobilization of platelet antigens (MAIPA) assay.
Influence of the Xanthine Derivate HWA 138 on Endotoxin-Related Coagulation Disturbances: Effects in Non-Sensitized vs D-Galactosamine Sensitized Rats
Authors: Bahrami Soheyl , Redl H , Buurman W A , Schlag G
Published: July 2018

We have evaluated the effects of the xanthine derivate HWA 138 in rat endotoxemia in order to 1) prevent coagulation disturbances and other endotoxin-induced physiological abnormalities and 2) to reduce mortality. We performed two studies using two different models (sensitized vs non-sensitized rats) with a similar mortality but different severity of coagulation disturbances: a) LPS (15 mg/kg) alone or with HWA 138 (80 mg/kg) as a treatment modality 30 min pre LPS, b) galactosamine (500 mg/ kg) simultaneously with LPS (100 µg/kg) with or without HWA 138 (80 mg/kg) pretreatment. Experiments c) and d) employed D-galactosamine and/or LPS similar to experiments a) and b), while HWA 138 was applied simultaneously. We found significant 1) amelioration of life-threatening coagulation disturbances in non-sensitized rats, 2) prevention of liver dysfunction in sensitized rats, 3) reduction of TNF formation in both models, and 4) improvement of survival in all groups receiving HWA 138. Our data indicate protective effects of HWA 138 against clotting disturbances either directly via reduced LPS-induced formation of procoagulant activity or indirectly via reduced TNF formation.
Prevention of Thrombus Growth by Antithrombin III-Dependent and Two Direct Thrombin Inhibitors in Rabbits: Implications for Antithrombotic Therapy
Authors: Brill-Edwards Patrick , Van Ryn-McKenna Joanne , Cai Lu , Ofosu Frederick A , Buchanan Michael R
Published: July 2018

We compared the abilities of heparin and two direct thrombin inhibitors to prevent fibrin accretion onto pre-existing thrombi in rabbits. Inhibition of thrombus growth was measured as the ability of each test compound to inhibit the accretion of 125I-fibrin onto thrombi pre-formed in jugular veins of rabbits. When administered as a continuous infusion, the two direct (i. e. antithrombin III-independent) thrombin inhibitors, r-hirudin and a tripeptide, Ac(D)-Phe-Pro-bor-Arg (P-8714) inhibited fibrin accretion as effectively as heparin, but did so in doses which generated little systemic anticoagulation, as compared to the marked anticoagulation associated with the heparin effect. However, both r-hirudin and P-8714 were more effective when they were administered as a single bolus injection than as a continuous infusion. Under the former conditions, there was only a transient systemic anticoagulant effect. We conclude that direct or antithrombin III-independent thrombin inhibitors are more effective than heparin in preventing thrombus growth. The limited effect of heparin is likely due to fibrin impairing the ability of heparin/antithrombin III to inactivate thrombin.
The Anatomical Distribution of Plasma Fibrinolytic Activity in Man During Cardiac Catheterisation
Authors: Gough S C L , Smyllie J , Sheldon T , Rice P J S , Grant P J
Published: July 2018

Regional circulating plasma levels of fibrinolytic activity were assessed in 15 patients undergoing cardiac catheterisation. The euglobulin clot lysis time (ECLT) was longer in the abdominal aorta (AA) than the inferior vena cava (IVC), (median difference – 17.5 min, p = 0.008). This was associated with higher inhibition of plasminogen activator activity (PAI) in the AA than IVC, – 1.75 IU/ml, p = 0.002. In the venous circulation the ECLT was higher in the peripheral venous sample than in the IVC, –25.5 min, p = 0.003, with higher PAI peripherally than in the IVC, -1.9 IU/ml, p = 0.001. There were no differences in ECLT, PAI, PAI-l:Ag or t-PA:Ag throughout the arterial circulation. These results demonstrate higher fibrinolytic activity with lower inhibitor activity in the venous compared to the arterial circulation. Within the venous circulation fibrinolytic activity is lower peripherally with increased inhibitor activity.
The Interaction of Botrocetin with Normal or Variant von Willebrand Factor (Types IIA and IIB) and Its Inhibition by Monoclonal Antibodies that Block Receptor Binding
Authors: Fujimura Y , Miyata S , Nishida S , Miura S , Kaneda M , Yoshioka A , Fukui H , Katayama M , Tuddenham E G D , Usami Y , Titani K
Published: July 2018

We have recently shown the existence of two distinct forms of botrocetin (one-chain and two-chain), and demonstrated that the two-chain species is approximately 30 times more active than the one-chain in promoting von Willebrand factor (vWF) binding to platelet glycoprotein (GP) Ib. The N-terminal sequence of two-chain botrocetin is highly homologous to sea-urchin Echinoidin and other Ca2+-dependent lectins (Fujimura et al., Biochemistry 1991; 30: 1957–64).

