bluuebunny/crossref_metadata_2025_split huggingface dataset:
Double-Blind, Randomized Comparison of Systemic Continuous Infusion of 0.25 Versus 0.50 mg/kg/24 h of Alteplase over 3 to 7 Days for Treatment of Deep Venous Thrombosis in Heparinized Patients: Results of the European Thrombolysis with rt-PA in Venous Thrombosis (ETTT) Trial
Authors: Bounameaux H , Banga J D , Bluhmki E , Coccheri S , Fiessinger J N , Haarmann W , Lockner D , Mahler F , Ninet J , Schneider P A , de Torrente A , van der Meer J , Verhaeghe and R 
Published: July 2018

SummaryThirty-two patients with acute, proximal-vein thrombosis were treated with heparin and alteplase (0.25 versus 0.5 mg/kg/24 h during 3-7 days) in a randomized, double-blind, multicenter, European (ETTT) trial. The treatment resulted in a decrease of the venographic Marder’s score from 18 (6-25) to 13 (2-24) units (median, range) in Group I (0.25 mg/kg/24 h, n = 15, median decrease 3.0, p = 0.32) and from 17.5 (3-33) to 15.5 (0-27) in Group II (0.5 mg/kg/24 h, n = 16, median decrease 4.0, p = 0.23). Comparison of the sequential venograms could be performed in 14 cases of Group I and in 15 cases in Group II. A minority of patients showed substantial partial recanalization of the initially obstructed veins on the control venogram (one in each treatment group) and most of the control venograms showedThus, the results of the ETTT trial show that the used low dosages of alteplase administered intravenously over 3-7 days in heparinized patients cannot be recommended as a treatment for patients with deep venous thrombosis of lower limbs and/or pelvis. Further studies are needed to define a more suitable dosage regimen of alteplase in this indication.
In Vitro Inhibition of Blood Coagulation by Tripeptide Aldehydes - A Retrospective Screening Study Focused on the Stable D-MePhe-Pro-Arg-H · H2SO4
Authors: Bágdy Daniel , Barabás Èva , Bajusz Sándor , Széll Erzsébet 
Published: July 2018

SummaryA series of peptide aldehydes synthetized in our institute during the last 15 years were screened to detect their inhibitory effect on blood coagulation. Simple conventional clotting assays, platelet function tests and fibrinolytic methods were used to evaluate the inhibitory potency of the compounds in complex clotting systems as well as their supposed antifibrinolytic effect in vitro. Special attention was paid to the possible interactions with blood cells and plasma proteins, and to the functional stability of the inhibitors in several tissue homogenates. D-Phe-Pro-Arg-H (GYKI-14166, RGH-2958), Boc-D-Phe-Pro-Arg-H (GYKI-14451) and D-MePhe-Pro-Arg-H (GYKI-14766) were found to be the most potent inhibitors. The peptide aldehydes via formation of reversible complexes with thrombin impede the enzyme to react with the coagulation factors, platelet membrane and vessel wall. The compounds inhibit platelet aggregation induced by thrombin specifically without changing the sensitivity of platelets to other inducers. D-Phe-Pro-Arg-H and D-MePhe-Pro-Arg-H showed no antifibrinolytic effect. D-MePhe-Pro-Arg-H and Boc-D-Phe-Pro-Arg-H proved to be stable in dry state for years and in solution at room temperature for several days. The anticoagulant activity of the compounds was declared in NIH antithrombin units.
Availability of the Bβ(15-21) Epitope on Cross-Linked Human Fibrin and Its Plasmic Degradation Products
Authors: Chen Fabian , Haber Edgar , Matsueda Gray R 
Published: July 2018

SummaryThe binding of radiolabeled monoclonal antifibrin antibody 59D8 (specific for fibrin but not fibrinogen) to a series of degraded fibrin clots showed that the availability of the Bβ(15-21) epitope (against which 59D8 had been raised) was inversely proportional to the extent of clot lysis. Examination of digest supernatants revealed that the Bβ(15-21) epitope was released from clots as a high molecular weight degradation product in the presence of calcium ions but that the generation of low molecular weight peptides occurred in the absence of calcium ions. To address the question of epitope accessibility, we compared levels of fibrin clot binding among four radioactively labeled antibodies: antifibrin monoclonal antibody 59D8, two antifibrinogen monoclonal antibodies that cross-reacted with fibrin, and an affinity-purified polyclonal antifibrinogen antibody. We expected that the antifibrinogen antibodies would show enhanced binding to clots in comparison with the antifibrin antibody. However, the epitope accessibility experiments showed that all four antibody preparations bound fibrin clots at comparable levels. Taken together, these studies demonstrated that one fibrin-specific epitope, Bβ(15-21), remains available on clots as they undergo degradation by plasmin and, importantly, that the epitope is not solubilized at a rate faster than the rate at which the clot is itself solubilized. The availability of the Bβ(15-21) epitope during the course of plasminolysis assures the potential utility of antifibrin antibodies such as 59D8 for detecting thrombi and targeting plasminogen activators.
Toward Gene Therapy in Haemophilia A: Retrovirus-Mediated Transfer of a Factor VIII Gene into Murine Haematopoietic Progenitor Cells
Authors: Hoeben R C , Einerhand M P W , Briët E , van Ormondt H , Valerio D , van der Eb A J 
Published: July 2018

