bluuebunny/crossref_metadata_2025 huggingface dataset:
PARADOXICAL INCREASE IN HUMAN FACTOR VIII AFTER INFUSION OF PORCINE FACTOR VIII CONCENTRATE IN A PATIENT WITH ACQUIRED VARIANT VON WILLEBRAND'S DISEASE
Authors: Ball J , Malía R G , Greaves M , Preston F E
Published: June 2019

A patient with acquired variant von Willebrand's disease was given an infusion of 2000 units of high purity porcine factor VIII (Hyate). Quantitative factor VIII parameters were assessed following infusion and human factor VIII multimers were analysed by radioimmunoelectrophoresis and autoradiography. We have previously described the patient to have acquired von Willebrand's disease due to a circulating inhibitor to the factor VIII complex (B. J. Haematol-, 54,233,1983). Prior to infusion plasma from the patient contained factor VIIIC, RRCo, and vWFAg at less than 10 u/dl- Plasma factor VIII multimers showed an abnormal pattern with no high molecular weight bands present despite a normal triplet structure in the low molecular weight forms. After the infusion of porcine factor VIII concentrate a large increase in the levels of plasma VIIIC was detected with a disappearance half-life of 3.5 hours. A specific non-crossreacting immunoradiometric assay (IRMA) showed that plasma levels of porcine vWFAg did not rise significantly after the infusion. Despite this, human vWFAg levels were notably elevated at 1 hour (40 u/dl by Laurell) and 2 hours (30 u/dl by IRMA) post infusion. Similarly, ristocetin induced platelet aggregation and plasma RRCo levels showed significant elevations , 2 hours after the infusion. Factor VIII multimers assessed on plasma samples taken over a similar time period revealed the transient appearance of a normal compliment of human factor VIII multimeric forms 2 hours after the infusion of porcine factor VIII concentrate. This study indicates that the abnormal pattern of factor VIII multimeric bands present in inhibitor-related variant acquired von Willebrand's disease can be transiently normalised by infused porcine factor VIII concentrate. Whether this represents antibody displacement or de novo synthesis is yet to be determined.
VON WILLEBRAND FACTOR mRNA IS SEVERELY REDUCED IN PIGS WITH HOMOZYGOUS VON WILLEBRAND DISEASE
Authors: Wu Q Y , Bahnak B R , Coulombel L , Caen J P , Pietu G , Meyer D
Published: June 2019

Porcine von Willebrand disease (vWD), an autosomal recessive disorder, is similar to some of the severe forms of vWD in humans and is characterized by a prolonged bleeding time and very low or undetectable amounts of von Willebrand factor (vWF) antigen and activity in plasma, platelets and endothelial cells. The molecular events that control the lack of expression of vWF in the vWD pigs is not known and could be at the transcriptional or post-transcriptional level. Lungs from normal and two homozygous vWD pigs were extracted immediately after harvesting of the animals and placed on dry ice. Tissues were homogenized in 6 M guanidinium thiocyanate and RNA isolated by centrifugation through cesium chloride. Total RNA was analyzed by Northern hybridization including dénaturation in glyoxal, electrophoresis in 1.0 % agarose-2.2 M formaldehyde gels and transfer onto nitrocellulose. Messenger RNA was detected with a nick-translated human vWF cDNA probe or a human actin control probe. The vWF probe, cloned from a human lung library, was 2,280 bp in length and spanned nucleotides 960 to 3,240 of the human cDNA. These human probes were considered valid to detect levels of porcine vWF and actin mRNA because they hybridized with restriction enzyme digested genomic DNA from normal and vWD pig leucocytes under conditions of high stringency. The size of the vWF mRNA in the normal pigs after Northern hybridization was approximately 9.0 kb, similar to that of human vWF mRNA, and was easily detectable at the lowest concentration of RNA blotted (5 ug). In contrast, vWF mRNA from vWD pigs was at the lower limit of detection even at 10 ug of total RNA blotted. Nevertheless, although at extremely low levels, vWF mRNA from vWD pigs appeared to be the same size as the normal mRNA. These results agree with observations on the relationship of vWF secreted from 24 hr. cultures of endothelial cells from the pulmonary artery of normal and vWD pigs where the vWF levels were 0.90 and 0.06 U/108 cells, respectively. Therefore, it appears that the very low expression of vWF in the vWD pigs is due to a lack of transcription of the vWF gene. At this time, however, turnover of unstable transcripts in the vWD pigs can not be ruled out.
FACTOR XII, PLASMA PREKALLIKREIN ,α2-MACROGLOBULIN AND C1-INHIBITOR LEVELS IN RENAL ALLOGRAFT RECIPIENTS DURING IMMUNOSUPPRESSION WITH CYCLOSPORIN A
Authors: Lohri A , Huser B , Lämmle B , Oberholzer M , Thiel G , Duckert F
Published: June 2019