Present data indicate that purified two-chain botrocetin binds to vWF from plasmas of patients with type IIA or IIB von Willebrand disease and its interaction is indistinguishable from that with vWF from normal individuals. However, an “activated complex” formed between botrocetin and IIB vWF expresses an enhanced biological activity for binding to GP Ib whereas the complex with IIA vWF has a decreased binding activity. Among several anti-vWF monoclonal antibodies (MoAbs) which inhibit ristocetin-induced platelet aggregation and/or vWF binding to GPIb, only two MoAbs (NMC-4 and RFF-VIII RAG:1) abolished direct binding between purified botrocetin and vWF. This suggests that they recognize an epitope(s) on the vWF molecule in close proximity to the botrocetin binding site.
Heterozygous Abnormal Fibrinogen Osaka III with the Replacement of γ Arginine-275 by Histidine Has an Apparently Higher Molecular Weight γ-Chain Variant
Authors: Yoshida Nobuhiko , Imaoka Shingi , Hirata Hajime , Matsuda Michio , Asakura Shinji
Published: July 2018

Congenitally abnormal fibrinogen Osaka III with the replacement of γ Arg-275 by His was found in a 38-year-old female with no bleeding or thrombotic tendency. Release of fibrinopeptide(s) by thrombin or reptilase was normal, but her thrombin or reptilase time in the absence of calcium was markedly prolonged and the polymerization of preformed fibrin monomer which was prepared by the treatment of fibrinogen with thrombin or reptilase was also markedly defective. Propositus' fibrinogen had normal crosslinking abilities of α- and γ-chains. Analysis of fibrinogen chains on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in the system of Laemmli only revealed the presence of abnormal γ-chain with an apparently higher molecular weight, the presence of which was more clearly detected with SDS-PAGE of fibrin monomer obtained by thrombin treatment. Purified fragment D1 of fibrinogen Osaka III also seemed to contain an apparently higher molecular weight fragment D1 γ remnant on Laemmli gels, which was digested faster than the normal control by plasmin in the presence of [ethy-lenebis(oxyethylenenitrilo)]tetraacetic acid (EGTA).
Antithrombotic Properties of Dermatan Sulphate (MF 701) in Haemodialysis for Chronic Renal Failure
Authors: Ryan K E , Lane D A , Flynn A , Ireland H , Boisclair M , Shepperd J , Curtis J R
Published: July 2018

The therapeutic potential of the glycosaminoglycan (GAG), dermatan sulphate (DS), as an antithrombotic agent in humans has yet to be established. We have performed dose ranging studies of DS to determine its effectiveness as an antithrombotic agent in patients (n = 6–8) undergoing haemodialysis for chronic renal failure. In an initial study, Study 1, i.v. bolus doses of 2–4 mg/kg and 5–6 mg/kg DS were given to patients dialysing with polyacrylonitrile hollow fibre (PAN HF) membranes. In a second crossover study, Study 2, performed using cuprophane hollow fibre (CHF) membranes, i. v. bolus doses of 3 mg/kg and 6 mg/kg DS were compared to a standard unfractionated heparin (UFH) regime that has been shown previously to inhibit fibrin formation. Further infusion studies, Study 3 and Study 4 evaluated the antithrombotic efficacy of an i. v. DS bolus of 3 mg/kg plus an i. v. infusion of DS 0.6 mg kg-1 h-1 and a DS bolus of 5 mg/ kg plus an infusion of 1 mg kg-1 h-1 over 5 h, respectively. These studies were compared to standard UFH regimes in a randomised crossover design. Plasma levels of fibrinopeptide A (FPA) and thrombin-antithrombin (TAT) were used as markers of fibrin formation and thrombin generation during dialysis using both membranes.

The changes in DS concentration following administration of the different doses were similar in Studies 1 and 2. However, the effectiveness of DS as an anticoagulant appeared to depend markedly on the different dialyser types used in the two studies. In Study 1, 13/14 dialyses required additional UFH to complete a normal ~6 h session and DS was unable to prevent thrombin and fibrin formation, as determined by measurement of plasma FPA and TAT. However, some dose related effects were observed in the levels of these markers. Furthermore, DS levels correlated with those of FPA and TAT. In Study 2, increasing doses of DS (3 mg/kg and 6 mg/kg), allowed longer dialysis sessions (mean 4.57 h c.f. 5.25 h), approaching that obtained with UFH regime (5.86 h). FPA and TAT generation were incompletely suppressed by both doses of DS; FPA rose significantly compared to that observed with the UFH regime, while TAT did not. While no significant differences in the activation markers were observed between the two DS doses, DS levels, taken as a whole, showed significant negative correlations with those of FPA and TAT Little effect on the KCCT was seen.

In Study 3, 3/6 patients required additional UFH (mean dialysis duration with DS 4.33 h c. f. 5.67 h with UFH). Mean DS levels were maintained between 35–40 µg/ml. Mean plasma FPA levels were maintained at constant levels throughout dialysis following DS administration but were higher than those observed following the UFH regime. In Study 4 mean DS levels were
Recurrent, Isolated Factor X Deficiency in Myeloma: Repeated Normalization of Factor X Levels after Cytostatic Chemotherapy Followed by Late Treatment Failure Associated with the Development of Systemic Amyloidosis
Authors: Schwarzinger I , Stain-Kos M , Bettelheim R , Pabinger I , Kyrle P , Kalhs P , Kapiotis S , Jäger U , Lechner K
Published: July 2018