SummaryTo study and evaluate the potential of the haematopoietic system as a target for gene therapy in haemophilia A, we have infected murine bone-marrow cells with a recombinant retrovirus encoding blood-coagulation factor VIII and the bacterial enzyme neomycin-phosphotransferase. After transplantation of the infected bone marrow into lethally irradiated mice, the presence of intact vector could be demonstrated in DNA isolated from individual haematopoietic progenitor-cell-derived spleen colonies. About 8% of the spleen colonies were shown to contain the intact vector. Selection for resistance to the neomycin analogue G418 prior to transplantation specifically killed the uninfected bone-marrow cells and, as a result, over 90% of the spleen colonies contained the factor VIII vector. However, expression of factor VIII in vivo, either at the RNA or at the protein level could not be demonstrated. From these data we conclude that: 1) retroviral vectors can be used to transfer factor-VIII cDNA into haematopoietic progenitor cells; 2) the vector sequences are expressed immediately after integration; and 3) transcription of the vector is repressed in the progenitor-cell-derived cells.
Biochemical, Thrombolytic and Pharmacokinetic Properties of rt-PA P47G, K49N, a Substitution Variant of Human Tissue-Type Plasminogen Activator
Authors: Nelles L , Li X-K , Vanlinthout I , De Cock F , Lijnen H R , Collen D 
Published: July 2018

Summaryrt-PA P47G, K49N, a substitution variant of recombinant human tissue-type plasminogen activator (rt-PA), in which proline at position 47 and lysine at position 49 were replaced by glycine and asparagine respectively, was previously described by Ahem et al. (J Biol Chem 1990; 265: 5540—5) to have an extended in vivo half-life with unaltered in vitro fibrinolytic properties. Because this variant might possess an increased in vivo thrombolytic potency, we have constructed its cDNA, expressed it in Chinese hamster ovary cells and determined its biochemical, thrombolytic and pharmacokinetic properties relative to those of home-made rt-PA and of alteplase (Actilyse®).The specific fibrinolytic activities on fibrin plates were 160,000 ± 17,000, 210,000 ± 88,000 and 460,000 ± 72,000 IU/mg (mean ± SEM) for rt-PA P47G, K49N, rt-PA and alteplase, respectively, while the catalytic efficiencies for plasminogen activation (k
2
K
m) in the absence of fibrin were comparable (1.1 to 1.7 × 10-3 μM-1s-1). Fibrin enhanced the rate of plasminogen activation by rt-PA P47G, K49N 100-fold and by both wild-type molecules 390-fold. Binding of the variant rt-PA to fibrin was significantly reduced, but its affinity for lysine-Sepharose was unaltered. In an in vitro clot lysis system, consisting of a radiolabeled human plasma clot submersed in plasma, 50% clot lysis in 2 h required 0.67 ± 0.14 pg/ml rt-PA P47G, K49N, 0.36 ± 0.01 pg/ml rt-PA and 0.17 ± 0.01 pg/ml alteplase, respectively (mean ± SEM; n = 3 or 4). At these doses residual fibrinogen levels at 2 h were in excess of 80%.The thrombolytic properties were examined in a hamster pulmonary embolism model. The thrombolytic potency, expressed as percent lysis per mg activator administered per kg body weight, was 160 ± 28 for rt-PA P47G, K49N, 150 ± 36 for wild-type rt-PA and 310 ± 42 for alteplase. The specific thrombolytic activity, expressed as percent lysis per pg steady-state t-PA-related antigen level per ml plasma, was 49 ± 18% for rt-PA P47G, K49N, 160 ± 49 for rt-PA and 500 ± 79 for alteplase. The clearance rates following bolus injection in hamsters were 0.18 ± 0.02, 1.9 ± 0.2 and 2.1 ± 0.1 ml/min respectively.Thus, the substitution of only two residues in wild-type rt-PA markedly reduces its clearance, but it also significantly alters its specific thrombolytic activity, resulting in a virtually unaltered thrombolytic potency.
Lack of Stability of Aggregates after Thrombin-Induced Reaggregation of Thrombin-Degranulated Platelets
Authors: Kinlough-Rathbone Raelene L , Packham Marian A , Perry Dennis W , Mustard J Fraser , Cattaneo Marco 
Published: July 2018