Factor XII clotting activity (FXII), Plasma Pre-kallikrein amidolytic activity (PK), 2-Macroglobulin ( α2-M) and cT-Inhibitor (cT-Inh) antigen have been measured in 17 patients immediately before and sequentially up to four months after kidney transplantation. Based on suspected Cyclosporin A (CyA) induced endothelial damage, activation of the contact system with resulting consumption of the contact activation factors was^evaluated. Before transplantation, FXII, PK, α2-M, Cl-Inh levels were 99±27%, 102±21%, 115±55%, and 129±32%, respectively. In the first two weeks after transplantation FXII decreased to 65±27%, PK to 67±20% and α2-M to 88±42%; Cl-Inh rose to a maximum of 201±44% (mean ± S. D.)(2p<0.001). Mean FXII levels correlated positively with PK, α2-M and albumin and negatively with CyA level and dose and serum bilirubin. PK and α2-M correlated positively with each other and with albumin and negatively with creatinine, bilirubin and CyA (p<0.01). The changes of FXII, PK and α2-M after transplantation suggest an influence of CyA on production or consumption of these factors. The behaviour of the cT-Inh may be unspecific and related to its action of an acute phase reactant.
DDAVP IN DIABETES INSIPIDUS
Authors: Vicente V , Corrales J , Miralles J , Alberca I
Published: June 2019

In order to investigate whether the response of von Willebrand factor (vWF), Factor VIII (FVIII) and tissue plasminogen activator (t-PA) to DDAVP infusion is governed by the integrity of the hypothaLamo neurohypophyseal axis, we studied the behaviours >of these proteins (FVIII, one stage; vWF antigen by electroimmunoassay and t-PA was measured in the plasma auglobulin fraction with added C-1 inactivator on fibrin plates) after DDAVP infusion (0.3 ug/Kg) in five patients with cranial diabetes insipidus, comparing them with the responses obtained in six healthy subjects.

In spite of receiving a daily therapeutic dose of 10-20 ug of DDAVP the patients with diabetes insipidus showed normal basal levels of FVIII, vWF and t-PA. The increase in these parameters following DDAVP infusion were not significantly different in the two groups. These findings suggest that the integrity of the hypothalamo-hypophyseal axis is not neccessary for a response by vWF, FVIII and t-PA to occur after DDAVP infusion.
EFFECTS OF DDAVP AT COAGULATION AND FIBRINOLYTIC LEVELS : DIFFERENT MECHANISMS OF ACT ION-AN EXPERIMENTAL STUDY IN THE DOG
Authors: Pina-Cabral J M , Cunha-Monteiro A , Sousa-Dias M C , Agular-Andrade J
Published: June 2019

After the injection of DDAVP in 39 non-anesthezised dogs (0.4 μg/kg) there was an average increase of factor VIII:C activity up to 145% (p<0.0001) and of fibrinolytic potential of euglobulin precipitate (t-PA) up to 196% (p<0.0001). The injection of DDAVP was repeated in each dog of a group of good responder animals at weekly intervals , but after : -A) Pentobarbital anesthesia (30 mg/kg)- the increase of factor VIILC was reduced from 164% to 116% (n=11; p<0.0005) and the increase in t-PA was reduced from 270% to 192% (n-11; p<0.05). -B) Injection of propranolol (tmq/kq) - the increase of factor VIILC was reduced from 167% to 110% (n=13; p<0.0005) and there was no decrease of fibrinolytic activity, (n-13; n.s.). -C) Splenectomv-the increase of factor VIILC was reduced from 166% to 122% (n=10; p<0.0005) and t-PA was increased from 196% to 256% (n=9; n.s.). There were no statistical significant differences in factor VIILC and fibrinolytic activities after repeating only the injection of DDAVP three times in the same animal at weekly intervals (n=5; n.s.).

We conclude that the increase in F. VIII:C and fibrinolytic activities observed after DDAVP infusion are due to different mechanisms of action. On the one hand, pentobarbital anaesthesia reduced the increase of factor VIILC and t-PA , but on the other hand , beta-blockade and splenectomy influenced differently the behaviour of both biological activities.
DIFFERENTIAL CHANGESOF PLASMATIC MARKERS OF THE ENDOTHELIAL FOLLOWING dDAVP
Authors: Scrobohaci M L , Drouet L , Baudin B , Rodriguez A
Published: June 2019

Plasmatic levels of tissue plasminogen activation is commonly assumed to be of endothelial origin ; Angiotensin converting enzyme can be of endothelial or monocytic origin;Fibronectin is mostly from hepatic origin but endothelium can participate. Protein S is a newly recognized endothelial protein. Plasmatic von Willebrand factor is mostly from endothelial origin as the factor megakaryocytic origin is stored in platelet αd granules.

dDAVP by indirect effect stimulates release of endothelial factors,13 patients with moderate von Willebrand disease were submitted to an infusion of 0.4 ug/kg dDAVP and were followed for four hours after infusion.

Interstingly the kinetics of release of each of the studied factors is totally different.