We describe the case of a 64-year-old woman with isolated severe factor X deficiency associated with kappa light chain myeloma. At the time of diagnosis there was no evidence for amyloidosis. Complete remission (CR) of myeloma as well as normalization of factor X levels were achieved after cytostatic chemotherapy. Subsequently, factor X deficiency recurred twice without any evidence for relapse of myeloma. The first time factor X normalized again following cytostatic treatment, the second time, however, factor X deficiency was refractory to chemotherapy. Finally, relapse of myeloma became evident associated with rapidly progressing, systemic amyloidosis, which was fatal within a few months. Initially, factor X infusion studies showed a normal recovery, but when amyloidosis became overt the recovery decreased to 0%. We assume that factor X deficiency was due to a binding of factor X to kappa light chains associated with the proliferation of the malignant myeloma cell clone.
Use of Low Molecular Weight Heparin in Pregnancy
Authors: Melissari Euthemia , Parker Christopher J , Wilson Noel V , Monte Giovanni , Kanthou Chryso , Pemberton Kieran D , Nicolaides Kypros H , Barrett John J , Kakkar Vijay V
Published: July 2018

In a controlled study of 15 pregnant patients undergoing therapeutic termination of pregnancy, seven received subcutaneously 5,000 anti-FXa units of low molecular weight (LMW) heparin 15 and 3 h prior to the termination, and eight patients acted as controls. Paired maternal and fetal blood samples were taken (before or immediately after the termination) for assay of heparin activity by a chromogenic anti-FXa method sensitive to levels of 0.02 anti-FXa U/ml. LMW heparin was detected in all maternal samples of the test patients but was not detected in any of the fetal samples.

The use of LMW heparin as a thromboprophylactic agent was then evaluated in 11 patients who were known to have a severe thromboembolic tendency, had suffered recurrent miscarriages and had responded poorly to conventional anticoagulation (oral anticoagulant, conventional heparin). All patients receiving LMW heparin in thromboprophylactic doses completed uneventful pregnancies and gave birth to healthy babies (three for the first time) without complication. Bone density scans performed in all patients shortly after the delivery showed normal mineral mass. We conclude that LMW heparin does not cross the placental barrier, and in addition offers satisfactory antithrombotic protection for both maternal and placental circulation. In addition, this study provides preliminary data from 11 patients suggesting LMWH may not give rise to maternal osteoporosis, a finding that now needs further investigation.
Significance of Platelet β-Adrenoceptors for Platelet Responses In Vivo and In Vitro
Authors: Larsson P T , Wallén N H , Martinsson A , Egberg N , Hjemdahl P
Published: July 2018

The significance of platelet β-adrenoceptors for platelet responses to adrenergic stimuli in vivo and in vitro was studied in healthy volunteers. Low dose infusion of the β-adrenoceptor agonist isoprenaline decreased platelet aggregability in vivo as measured by ex vivo filtragometry. Infusion of adrenaline, a mixed α- and β-adrenoceptor agonist, increased platelet aggregability in vivo markedly, as measured by ex vivo filtragometry and plasma β-thromboglobulin levels. Adrenaline levels were 3–4 nM in venous plasma during infusion. Both adrenaline and high dose isoprenaline elevated plasma von Willebrand factor antigen levels β-Blockade by propranolol did not alter our measures of platelet aggregability at rest or during adrenaline infusions, but inhibited adrenaline-induced increases in vWf:ag. In a model using filtragometry to assess platelet aggregability in whole blood in vitro, propranolol enhanced the proaggregatory actions of 5 nM, but not of 10 nM adrenaline. The present data suggest that β-adrenoceptor stimulation can inhibit platelet function in vivo but that effects of adrenaline at high physiological concentrations are dominated by an α-adrenoceptor mediated proaggregatory action.
Biochemical and Ultrastructural Studies Suggest that the Effects of Thapsigargin on Human Platelets Are Mediated by Changes in Intracellular Calcium but not by Intracellular Histamine
Authors: Saxena Satya P , McNicol Archibald , Becker Allan B , Brandes Lorne J , Thastrup Ole , Gerrard Jon M
Published: July 2018

The involvement of intracellular histamine in thapsigargin (Tg)-induced platelet aggregation was studied. Platelet aggregation induced by 0.25 and 0.5 µM Tg was not accompained by a rise in intracellular histamine but a significant (p <0.01) increase in the level of intracellular histamine was observed at 1 µM Tg. Preincubation of platelets with inhibitors of histamine metabolizing enzymes had little effect on intracellular histamine levels in platelets stimulated by 0.5 µM Tg. In addition, the inhibitors of histidine decarboxylase (HDC), α-methyl histidine (α-MH) and α-fluoromethyl histidine (α-FMH) failed to inhibit Tg-induced aggregation. The intracellular histamine receptor antagonist, N,N-diethyl-2-[4-(phenylmethyl)phenoxy] ethanamine. HC1 (DPPE), inhibited Tg-induced aggregation but with IC50 values dependent on the concentration of agonist used. The inhibitory effects of DPPE on Tg-induced aggregation were not reversed by the addition of histamine to saponin-permeabilized platelets suggesting non-histamine mediated effects of DPPE on Tg-induced aggregation. Tg stimulated an increase in the cytosolic free calcium concentration which was unaffected by DPPE indicating that the effects of DPPE are also not due to the inhibition of mobilization of cytosolic calcium. The ultrastructural studies suggest that the major Tg-induced changes (pseudopod formation and granule centralization) are consistent with a primary role for Tg to mobilize calcium; DPPE had very little effect on these ultrastructural changes. The results indicate that the effects of Tg on human platelets are mediated by an increase in cytosolic calcium but not by intracellular histamine.
Antibodies to Thrombomodulin Induce Receptor-Mediated Endocytosis in Human Saphenous Vein Endothelial Cells
Authors: Brisson Christine , Archipoff Gisèle , Hartmann Marie-Louise , Hanau Daniel , Beretz Alain , Freyssinet Jean-Marie , Cazenave Jean-Pierre
Published: July 2018