SummaryThe stability of platelet aggregates is influenced by the extent of the release of granule contents; if release is extensive and aggregation is prolonged, deaggregation is difficult to achieve. The relative importance of the contributions of released substances to aggregate stability are not known, although stable thrombin-induced aggregates form in platelet-rich plasma from patients with barely detectable plasma or platelet fibrinogen, and ADP stabilizes thrombin-induced aggregates of platelets from patients with delta storage pool deficiency which otherwise deaggregate more readily than normal platelets. We degranulated platelets with thrombin (0.9 U/ml caused greater than 90% loss of delta and alpha granule contents) and recovered them as individual platelets in fresh medium. The degranulated platelets were reaggregated by thrombin (2 U/ml). To prevent continuing effects of thrombin, FPRCH2C1 was added when thrombin-induced aggregation of thrombin-degranulated platelets reached its maximum. EDTA (5 mM) or EGTA (5 mM) added at maximum aggregation did not deaggregate these platelets, indicating that the stability of these aggregates does not depend on Ca2+ in the medium. Whereas with control platelets a combination of PGE1 (10 μM) and chymotrypsin(10 U/ml) was required for deaggregation, with thrombin-degranulated platelets either PGE1 or chymo-trypsin alone caused extensive deaggregation. The rate and extent of deaggregation of thrombin-degranulated platelets by a combination of PGE1 and chymotrypsin was greater than with control platelets.Electron microscope gold immunocytochemistry using antihuman fibrinogen IgG, anti-von Willebrand factor and anti-fibronectin showed a) that fibrinogen in the vacuoles of degranulated platelets was visible at focal points of platelet contact in the aggregates, but that large areas of platelet contact had no fibrinogen detectable between them; and b) in comparison to fibrinogen, little fibronectin or von Willebrand factor (vWf) was detectable in the platelets.Since the linkages between thrombin-degranulated platelets reaggregated by thrombin can be disrupted either by raising cAMP (thus making glycoprotein IIb/IIIa unavailable) or by proteolysis, these linkages are less stable than those formed between normal platelets. It might therefore be expected that platelets that take part in thrombus formation and then recirculate are likely to form less stable thrombi than platelets that have not released their granule contents.
ADP Receptor Antagonists and Converting Enzyme Systems Reduce Platelet Deposition onto Collagen
Authors: Wagner William R , Hubbell Jeffrey A 
Published: July 2018

SummaryThe effect of the ADP receptor antagonists ATP and adenosine 5’-(β, γ-methylene)triphosphate (AMP-PCP), and the ADP-utilizing enzyme systems creatine phosphokinase/creatine phosphate (CPK/CP) and pyruvate kinase/phosphoenol pyruvate (PK/PEP) on platelet deposition onto type I collagen was examined. An in vitro perfusion system was used, which allowed continuous visualization of the deposition of fluorescently labelled platelets. This system also provided well-controlled rheology, precise quantification of deposition, and allowed the use of heparinized whole human blood (3 u/ml). Heparinization at this level permits the local generation of thrombin near surface platelet aggregates. The contribution of ADP is thus studied with the combined effects of thrombin, thromboxane A2, and other aggregating agents present. Results from these studies indicate that ATP was capable of inhibiting deposition by 60% at 1 pM and 90% at 5 μM (whole blood cone.). AMP-PCP inhibited deposition in a dose dependent manner with a Ki
of approximately 80 μM and a maximum inhibition of 60%. Inhibition by CPK/CP was measured at 20, 40, and 60 u/ml, with approximately 45% inhibition achieved for the latter two concentrations. PK/ PEP at 60 u/ml resulted in 70% inhibition. These results support a role for ADP in mediating platelet recruitment in thrombus growth on collagen. Previous work utilizing animal bleeding times supports this conclusion; the present study demonstrates that this role is not dependent upon endothelial or vasoconstrictive effects. Intraplatelet cAMP levels were raised with respect to controls upon exposure to ATP at 8.3 μM (p <0.025), and 15 μM (p <0.005), as well as AMP-PCP at 42-500 μM (p <0.005). Since increasing intraplatelet cAMP may affect platelet function, these results raise another possible interpretation for some of our platelet deposition data as well as that of previous experimenters. However, since ATP at 1 μM was not shown to affect intraplatelet cAMP, it is believed that at least part of the ATP-mediated inhibition was due to ADP receptor competition.
Induction of Tissue Factor Synthesis in Human Umbilical Vein Endothelial Cells Involves Protein Kinase C
Authors: Pettersen Kjell Sverre , Wiiger Merete Thune , Narahara Nobuhiro , Andoh Kiyoshi , Gaudernack Gustav , Prydz Hans 
Published: July 2018