-Tissue plasminogen activator israpidly released and come back alsoquickly to its original level,- Release of von Willebrand factor is delayed comparatively to tPA and stayed for a longer time at increased level,- Angiotensin converting enzyme is not affected by dDAVP while its level is stimulated by veno-occlusion- Fibronectin and Protein S arenot significantly modified by dDAVP

From these results mechanism of action of dDAVP cannot be hypothesized, but it seems evident that the release (and so the endothelial metabolism) of each of the studied factorsis different and not linked.
IMMUNOLOGIC EVALUATIONOF ITALIAN HAEMOPHILIACS TREATED WITH COMMERCIAL CONCENTRATES
Authors: Rossi L , Gianregorio A M , Bartolai S , Lecchini L , Bendinelli M , Panicucci F
Published: June 2019

The presence of the antibody to human immunodeficiency virus (HIV) was investigated in 487 serum samples withdrawn from 216 haemophiliacs (186 haemophiliacs A and 30 B) who came to the Pisa Centre between 1977 and 1986. Results show that a considerable proportion of haemophiliacs (15%) were already positive in 1982. In haemophiliacs A, who were treated up to 1983 with concentrates made from US plasma, but from 1984 on began to use concentrates made from Italian plasma, this proportion in 1986 was 16%. In haemophiliacs B who have continued using concentrates from US plasma, the prevalcence rate of Anti-HIV was found to be much higher (63%). As regards the contact with the virusof hepatitis B (HBV), antibodies werefound in 64% of haemophiliacs and theviral antigens in 6%. 137 of these haemophiliacs (112 A and 25 B) were examined between 1985 and 1986 for indexof hepatic cytoli-sis, immunological and clinical status. In 65% of haemophiliacs the level of serum glutamic pyruvate transaminase (SGPT) was mildly increased, perhaps for the presenceof chronic hepatitis. In Anti-HIV negative subjects we found a decrease of T helper/T suppressor (Th/Ts) ratio, with a mean of 1.3 (in controls, 1.7),due especially to an increase of Ts (mean 0.8 × 103Vcu.mm.; incontrols, mean 0.6). Also in Anti-HIVpositive haemophiliacs there was a decrease ofTh/Ts ratio (mean 1.1), but this was mainly due to a decrease of Th (mean 0.7 × 103/cu.mm.;in controls, mean 1.1). Clinical evaluation of the Anti-HIVpositive subjectsshowed 2 patients with AIDS related illness with opportunistic infections and 13 haemophiliacs (33% of seropositive subjects) withone or more of these abnormalities: thrombocytopenia, lymphoadenopathy, slight persistentfever, diarrhea.
INACTIVATION OF VIRUSES IN PLASMA ON TREATMENT WITH TRI(N-BUTYL) PHOSPHATE (TNBP) DETERGENT MIXTURES
Authors: Piët M P J , Chin S , Prince A M , Horowitz B
Published: June 2019

Treatment of plasma with organic solvent/ detergent mixtures at the time of plasma collection or plasma pooling could reduce the exposure of technical staff to infectious virus and enhance the viral safety of final product. Treatment of plasma for 4 hours with 2% TNBP at 37°C or with 1% TNBP and 1% Tween 80 or Triton X-45 at 30°C resulted in the rapid and complete inactivation of ≥104 tissueculture infectious doses (TCID-50) ofvesicularstomatitis and Sindbis viruses, used as surrogates. TNBP and TNBP/Tween treatment of plasma was shown to inactivate ≥104 TCID-50 of human immunodeficiency virus. TNBPtreatment of plasma contaminated with 106 chimpanzee infectious doses (CID-50) of HBV and 105 CID-50 of NANBHV prevented the transmission of hepatitisto chimpanzees through 6 months follow-up.

Immediately following treatment with 2% TNBP, the recovery of AHF, factor IX, factor V, and antithrombin IIIwas 75%,90%, 65% and 100%,respectively. A ⊕90% recovery of AHF was observed with TNBP/detergent mixtures. Treated plasma was fractionated intoAHF and prothrombin complex concentrates, immune globulin, and albumin by published techniques. Prior treatmentwithTNBP or TNBP and detergent did not affect the separations of desired proteins. Therefore, it appears possible to inactivate viruses in plasma prior to execution of standard fractionation procedures. If desirable, products prepared from TNBP-treated plasmacan be subjected to additional procedures to further insure virus safety.
EFFICIENCY OF STEAM TREATMENT PROCEDURES USED FOR VIRUS INACTIVATION IN HUMAN COAGULATION FACTORS
Authors: Barrett P N , Wöber G , Eibl J , Dorner F
Published: June 2019

The transmission of viral agents from blood products derived from humanplasma has long been a serious healthproblem for the recipients. The majoragents involved are HBV, nonA nonB, Delta agent and HIV (LAV/HTLV-III). Wehave attempted to design product-specific viral inactivation procedures toprevent such transmission in blood products. Experiments were carried out with HIV, which is the only one of the above viruses which can be propagated and titrated in cell culture, and with a wide range of model viruses. High titre virus was added to blood products, and they were then subjected to different virus inactivation procedures.

Heat treatment was carried out in aqueous solution with and without protein-stabilising agents, and it was demonstrated that such agents added toprevent loss of biological activity of blood products also resulted in a stabilisation of contaminating viruses.

Therefore steam treatment inactivation procedures have been developed without the addition of such agents. Steam pressure, temperature and time of inactivation are the main variablesand these can be altered for each product to achieve an optimal balance between the degree of virus inactivation and retention of biological activity.

It has been shown that each of thedifferent product-specific treatment procedures has a high HIV inactivation capacity.