The membrane glycoprotein thrombomodulin (TM) is an essential endothelial cell (EC) cofactor, which forms a 1:1 stoichiometric complex with thrombin. Binding of thrombin to the high affinity TM receptor transforms its procoagulant activity into an anticoagulant potential, by activating protein C. The fate of TM in the presence of thrombin is still unclear: some authors claim that the thrombin-TM complex is internalized in EC, while others find this complex to be stable for at least 2 h at 37° C on the EC surface. In the present study, we investigated the interactions of thrombin and Fab-fragments of anti-TM antibodies, coupled to 5 or 15 nm gold particles with saphenous vein endothelial cells. Our results demonstrate that TM can be observed both on the plasma membrane and in coated structures only in the presence of anti-TM antibodies. Addition of thrombin decreased the extent of this labeling, while in double labeling experiments, where cells were incubated with 5 nm gold coupled thrombin and 15 nm gold coupled Fab fragments of anti-TM antibodies, thrombin was cointernalized only when anti-TM antibodies were present. These results show that thrombin-TM complex is not significantly internalized in EC. The internalization of this complex induced by anti-TM antibodies could play an important role in the thrombotic complications induced by anti-EC autoantibodies.
Pharmacokinetics and Pharmacodynamics of Saruplase, an Unglycosylated Single-chain Urokinase-type Plasminogen Activator, in Patients with Acute Myocardial Infarction
Authors: Koster Rudolph W , Cohen Adam F , Hopkins Gwyn R , Beier Horst , Gunzler Wolfgang A , van der Wouw Poll A
Published: July 2018

We examined in patients with acute myocardial infarction (AMI) the pharmacokinetics of saruplase, an unglycosylated, single chain, urokinase-type plasminogen activator (rscu-PA) by measuring urokinase-type plasminogen activator (u-PA) antigen and total u-PA activity, its conversion to active two-chain urokinase-type plasminogen activator (tcu-PA) and evaluated its effect on haemostatic parameters.

Twelve patients were studied during and after administration of 20 mg bolus plus 60 mg continuous 1 h i.v. infusion of saruplase. For u-PA antigen and total u-PA activity (expressed as protein equivalents), where 234 U corresponds to 1 μg, respectively, steady state plasma concentrations were 2.75 ± 8.3 and 2.50 ± 7.0 μg/ml (mean ± standard deviation) and were reached within 20min, t1/2M was 9.1 ± 1.8 and 7.8 ± 1.3 min, t1/2λ2 1.2 ± 0.2 and 1.9 ± 0.5 h, and the total clearance was 393 ±110 and 427 ± 113 ml/min. Inactivation of saruplase in plasma was negligible.

After 15 min, tcu-PA was detected in plasma. From the ratio of the areas under the curve of tcu-PA and total u-PA activities it was calculated that 28 ± 9.3% of the saruplase dose is converted into active tcu-PA. Systemic plasminaemia occurs as shown by a decrease in α2-antiplas-min and fibrinogen and an increase in fibrinogen degradation products. Thrombin-antithrombin complex formation indicated activation of the clotting system.

Saruplase is eliminated rapidly from plasma in AMI patients. A variable, but significant proportion of saruplase is converted into active tcu-PA. At the end of the infusion tcu-PA accounted for 55% of total u-PA activity. Systemic plasmin generation and activation of the clotting system occur during saruplase treatment for AMI.
Thrombin-stimulated Human Platelets Express Molecular Forms of α-Granule Factor V (FV), which Differ from FVa
Authors: Wyshock Edward G , Stewart Gwendolyn J , Colman Robert W
Published: July 2018

Binding of 125I-Fab fragments of chain-specific antibodies indicate that both heavy and light chains of a-granule factor Va (FVa) were externalized on the platelet membrane after stimulation with thrombin. Using a Mab against the activation peptide of factor V (FV), the epitope appears on the stimulated platelet surface. Half as much light chain and heavy chain (FVa) was expressed compared to the activation peptide, suggesting that expression of α-granule FV occurs after thrombin stimulation. Using an ELISA, we find that 32% of a-granule FV was released and 68% is retained in the platelet pellet. Immunoblots of platelets indicate that FV exists in 200 kDa und 150 kDa forms, representing incomplete cleavage, while the releasate demonstrates a more complete cleavage by proteases. We conclude that expression of α-granule FV is quantitatively greater than that released and exists in molecular forms which cannot be completely explained by the binding of FVa.
Thrombin Generation during Cardiac Surgery: Is Heparin the Ideal Anticoagulant?
Authors: Brister S J , Ofosu F A , Buchanan M R
Published: July 2018