SummaryIncubation of human umbilical vein endothelial cells with one of the following compounds: endotoxin, recombinant interleukin-1β, recombinant tumor necrosis factor α, allogenic lymphocyte subpopulations or phorbol ester resulted in significant induction of tissue factor synthesis. Diacylglycerol had the same effect and also enhanced synergistically the induction caused by endotoxin and interleukin-1β. Two different inhibitors of protein kinase C, H7 and sphingosine, inhibited tissue factor synthesis at concentrations which did not depress protein synthesis in general, suggesting that protein kinase C is involved in the processes leading to tissue factor synthesis. Cells down-regulated for the tissue factor response to TPA responded essentially normally to endotoxin and interleukin-1 with regard to tissue factor synthesis.
On the Relationship Between Molecular Mass and Anticoagulant Activity in a Low Molecular Weight Heparin (Enoxaparin)
Authors: Bendetowicz Ana-Victoria , Pacaud Elisabeth , Béguin Suzette , Uzan André , Hemker H Coenraad 
Published: July 2018

SummaryA low molecular weight heparin (enoxaparin, mean molecular weight ~ 4,400) was separated by gel chromatography into eight different fractions with a narrow distribution around the following mean molecular weights: 1,800, 2,400, 2,900, 4,200, 6,200, 8,600, 9,800 and 11,000. We compared the influence of enoxaparin on the generation of thrombin in plasma to that of the eight fractions.We determined: a) the % of material with high affinity to antithrombin III (HAM) and the % of HAM above the critical chainlength necessary to allow for thrombin inhibition (ACLM), b) the specific catalytic activity on the decay of endogenous thrombin, and c) the inhibition of over-all thrombin formation in the extrinsic and the intrinsic pathway. From b and c we calculated the inhibition of prothrombin conversion in these pathways.We found that a) there is a gradual decrease of the HAM fraction with decreasing molecular weight; b) the specific catalytic activity for the inactivation of thrombin does not vary significantly between the fractions when expressed in terms of ACLM; c) the potency to inhibit prothrombin conversion does not vary significantly between the fractions when expressed in terms of HAM.
Effects of Splenectomy on Immune Thrombocytopenic Purpura in (NZW x BXSB) F1 Mice: Analyses of Platelet Kinetics and Anti-Platelet Antibody Production
Authors: Mizutani Hajime , Furubayashi Takayasu , Kashiwagi Hirokazu , Honda Shigenori , Take Hironori , Kurata Yoshiyuki , Yonezawa Takeshi , Tarui Seiichiro , Ikehara Susumu 
Published: July 2018

SummaryEffects of splenectomy on platelet kinetics and production of anti-platelet antibodies were studied in male (NZW × BXSB) F1 (W/B F1) mice, which are known as the animal model of immune thrombocytopenic purpura (ITP). Studies on organ localization of radiolabeled platelets revealed that splenic uptake significantly increases in W/B F1 mice in comparison with that of normal controls. W/B Fj mice showed a significant increase in platelet counts and, in contrast with sham-operated controls, high levels of platelet counts were maintained up to 6 weeks after splenectomy. Platelet lifespans (PLSs) did not reach normal levels, although prolonged PLSs were observed. In addition, platelet-associated antibody (PAA) values showed a tendency towards transient decrease, but there was no change in platelet-bindable serum antibodies (PBAs). These findings indicate that the suppression of anti-platelet antibody production is essential to the treatment of ITP; splenectomy may not be effective in treating severely affected ITP patients because, although the spleen is one of the major sites of platelet sequestration and antibody production, reticulo-endothelial systems (RESs) (liver, bone marrow, lymphnodes, etc.) other than the spleen are also responsible for the destruction of platelets. We therefore consider the W/B F1 mouse to be a useful model of human ITP, and believe that it provides valuable information for the development of new therapeutic agents in patients with ITP, especially those who do not respond to splenectomy.
Assessment of Lumiaggregometry for Research and Clinical Laboratories
Authors: White Melanie McCabe , Foust John T , Mauer Alvin M , Robertson James T , Jennings Lisa K 
Published: July 2018