Studies with various model virusesalso demonstrated that the efficiencyof these procedures could be furtherimproved if necessary by altering various parameters such as steam pressure.
A STUDY ON THE RECOVERY OF FACTOR VIII PROCOAGULANT ACTIVITY FROM RECALCIFIED, HEPARINISED, CITRATED PLASMA
Authors: Cumming A M , Wensley R T , Cottrell S E , Delamore I W
Published: June 2019

This study has been carried out to investigate the potential for increasing the recovery of factor VIII procoagulant activity (factor VIII:C),in cryoprecipitates and concentrates, by the use of heparin anticoagulant. Jhctor VIII:C in citrated plasmahas been shown to be stabilised by recalcification and heparinisation of the plasma. Donations of substantially platelet-free and platelet-product-free plasma, anticoagulated by acid citrate dextrose formula A anticoagulant (ACD A), were collected using a combined membrane filtration/centrifugation plasmapheresis device (the HemaSciences "Autopheresis C"(R)). Samples of this plasma were heparinised (over a range from0.1 to 12.8 iu/ml) and physiological Ca2+ levels were restored, levels of fibrinopeptide A (FpA) of less than 5 ng/ml were measured in all samples(including following the incubation of plasmas at 21 °C for 24 hours). This indicated minimal thrombin generation at plasma heparin concentrations as low as 0.1 iu/ml. Fibrinogen degradation fragment BJ3 1542 levelsin these plasmas were comparable to those in ACD A control plasmas and there was no upward trend with increasing heparin concentration. This suggested that heparin-induced formation of plasmin would not adversely affect the stability of factor VIII:C in plasma treated in this way. Cryoprecipitates were prepared in a routine manner from ACD A plasmas collected, by plasmapheresis, into bags containing sufficient heparin and Cacl2 to achieve plasma heparin levels of 0.1 iu/ml and physiological Ca2+ concentrations. (As a result of thissecondary heparinisation, there was no infusion of heparin into the donors). Analysis of the cryoprecipitates revealed no resolubilisation problems, a significant (P<0.02) gain inthe yield of factor VIIIsC comparedwith ACD A control cryoprecipitates, and a mean factor VIIIsC specific activity of 0.11 iu/mg of total protein (S. D. 0.03, 6 experiments). Pools of plasma collected in the above manner are currently being fractionated to evaluate the resulting factor VIII concentrates.
MEASUREMENT OF HEPARIN IN PLASMA: INFLUENCE OF INTER-SUBJECT AND CIRCADIAN VARIABILITY IN HEPARIN SENSITIVITY ACCORDING TO METHOD
Authors: Scully M F , Decousus H A , Ellis V , Girard P , Parker C , Kakkar V V
Published: June 2019

Heparin was measured, with respect to standard curves prepared with normal pooled plasma, by five methods, APTT, (Auto-APTT), thrombin time (Thromboquik), one (Heptest) and two-stage (Hepaclot) coagulation, anti-factor Xa and chromogenic anti-factor Xa after addition at three concentrations (0.2, 0.4, 0.6iu/ml) to plasma prepared from three groups of ten individuals: normal young volunteers, hospitalized patients with malignancy and hospitalized geriatric patients. By the APTT and TT, differences in sensitivity were observed, for example, at 0.4iu heparin/ml corresponding to an apparent difference in heparin level of 10- and 14-fold between high and low responding individuals. All low responders, by APTT and TT, were in the older hospitalized group, tending to overheparinization when monitoring by these methods. Such large differences were not apparent by any of the anti-factor Xa assays. A circadian difference (1.9 and 1.8 fold) in sensitivity was also observed in the patient group such that in samples taken at night (12 pm), heparin levels were 30-50% higher than those taken during the morning on average when measured in the APTT and TT (p,0.001, analysis of variance). Again, such large differences were notapparent by anti-factor Xa methods. In light of recent findings about the usefulness of anti-factor Xa methods for efficient monitoring of heparin, it issuggested that this conclusion may arise from the tendency for anti-factor Xa methods to determine actual concentrations of heparin.
QUALITY ASSURANCE OF THERAPEUTIC HEPARIN/ORAL ANTICOAGULANT THERAPY IN A GENERAL TEACHING HOSPITAL
Authors: Brien W , Inwood M , Dillon K
Published: June 2019

In preparation for the development of a standard anticoagulation protocol and as part of a quality assurance programme, a retrospective analysis of anticoagulation therapy in a teaching hospital was undertaken. The results of this audit were compared to the results of other workers who have previously emphasized controversial areas of anticoagulation therapy and potential benefits of a standard anticoagulation protocol.

Charts from seventy-nine patients with a diagnosis of venous thrombosis were analyzed. Fifty-five percent of the patients were at increased risk of bleeding and forty-five percent had an extensive proximal vein thrombosis, but not apparent consideration of a heparin dose adjustment was noted. Patients received heparin for an average 9.7 days, with concomitant coumadin therapy for an average 4.8 days. Patients on heparin were over/ or undercoagulated on average 5.4 days and ninety-four percent of patients on coumadin had a PT (rabbit brain) greater than 18 seconds for at least one day. Sixty percent of patients had their dose changes noted in a tabular form for easy reference. Less than fifty percent had their hemoglobin or platelet counts followed while on heparin. The APTT and TCT are used to follow heparin therapy and discrepant results were noted in thirty-three percent. All patients' discrepancies were investigated automatically by the coagulation laboratory. Less than thirty percent of patients received formal teaching from the clinical pharmacist concerning coumadin therapy.