Blood samples were collected from 43 patients undergoing elective cardiac surgery to determine the extent of thrombin generation and inhibition in patients when receiving heparin while undergoing cardiopulmonary bypass (CPB). Plasma prothrombin fragment F1 + 2 and thrombin-antithrombin III (TAT) levels were measured as markers of thrombin generation and inhibition, respectively. Both F1 + 2 and TAT levels increased significantly during the course of CPB despite the heparin causing significant systemic anticoagulation, i.e. the activated coagulation time (ACT) was prolonged to greater than 400 s throughout the entire surgical procedure. The extent of thrombin generation increased with time on CPB but did not differ between patients receiving normothermic and hypothermic cardioplegia during CPB. Furthermore, thrombin generation increased following the neutralization of the heparin with protamine sulphate, and continued to be elevated significantly 24 h post surgery. The observation that high dose heparin did not prevent thrombin generation during CPB, is consistent with previous experimental studies demonstrating that thrombin bound to fibrin or other surfaces (e.g. the CPB conduit) is resistant to antithrombin III/heparin inhibition, and thus able to facilitate further thrombin generation. The observation that thrombin generation continued to be elevated post surgery i.e. 24 h after neutralizing the heparin with protamine sulphate, suggests that the high dose heparin did not inhibit effectively all of the thrombin that had been generated. Thus, CPB patients may be at risk not only of bleeding and other side-effects associated with the acute use of high dose heparin, but may also be at risk of further thrombosis-related events either acutely or chronically.
The Use of Active Centre Acylation to Control the Pharmacokinetic Profile of a Recombinant Chimaeric Plasminogen Activator
Authors: Wilson S , Cronk D W , Dodd I , Esmail A F , Kalindjian S B , McMurdo L , Browne M J , Smith R A G , Robinson J H
Published: July 2018

Recombinant hybrid plasminogen activators consisting of the “A” chain of plasminogen linked to the “B” chain of t-PA that are inhibited rapidly by plasma protease inhibitors have recently been described (Robinson et al. Circulation 1992; 86: 548-552). We have now shown that following bolus administration of native hybrid to guinea pigs, fibrinolytic activity was cleared rapidly from the circulation. Active centre acylation appeared to protect the hybrid from inhibition and allowed material to circulate as potentially active species for prolonged periods. Clearance rates of a range of acyl derivatives of the hybrid were 7-35-fold slower than for native hybrid and 20-100-fold slower than for t-PA. Clearance rates were influenced markedly by deacylation rate, such that clearance half-life correlated well with deacylation halflife. We have thus shown that it is feasible to control the pharmacokinetic profile of a recombinant hybrid plasminogen activator over a wide range by selection of an appropriate acyl group for attachment to the active site. Such control is not possible with plasminogen activators that are cleared predominantly by mechanisms other than inhibition.
Inhibitors to Factor VIII in a Family with Mild Hemophilia: Molecular Characterization and Response to Factor VIII and Desmopressin
Authors: Santagostino E , Gringeri A , Tagliavacca L , Mannucci P M
Published: July 2018

Inhibitor development in patients with mild hemophilia is a rare event. We report the occurrence of a persistent, high-responding inhibitor in two affected members of a mild hemophilia A family and discuss the therapeutic approaches employed in these patients in terms of their efficacy and effect on antibody titer. Desmopressin was an effective option for bleeding management, because endogenous factor VIII released by DDAVP was less immunogenic than exogenous factor VIII replacement, which invariably triggered anamnestic responses. Genetic analysis performed to investigate whether or not a peculiar molecular lesion accounted for this particular phenotype identified a G-A transversion at nucleotide 6507 in exon 23. This missense mutation has been already described in mild hemophilia A, but not in patients with inhibitors.
Some Characteristics of Mouse Platelet Aggregation and a Comparison of the Activity of a Range of Compounds in Mouse and Human Platelet-Rich Plasma in Vitro
Authors: Nunn Barbara
Published: July 2018

Collagen-induced platelet aggregation in mouse citrated platelet-rich plasma (PRP) was associated with 5-hydroxytryptamine (5-HT) release and degranulation. Adenosine 5’-diphosphate (ADP) induced monophasic and reversible aggregation with no degranulation. Mouse platelets gave no response to adrenaline but changed shape and sometimes aggregated in response to 5-HT. Both amines potentiated the effect of ADP. The respective potencies of prostaglandin E1, papaverine, VK 774, BL 3459 and adenosine as inhibitors of ADP-induced aggregation were similar in mouse and human PRP. Although ticlopidine had similar activity in the two species, aspirin and flurbiprofen were considerably less potent in mouse than human PRP as inhibitors of collagen-induced aggregation. It is suggested that collagen-induced aggregation of mouse platelets in vitro occurs by a mechanism largely independent of arachidonic acid metabolites.
Studies in Man and Experimental Animals of a Low Molecular Weight Heparin Fraction
Authors: Thomas D P , Merton R E , Lewis W E , Barrowcliffe T W
Published: July 2018