SummaryPlatelet aggregometry is often used to help diagnose storage pool disease (SPD-reduced amounts of granule nucleotides) and release defects (abnormal release of granule nucleotides). The general assumption that normal aggregation patterns are sufficient to rule out the diagnosis of one of these disorders has been invalidated by the recent publication of two papers describing patients with clinical bleeding, prolonged bleeding times and normal aggregation patterns in spite of defective release. The lumiaggregometer provides a tool for measuring platelet release and aggregation simultaneously. This paper presents a standardized, reproducible method for the use of the lumiaggregometer based on a “standard curve”. Data obtained during the development of the procedure are presented including normal ranges of release at different concentrations of agonists, release measured in intrinsic disorders as well as in patients on aspirin, and values for release relative to varying platelet counts. A monoclonal antibody (anti-p24/CD9; MAb7) which activates platelets similarly to thrombin and may be a useful reagent for distinguishing SPD and release defects is also introduced.
Effects of Injected Antibody against the Platelet Glycoprotein llb/llla Complex on Monkey Platelet Fibrinogen
Authors: Suzuki Masahiko , Kawakatsu Toshihiro , Nagata Hirokazu , Hamamoto Kenjiro , Iwata Koji , Ohga Shigetoshi , Nomura Shosaku , Kokawa Terutoshi , Yasunaga Kojiro 
Published: July 2018

SummaryFour monkeys were injected for a 10-day period with the Fab fragment of a murine monoclonal antibody (NNKY 1-32) which inhibits the binding of fibrinogen to the platelet glycoprotein (GP) Ilb/IIIa complex. Platelet fibrinogen levels were assessed quantitatively by electroimmuno assay and qualitatively by immunoelectron microscopy.The platelet fibrinogen level fell to 9.0 ± 2.8% of the control level after antibody administration. Immunoelectron microscopy showed that the injected antibody was localized on the inner surface of the platelet a-granule membrane.Our findings suggest that the GP Ilb/IIIa complex can be internalized by a-granules and that it may mediate the endocytosis of plasma fibrinogen by platelets.
Prevalence of Inhibitors in a Population of 3435 Hemophilia Patients in France
Authors: Sultan Y ,   French Hemophilia Study Group
Published: July 2018

SummaryA cooperative study between the 37 centers of the French Hemophilia Study Group was undertaken to establish the prevalence of inhibitor patients in the French hemophilia population. The prevalence reported in the literature varies widely from 3.6% to 17.5%. Some of the studies are dealing with a small number of patients and inhibitor patients are reported either to the total number of hemophiliacs or to the severely affected ones. The French study provided information concerning 3,435 hemophiliacs and showed a prevalence of 6.2% for the overall population. Prevalence of inhibitors was found to be 7% in the population of hemophilia A patients and 12.8% in the population of severely affected ones. The prevalence of inhibitors in the population of hemophilia B patients was 2% and 4% in the population of severely affected hemophilia B patients. The cooperative study also showed that 47.5% of inhibitors are detected before 10 years of age and that 82% of inhibitor patients are high responders. Analysis of inhibitor detection in patients under the age often showed that there was a peak in the population of 2 years old children. Although not comparable to the present study the high incidence of inhibitors with ultrapurified and recombinant FVIII reported in previously untransfused patient may be borne in mind.
Effects of Heparin and Endothelial Cell Growth Supplement on Haemostatic Functions of Vascular Endothelium
Authors: Minter Allison J , Dawes Joan , Chesterman Colin N 
Published: July 2018

SummaryHeparin in combination with endothelial cell growth factor (ECGF) affects physiological responses and growth of human umbilical vein endothelial cells (HUVEC). We have examined the effect of heparin, crude ECGF (endothelial cell growth supplement [ECGS]), or both on the basal and thrombin challenged output of metabolites by HUVEC. The supernatant and/or cell lysate was assayed for released prostacyclin, von Willebrand factor, tissue plasminogen activator, plasminogen activator inhibitor and thrombospondin. Heparin modified release of all these metabolites when in combination with ECGS, and in general these responses were the opposite of those generated by inflammatory mediators such as interleukin-1. It has been postulated that heparin acts by potentiating the effect of ECGF, but heparin inhibited thrombospondin release and enhanced that of von Willebrand factor in the absence of ECGS, while ECGS alone inhibited release of plasminogen activator inhibitor. Thus, under our experimental conditions it would appear that heparin and crude ECGF can affect HUVEC independently of one another.