We feel that these results show a need for a standard protocol for heparin coumadin therapy. It has been shown that a standard protocol favourable chemical, financial and therapeutic benefits
PHARMACOKINETIC BEHAVIOUR OF ANTITHROMBOTIC AND BLEEDING ENHANCING EFFECTS OF VARIOUS HEPARIN(OID)S IN RATS: THEIR RELATION WITH INTERACTIONS WITH FACTORS IIa AND Xa
Authors: Hobblen P M J , Vogel G MT , Meuleman D G
Published: June 2019

The effects on thrombus formation and bleeding of various heparin(oid)s were studied at several time-intervals: in addition, the αXa-, αiia-, and thrombin generation inhibitory (IIaGI)-activities were measured amidolytically in plasma from these animals. The following substances were studied: heparin (HEP), the fraction of heparin with high affinity for ATIII (HA-HEP), a low molecular weight fraction of heparin (LMW-HEP), nitrous acid degraded heparin (NAD-HEP), FragrainR (FRA), Org 10172 and the fraction of Org 10172 with high affinity for ATIII (HA-10172). The substances were given in a dose of 800 αXa U/kg i.v. in the thrombosis model and in the double dose (1600 αXa U/kg i.v.) in the bleeding model because of the lower intrinsic effect on bleeding. The initial magnitude and the duration of the anti-thrombotic and bleeding enhancing effects of these substances are summarized in the table together with the half-lives of the αXa-, αia- and IIaGI-activities. NE: not estimated due to too low alla-activities.

The following conclusions could be drawn concerning the duration of the effects: 1. in contrast with HEP most lower molecular weight substances showed different time courses of the anti-thrombotic and the bleeding enhancing effect 2. the duration of the antithrombotic effect of Org 10172 and HA-10172 is considerably longer than that of the other substances 3. the duration of the bleeding enhancing effect of Org 10172, HA-10172, NAD-HEP and FragminR is shorter than that of HEP, HA-HEP and LMW-HEP. Additional conclusions could be drawn by inspecting the time response curves and taking the magnitude of the effects into account: 4. Org 10172 and its high affinity fraction have a better benefit/risk ratio in comparison with the other substances, which even increases in time 5. the time-response curves of the anti-thrombotic effects seem to be associated with those of the aXa-activities and 6. the time-response curves of the bleeding enhancing effects seem to be related to those of thrombin inactivation.
BLEEDING EFFECTS ASSOCIATED WITH HEPARIN contaminants
Authors: Pangrazzl J , Oreste P , Torri G , Maggi A , Donati M B , Casu B
Published: June 2019

Clinical use of heparin as an antithrombotic drug Is limited by the risk of excessive bleeding, generally ascribed to the anticoagulant activity of this mucopolysaccharide.

Unexpectedly, some low molecular weight heparins with reduced anticoagulant activity are reported to cause bleeding in clinical and experimental studies. To approach this problem, a number of heparin preparations with various molecular weights were characterized by analysis of their 13C-NMR spectra. The bleeding potential of the same heparins was tested by measuring the "template" bleeding time (BT, a method exploring the mechanisms of primary haemostasis) in the rat tail, 15 min after i.v. administration of 0.75 mg/kg b.w. of the drug.

Besides major NMR signals associated with known units of the regular and irregular regions of heparins, some of the preparations display 13C-NMR "extra-signals", which have a higher degree of relative intensity in the spectra of samples with higher haemorrhagic potential; indeed, all the samples exceeding in "extrasignals" a conventional intensity level of 2 had at least 70% prolongation of the BT over controls.

These results suggest that specific contaminants, responsible for these13C-NMR "extra-signals", may also be responsible for the unusually high bleeding expressed by some heparin preparations. Work is in progress to identify these components and to evaluate which mechanism of primary haemostasis is involved in their haemorrhagic effect.
THE EFFECT OF MOLECULAR WEIGHT ON THE RELATIVE BIOAVAILABILITY OF HEPARIN
Authors: Emanuele R M , Fareed J
Published: June 2019

Three fractions of different molecular weight (M. W.) were obtained from gel-filtration of porcine mucosal heparin. These fractions along with unfractionated and a low M. W. heparin (CY 216) were compared for relative bioavailability in primates (Macaca mulatta: n = 5). The M. W.'s of all fractions were determined using high performance liquid chromatography - gel permeation and characterizied in terms of mean M. W., peak M. W. and M. W. distribution. Area under the plasma concentration time curve (AUC) was calculated by the trapezoidal method after intravenous and subcutaneous administration and used as an index of bioavailability. For the AUC calculations, plasma heparin concentrations were determined using a Xa/IIa clotting assay (Heptest) and individual callibration curves. Using the AUC data, relative bioavailability was calculated using the following formula:

Bioavailability = subcutaneous AUC / dose intravenous AUC / dose Significant differences in the bioavailability of the various molecular weight fractions were observed and inversely related to M. W. The highest M. W. fraction (23,000) showed the least bioavailability at 3 %. The bioavailability of the 13,300 M. W. fraction was 9 % while the 5,100 M. W. fraction was 93 % bioavailable. CY 216 (M. W. 5,400) displayed a relative bioavailability of 88 % while unfractionated heparin (M. W. 12,600) was 40 %. The apparent discrepancy in the bioavailability of the unfractionated heparin is likely due to differences in molecular weight distribution.