In vitro and in vivo studies were carried out on a commercially prepared low molecular weight heparin fraction. By APTT assay the fraction had a specific activity of half that of unfractionated mucosal heparin, yet retained full potency by anti-Xa assay (both clotting and chromogenic substrate). When administered intravenously to human volunteers, the anti-Xa/APTT ratio remained the same as it was in vitro. However, after subcutaneous injection, the ratio increased and anti-Xa activity could not be fully neutralized ex vivo by PF4. The fraction was as effective as unfractionated heparin in preventing experimental serum-induced thrombosis, suggesting that a heparin fraction with high specific activity by anti-Factor Xa assay compared to APTT activity may be an effective drug for the prophylaxis of venous thrombosis.
Comparison of the Cross-Linking Patterns of Lamprey Fibrinogen and Fibrin by the Action of the Intrinsic Lamprey Factor XIII and Human Factor XIII during the Process of Blood Coagulation
Authors: Murtaugh Patricia A.
Published: July 2018

Intrinsic lamprey factor XIII cross-links the γ-chain of lamprey fibrin (50,000 daltons) to the γ-dimer (100,000 daltons). The a-chain (110,000 daltons) is cross-linked very slowly to a-dimer (210,000 daltons) and a-trimer (330,000 daltons). In contrast, human factor XIII, when added in combination with intrinsic lamprey factor XIII, cross-finks the a-chain of lamprey fibrin to a high molecular weight polymer, and any remaining γ-chain is also cross-linked to a polymer. However, the γ-chain that has previously cross-linked to the γ-dimer by the intrinsic lamprey factor XIII remains as a γ-dimer. Factor XIII-free lamprey fibrin cross-links all its subunits (α, β, γ) to high molecular weight polymers when human factor XIII is added. In contrast to human and bovine fibrin where α-chain cross-linking in the process of blood coagulation commences when all of the γ-chain has cross-linked, the lamprey α-chain will begin to cross-link when approximately half of the γ-chain has cross-linked to the γ-dimer.
Evolution of Urokinase-Type Plasminogen Activator (u-PA) and Tissue-Type Plasminogen Activator (t-PA) in Orthotopic Liver Transplantation (OLT)
Authors: Himmelreich G , Dooijewaard G , Breinl P , Bechstein W O , Neuhaus P , Kluft C , Riess H
Published: July 2018

In orthotopic liver transplantation (OLT) hyperfibrinolysis seems to be of causative importance for intra- and postoperative bleeding. Although recently hyperfibrinolysis has been successfully reduced by intraoperative aprotinin treatment, small increases of fibrinolysis still remain during OLT. Originally, tissue-type plasminogen activator (t-PA) was considered to be responsible for the increases, but the efficacy of aprotinin which inhibits besides plasmin also kallikrein and urokinase-type plasminogen activator (u-PA) suggested also a role for the intrinsic and contact system-dependent plasminogen activators. We investigated the role of u-PA. From 29 patients undergoing OLT with intraoperative aprotinin infusion arterial blood samples were taken at 7 different time points. The preoperative median values for u-PA antigen (u-PA Ag) and plasmin-activatable single-chain u-PA (scu-PA) levels, which were more than 2-fold above normal (both: p <0.01), decreased slightly during the preanhepatic phase and remained unchanged during the anhepatic phase. With reperfusion of the graft liver the two levels decreased significantly (p = 0.0003 and p = 0.006, respectively) to almost normal values, probably due to clearance by the graft liver. Active two-chain u-PA (tcu-PA) was preoperatively 2-fold above the detection limit, remained stable during the preanhepatic phase and increased 2-fold in the anhepatic phase (p = 0.0018). As expected tcu-PA also relapsed upon reperfusion, but to the preoperatively enhanced level, possibly caused by sustained activation of scu-PA by cathepsin B. t-PA activity levels were at the upper end of the normal range preoperatively, slightly increased during preanhepatic and anhepatic phases and decreased significantly with reperfusion. The increases in tcu-PA and t-PA activities during the anhepatic phase coincided with greatly increased fibrinolysis as demonstrated by thrombelastography, indicating that both u-PA and t-PA are involved in the development of fibrinolysis during OLT.

One patient was excluded from statistical evaluations because preoperative u-PA Ag, scu-PA, tcu-PA and t-PA activity levels were much higher than in the other 28 patients. In the investigated group this patient was the only one with diffuse peritonitis intraoperatively and severe bleeding complications postoperatively which made retransplantation mandatory.
Plasma Lipids and Platelet Membrane Fluidity in Essential Hypertension
Authors: Quan Sang Kim Hanh Le , Mazeaud Madeleine , Astarie Catherine , Duranthon Véronique , Driss Fathi , Devynck Marie-Aude
Published: July 2018

Essential hypertension is often associated with high levels of plasma cholesterol or triglycerides. The relationships between plasma lipids and platelet lipids, membrane fluidity and functions in untreated hypertensive patients were investigated by measuring the fluorescence anisotropies of two fluorescent dyes (DPH and its cationic derivative, TMA-DPH, with different subcellular localization), cytosolic Ca2+ and pH, cyclic AMP content and aggregation to ADP and collagen. Hypercholesterolemia was found to be accompanied by a rise in platelet cholesterol content without changes in TMA-DPH or DPH anisotropies whereas hypertriglyceridemia was associated with a decreased cholesterol to phospholipid molar ratio, a decreased DPH anisotropy and a tendency of the cytosol to alkalinize. These results point out the differences between the effects of an acute cholesterol load and those of chronic hypercholesterolemia on platelet membrane microviscosity and aggregation. They demonstrate a strong association between plasma triglyceride levels and platelet membrane structure.
Increased Activity of Vitamin K-Dependent Clotting Factors in Human Hyperlipoproteinaemia – Association with Cholesterol and Triglyceride Levels
Authors: Constantino M , Merskey C , Kudzma D. J , Zucker M. B
Published: July 2018