These studies suggest that the M. W. of heparin preparations influences the relative bioavailability. Heparins of lower M. W. exhibit greater relative bioavailability compared to high M. W. preparations. Furthermore, characterization of both mean M. W. and M. W. distribution are necessary for predicting the bioavailability characteristics of heparin. The relationship between relative bioavailability and the M. W. of heparin may help identify low M. W. heparins as a more efficient agent for subcutaneous regimens in the prophylaxis of venous thrombosis.
THE ROLE OF A 3-0 SULFO GROUP IN THE GLUCOSAMINE RESIDUE OF A SYNTHETIC OLIGOSACCHARIDE FOR THE DETERMINATION OF ANTITHROMBOTIC ACTIONS
Authors: Walenga J M , Fareed J , Petitou M , Lormeau J C , Samama M , Choay J
Published: June 2019

We have previously reported on the antithromboticaction of a chemically synthesized heparin pentasaccharide which exhibits high affinity to anti thrombinIII and sole anti-factor Xa activity. In order to investigate the relative importance of the 3-0 sulfo group of this pentasaccharide, we evaluated the in vitro and in vivo antithrombotic activity of a synthetic pentasccharide devoid of the sulfo group at the third position of the glucosamine residue. In amidolytic and clot-based assays the 3-0 de- sulfated pentasaccharide (3-0-DP) failed to exhibit any antifactor Xa actions at concentrations <100 ug/ml in humanor rabbit plasmas, whereas pentasaccharide showed strong factor Xa inhibition at 1.0 ug/ml IK-=3.2x10 M)and at 10.0 ug/ml in rabbit plasma (K.=9.0×10™7 M). Using a rabbit stasis thrombosis model in which thrombosis was induce by human serum or an activated pro-thrombin complex concentrate, 3-0-DP failed to produce any antithrombotic action in acute intravenous regimens at dosages up to 200 ug/kg. In these two models, pentasaccharide produced >80% inhibition of induced thrombosis. These studies demonstrate the critical importance of the 3-0 sulfo group in this heparin pentasaccharide for the determination of antithrombotic activity, and that in this type of oligosaccharide, anti-factor Xa activity is responsible for producing the antithrombotic effect.
LOW DOSE HEPARIN PROPHYLAXIS IN HIP FRACTURE SURGEY -HEPARIN EFFECT; INHIBITORS; FIBRINOLYSIS AND INCIDENCE OF DVT
Authors: Kußmann J , Spannaql M , Boehnke J , Kückel-haus H G , Schramm W
Published: June 2019

In a prospective studie 129 patients with hip fracture surgery under LDH-prophylaxis (3x5000 U Na-heparinat) were examined in order to find an answer tothe question, if there is a correlation between inhibitor activity, parameters of fibrinolysis, plasma heparin activity and the incidence of DVT.

100 patients with ascending phlebography on day 7 to9 post op. were taken into final consideration (blood collection on admission and on day 1, 2, 4, 7 after surgery):

1)Incidence of DVT : 17 %.

2)Inhibitors: No difference between patients with and without DVT with respect to AT III activity and prot. C concentration ( prot. C activity in progress).

3)Fibrinolysis: Elevated levels of DD-fragment (x = 1 780 mg/ml), t-PA Inhibitor (x = 31 AU) and fibrinogen (417 rng/dl) before operation due to preceding trauma. No significant difference between t-PA, t-PA inhibitor and antiplasmin with respect to DVT. Whileplasminogen concent was significantly increased in patients with DVT on day 4 and 7, DD-fragments had lower values on day 7 (x = 1395 / x = 2140 ng/ml).

4)Heparin effect: Plasma heparin activity was assesed by an amidolytic anti ll^assay. Although plasmaticheparin action only represents one aspect of thromboprophylactic heparin-activity, there is an obvious difference between patients with and without DVT withrespect to plasmatic heparin activity( p <0.005).
PREDICTION OF POST-OPERATIVE DVT BY SALINE DILUTION
Authors: Chayen D , Blair S D , McCollum C N , Greenhalgh R M
Published: June 2019

Clinically, it is difficult to predict deep vein thrombosis (DVT), but the in vitro saline dilution test using the Thrombo-elastograph (TEG) is reported to identify the risk for individual patients [1]. The Biobridge Impedance Clotting Time (ICT) is more sensitive and reproducible than the TEG [2], and we therefore studied 33 patients undergoing elective laparotomy to see if pre-operative saline dilution tests using both the TEG and ICT predicted post-operative DVTs. Post-operatively, both legs were scanned daily for 7 days using 125I Fibrinogen to detect DVTs.

The mean age of the patients was 65.7±2.4 years and 17 had malignant disease. In this clinically high risk group, 24 developed a DVT.

Fifty-one percent were predicted correctly by TEG. The ICT was significantly better as a predictor with 79% of all patients correctly predicted (p<0.01).

The saline dilution test using the ICT is a significant improvement on the TEG, and may enable us to tailor DVT prophylaxis policy to each patient’s specific requirements.

Heather BP, Jennings SA, Greenhalgh RM. The saline dilution test - a preoperative predictor of DVT. Br J Surg 1980; 67: 63-65

Blair SD, Menashi S, Samson D, Greenhalgh RM. Can the hypercoagulability of surgery be measured? Br J Surg 1986; 73: 500.
EFFECT OF PIRACETAM ON THE MEAN INTRASPLENIC RED CELL TRANSIT TIME (MST) IN SICKLE CELL DISEASE AND SICKLE CELL THALASSAEMIA
Authors: Ferrant A , Leners N , Gini E K , Osselaerey J P , Sonnet J
Published: June 2019

The effect of Piracetam on the MST has been evaluated in 3 children with homozygous sickle cell disease, and in 1 child and 2 adults with sickle cell β thalassaemia. The MST was measured using 99 Tern labelled autologous red cells before and during treatment with Piracetam (160 mg/kg/d). Tests for in vitro fi1terabi1ity of red cells were performed and an improvement of the in vitro deformabi1ity was observed in all the patients.

A shortening of the MST was also observed :
EFFECTS OF FLOW PULSATILITY ON PLATELET ADHESION TO SUBENDOTHELIUM
Authors: van Breugel Hans H F I , Sixma Jan J , Heethaar Robert M
Published: June 2019

Platelet adhesion studies in perfusion chambers are generally performed with constant flow rate or with pulsatile flow generated by a roller pump. No information is available about the effect of flow pulsatility in the physiological range. For this purpose we devised a system consisting of a eccentric rotating disc which forces a driver toward a flexible tube. This movement causes a sinusoidal laminar flow component. The effect of this sinusoidal pulse on platelet adhesion was studied with the annulai^perfusion chamber of Baumgartner using umbilical arteries and In-labeled platelets. Conditions used are described below, where γ [1/s] is the constant flow component, γ0 [1/s] is the amplitude of the sinusoidal component and f [beats/min] is the frequency of the pulse (70 beats/min standard). Adhesion values are expressed as 105 III In platelets/cm2 (n=4).

When osciallatory flow (abence of constant component) was generated, platelet adhesion incersed with increasing amplitude of the pulse.

These data indicate that pulsatility as persent in the the aorta and large arteries has no effect on platelet adhesion.
PLATELET DEPOSITION ONTO FIBRIN-COATED SURFACES UNDER FLOW CONDITIONS
Authors: Jen C J , Chiu Y L
Published: June 2019

Fibrin solubilized in NaBr/acetic acid was used to coat glass tubes. Platelet deposition on fibrin-coated surface and release from these adherent platelets were studied in an in vitro flow system. When a mixed suspension of washed platelets and red cells flowed through a fibrin-coated glass tube, only platelets deposited onto the fibrin-coated surface. The density of adhered platelets increased with flow time and decreased with distance from the tube inlet. The adhesion rate increased with increasing shear rates from 45 s−1 to 180 s−1. This adhesion process appears to fit a diffusion-limited mathematical model. Comparing with glass and other protein-coated surfaces such as collagen, fibrinogen, or albumin coated surfaces, the number of adhered platelets per unit area decreased in the following order: collagen > fibrin> fibrinogen > glass > albumin. On the other hand, the degree of release reaction from these platelets decreased by another order: collagen > glass > fibrinogen > fibrin. We observed little release from platelets that were in contact with a fibrin-coated surface. Our results support that platelets specifically adhere to fibrin-coated surface and that this interaction does not induce platelet release.
COMPARISON OF A MODIFIED BLEEDING TIME/ “HAEMOSTASIS TIME” (HT), WITH THE DUKE AND IVY/TEMPLATE BLEEDING TIMES IN HAEMOPHILIA AND VON WILLEBRAND’S DISEASE
Authors: Janzarik H , Heinrich D , BÖdeker R H
Published: June 2019

The occlusion time of a Butterfly 25 short cannula inserted into the cubital vein was assessed as a new bleeding time modification. The method excludes the influences of vessel wall and tissue factors since primary haemostasis takes place on the standard surface of the cannula. Avoiding scar formation, the technique can be repeated deliberately. It could be shown that HT correlated better than the Duke and Ivy/ template bleeding times with reduced platelet counts and reflected the effects of aspirin and unfractionated heparin in the same way (Janzarik et al., Blut 1986, 52:345).

HT was applied simultaneously with the Duke and Ivy/ template or Simplate bleeding times in haemophilia and von Willebrand's disease (vWD). According to most of the other authors, the skin bleeding times were normal in patients with haemophilia. However, HT was prolonged in 9/16 patients irrespective of the factor VIII:C/IX:C values. The duration of HT correlated significantly to the onset of bleeding symptoms and to the degree of spontaneous bleeding (e.g., haemarthro-ses). In 31 patients with clinically mild or moderate vWD, the Duke bleeding time was prolonged in 3/31 patients, Simplate II in 13/31 patients and HT in 14/31 patients. With the two latter methods, corresponding results were observed in 20 cases, different results in 11 cases. HT and both the other methods correlated significantly to f. VIII:RCo and VIII:C but not to VIII: vWF

We conclude that the repeatable HT method is an interesting diagnostic parameter in haemophilia and vWD. With the Simplate and HT methods simultaneously performed, the influence of vessel wall factors on primary haemostasis may be differentiated.
EVALUATION AND STANDARDISATION OF THE “IN VITRO” BLEEDING TIME TECHNIQUE
Authors: Kratzer MA A , Knedel M
Published: June 2019

Recently a model of primary hemostasis has been introduced (Kratzer & Bom, Haemostasis 15:357;1985). This in vitro method detects very sensitively pathological platelet functions (Kratzer, Bellucci & Caen, Haemostasis 15:363; 1985). In order to use this technique routinely in the hospital the instrument now called “Thrombostat 4000” has been completely computerized and standardized by Kratzer and von der Goltz, VDG (8221 Seeon, West Germany).