Levels of blood coagulation factors, cholesterol and triglyceride were measured in human plasma. Prothrombin was significantly elevated in type Ha hyperlipidaemia; prothrombin and factors VII, IX and X in type lib; and prothrombin and factors VII and IX in type V. Multiple regression analysis showed significant correlation between the levels of these plasma lipids and the vitamin K-dependent clotting factors (prothrombin, factors VII, IX and X). Higher cholesterol levels were associated with higher levels of prothrombin and factor X while higher triglyceride levels were associated with higher levels of these as well as factors VII and IX. Prothrombin showed a significant cholesterol-triglyceride interaction in that higher cholesterol levels were associated with higher prothrombin levels at all levels of triglyceride, with the most marked effects in subjects with higher triglyceride levels. Higher prothrombin levels were noted in subjects with high or moderately elevated (but not low) cholesterol levels. Ultracentrifugation of plasma in a density of 1.21 showed activity for prothrombin and factors VII and X only in the lipoprotein-free subnatant fraction. Thus, a true increase in clotting factor protein was probably present. The significance of the correlation between levels of vitamin K-dependent clotting factors and plasma lipids remains to be determined.
Isolation of Antithrombin III from Normal and α1-Antitrypsin-Deficient Human Plasma
Authors: Borsodi Anna D. , Bradshaw Ralph A.
Published: July 2018

The plasma of individuals, hetero- or homozygous for α1-antitrypsin deficiency, contains greatly decreased amounts of antithrombin activity as assayed against factor Xa. However, heparin stimulation of the residual antithrombin activity is observed, which is comparable to that of normal plasma. Antithrombins isolated from both normal and α1-antitrypsin deficient plasma by a simplified procedure are indistinguishable in both properties and yields. The microheterogeneity observed on isoelectric focusing of both preparations can be eliminated by treatment with neuraminidase. Neither purified human antithrombin nor α1-antitrypsin, when assayed against bovine trypsin, is stimulated by heparin. These results clearly establish the unique natures of antithrombin and α1-antitrypsin and show that about 75% of the antithrombin activity measured in normal plasma is due to α1-antitrypsin. Estimates of anti thrombin III activity in normal plasma by assays dependent on enzymatic activity can probably be obtained only in the presence of heparin.
The Effect of Halofenate – Free Acid on Aggregation – the Release Reaction, Coagulant Activity, and Lipid Metabolism of Human Platelets
Authors: Akbar Huzoor- , Ardlie N. G
Published: July 2018

Halofenate – free acid (HFA), the major metabolite of the hypolipidemic drug, halofenate, inhibited platelet aggregation induced by collagen and sodium arachidonate and blocked the second phase of aggregation caused by ADP, thrombin and epinephrine in human platelet-rich plasma. The aggregation of washed platelets by thrombin and collagen was also blocked. HFA also inhibited the release by thrombin and collagen of 5-hydroxytryptamine from dense granules of platelets and the release by thrombin of β-glucuronidase from platelet α-granules. These inhibitory effects were concentration and time-dependent. HFA decreased platelet factor 3 activity by 31 % and also inhibited the incorporation of 14C-acetate and U-14C-glucose into platelet lipids by 89 % and 56 % respectively. Thrombin-induced lipid peroxidation and prostaglandin formation was investigated by measuring the by-product malonyldialdehyde, and this was found to be inhibited by HFA. It is suggested that the effect of HFA on aggregation is attributable to inhibition of the release reaction which may in turn be a consequence of the effects of the drug on platelet lipid synthesis.
Depression of Collateral Blood Flow Following Arterial Thrombosis
Authors: Schaub Robert G. , Sande Ronald , Meyers Kenneth M.
Published: July 2018

Permanent ligation of the feline aorta at the iliac bifurcation is followed by rapid opening of pre-existing collateral blood vessels. However, if ligation is combined with formation of a clot, these protective collateral vessels do not function. This study was undertaken to determine if drugs which alter serotonin function can improve collateral blood flow after arterial thrombosis. Permanent ligations were placed at the iliac bifurcation, circumflex iliac and sixth lumbar arteries in all cats. A clot was produced in the aorta of 27 cats by injection of 0.1 ml of thromboplastin. Ligated clot-occluded cats were untreated (10); had blood serotonin depleted using a single dose of reserpine (0.1 mg/kg i. m.) followed by para-chlorophenylanine (p-CPA) (100 mg/kg orally) every 3 days (9) ; or were treated prior to surgery with a serotonin antagonist cinanserin HC1 (4 mg/kg i. v.) (8). Control cats (18) were acutely ligated. 9 of these cats were untreated, 5 were cinanserin HC1-treated, and 4 were reserpine/p-CPA-treated. Extent of collateral development was assessed by aortograms 3 days after occlusion and by neurologic rating. Aortograms of acutely ligated cats indicated a significant collateral blood flow around the segment of ligated aorta, while ligated clot-occluded cats had a severely depressed hind-limb perfusion. Reserpine/p-CPA-treated ligation clot-occluded cats had aortograms similar to acutely ligated cats. The cinanserin HC1-treated ligation clot-occluded cats had aortograms which indicated hind-limb perfusion was not as adequate as the acutely ligated cats. However, the perfusion of these animals was improved over untreated ligation clot-occluded cats. Neurologic rating correlated with aortograms. These results suggest: 1) the clinical consequences of arterial thrombosis cannot be entirely attributed to mechanical occlusion of an artery, but may be due to depression of protective collateral blood flow induced by thrombosis, 2) serotonin is an important factor in this depression of collateral blood flow, and 3) isolation of the factors responsible for collateral inhibition could permit the development of therapeutic interventions.
Disseminated intravascular Coagulation in Status Epilepticus
Authors: Fischer Sharon P. , Lee Joseph , Zatuchni Jacob , Greenberg Jack
Published: July 2018

Disseminated intravascular coagulation is here reported as a complication of status epilepticus. Other features of this case were rhabdomyolysis, hyperthermia, myoglobinuria and renal failure.
Adenylate Cyclase and Phosphodiesterase Activity in the Platelet Release Abnormality
Authors: Chesney Carolyn Mci. , Baker Ann S. , Carvalho Angelina Ca. , Ozer Asli , Meissner George F. , Colman Robert W.
Published: July 2018

11 patients with histories of clinical bleeding were selected as examples of platelet release abnormality. Mean bleeding time was 18 ± 2.6 min (normal ±SEM; 6 ±0.44); mean platelet adhesiveness was 9.9 ± 4.3% (normal ±SEM; 30 ±2.2). Clot retraction and platelet factor 3 were normal. Platelet aggregation with adenosine diphosphate (ADP), epinephrine and collagen was decreased, as was 14C-serotonin release. Electron microscopic studies of platelets exposed to epinephrine showed 2 subgroups: one which failed to aggregate or have centralization of organelles and a second which developed pseudopodia and centralization of organelles, but rarely aggregated or degranulated.

Measurements of activity of adenylate cyclase and phosphodiesterase under basal conditions were performed on platelets from patients and control subjects. Adenylate cyclase activity was significantly lower and phosphodiesterase activity significantly higher in the patient group. Prostaglandin E1 was a potent stimulator of adenylate cyclase in both groups, as was NaF. It was concluded that the causative defects in patients with “platelet release abnormality” do not reside in either the activity of adenylate cyclase or of phosphodiesterase. Changes in formation and destruction of cyclic adenosinemonophosphate (AMP) may instead be regarded as a compensatory response to a defect in another effector system.
Molecular Cloning and Cell-Free Expression of Mouse Antithrombin III
Authors: Wu John K , Sheffield William P , Blajchman Morris A
Published: July 2018

The homology between antithrombin III (AT-III) of mouse, of man, and that of other species was investigated. Preliminary experiments showed that mouse AT-III inhibited human α-thrombin efficiently (second order rate constant [K
2nd] 5.8 × 103 M–1 s–1) as compared to human AT-III (K
2nd 6.7 × 103 M–1), but was not recognized on immunoblots by antibodies that recognized both human and rabbit AT-III. In order to compare AT-III from different species at the molecular level, a cDNA clone for murine AT-III was isolated from a λZAP mouse liver cDNA library on the basis of hybridization to a rabbit AT-III cDNA probe. The 1509 bp murine AT-III cDNA consists of a 1398 bp open reading frame, preceded by a 15 bp 5’ untranslated region, followed by a 75 bp 3’ untranslated region. The deduced primary protein structure consists of a 32 amino acid signal sequence, with a mature portion of 433 residues. Mature murine AT-III is 89% identical to its human counterpart, 86% identical to bovine AT-III, and 82% identical to that of the rabbit. Constructs lacking the nucleotides encoding the signal sequence were engineered and expressed in a cell-free system. The resulting 47 kDa non-glycosylated translation product was capable of being cleaved by human α-thrombin, of forming SDS-stable complexes with the protease, and of binding to immobilized heparin. Isolation of the murine AT-III cDNA will make feasible molecularly defined experiments with murine AT-III in the mouse system.
Effect of DDAVP on Endotoxin-Induced Intravascular Coagulation in Rabbits
Authors: Paloma M J , Páramo J A , Rocha E
Published: July 2018

We have evaluated the effect of l-Deamino-8D-arginine vasopressin (DDAVP) on an experimental model of intravascular coagulation (DIC) induced in rabbits by injection of 20 µg kg–1 h–1 during 6 h of E. coli lipopolysaccharide. DDAVP significantly ameliorated the platelet drop and fibrinogen decrease (p <0.01) induced by endotoxin in control animals. A significant reduction in factor XII consumption (p <0.01) and a decrease in the generation of endotoxin induced PAI-1 activity in rabbits circulation was also observed (p <0.005). Moreover, fibrin deposition in kidneys of rabbits receiving DDAVP was significantly reduced as compared to control animals. Finally, the mortality rate in the control group was significantly higher than in DDAVP-treated rabbits (p <0.01). The hemostatic changes induced by DDAVP correlated with lower fibrin deposition and reduction in mortality rates.