An artificial vessel (Aperture: cellulose acetate coated with collagen typ I, 2.2 ug/ mm2) with a single 150 urn hole; capillary: teflon, 200 urn, length 20 mm) was perfused with anticoagulated whole blood (Na-citrate 1 to 10) using a pressure of 40 mbar. To determine the reproducibility of the new technique, in vitro bleeding volume (V), time (T) and flow at the beginning of the experiment (IF), depending on blood viscosity, was measured in a healthy control person at different days (mean +− standard deviation, n= number of experiments).

The mean of the intraassay variance (%) was: V (7.1), T(6.8) and IF(3.3). It was possible to measure the mean from day to day with the following variances (%): V(6.2), T(4.7) and IF(4.1). The low variance, which approaches enzymatic determinations is astonishing because platelet cell functions depend on a complex interaction of several thousands of enzymes.

The excellent assistance of P. Gossweiler is greatly aknowledged
PILOT STUDY WITH AN “IN VITRO BLEEDING TIME” (“IN VITRO BT”) TO MONITOR THE USE OF A STABLE PROSTACYCLIN (PGI2 ANALOGUE IN HAEMODI ALYSIS (HD)
Authors: Maurin N
Published: June 2019

Administering PGI2 instead of heparin (H) during HD enables the risk of bleeding to be reduced. A serious disadvantage of this method of inhibiting coagulation by inhibiting thrombocyte function is however that, in contrast to anticoagu1 ation with H alone, there exists no quickly measurable monitoring parameter. Recently Kratzer and Born (Haemostasis 1985; 15: 357-52) presented a new method for measuring the primary haemostatis: the “in vitro BT”. Our pilot study was to determine whether this “in vitro BT” is concentration - d epe nd ent 1 y prolonged by administering a stable PGI2 analogue during HD. In 5 HD’s the stable P G I 2 analogue CG 4203 (dose gradually raised starting 45 rfnn before HD to 25 ng/kg b.w./min during HD; Gruenenthal GmbH, FRG) were given. Before, during and after the HD the “in vitro BT” (Thrombostat 4000, VDG, FRG) was measured under standardised conditions 15 min after taking 1 ml of citrate blood from the arterial branch of the extracorporeal system. The “in vitro BT” was concentration-dependent!y prolonged by CG 4203. The time interval between taking and measuring the blood sample affect the measured value. If the blood sample is taken directly before the dialyser, the “in vitro BT” is greater than if the sample is taken directly after the dialyser; from which it can be assumed that the dialyser activates the platelets. 1 h after ending HD and discontinuing the CG 4203, the “in vitro BT” is shorter than it was before starting HD; perhaps this is due to platelet activation during HD in the extracorporeal system and to removal of uraemic toxins. Furthermore, the “in vitro BT” was measured in a healthy volunteer 20 times consecutively. The coefficient of variability was 9.3 %. Further studies should be performed to see whether adjusting the “in vitro BT” to a given range allows the dosing of PGI2 or a stable PGI2 analogue to be so controlled as to prevent occlusion in the extracorporeal system.
THE EFFECT OF ANTICARDIOLIPIN POSITIVE SERA FROM SLE PATIENTS ON PLATELET AND ENDOTHELIAL PROSTANOID SYNTHESIS
Authors: Hasselaar P , Blokzijl L , Derksen RH W M , de Groot Ph G
Published: June 2019

A strong correlation has been found between the presence of lupus anticoagulant (LAC) and/or anticardiolipin antibodies (ACA) and the development of thrombotic complications in SLE patients. One of the mechanisms suggested for the increased risk for thrombosis is an inhibition of endothelial cell (EC) prostacyclin (PGI2) synthesis by ACA. In these studies, no attention has been paid to the effects of ACA on platelet thromboxane (TxA2) synthesis. To compare the effects of ACA on PGI2 and TxA2 synthesis we have incubated human EC and platelets with 3 different control sera and sera of 11 different patients with SLE (of which 9 had a history of thrombosis). EC PGI2 synthesis was stimulated with PMA (24 hours) or arachidonic acid (AA) (10 min), platelet TxA2 synthesis was stimulated with collagen (5 min) or AA (1 min). The results are summarized in the table.55% of the sle sera inhibited PGI2 release and 65% stimulated TxA2 production. There is no correlation between these phenomena. The presence of ACA/LAC in the sera does not always result in an abnormal prostanoid production. Both decreased PGI2 production and increased TxA2 synthesis could result in an increased risk for thrombosis. However, why some SLE sera inhibit PGI2 production in EC and stimulate TxA2 production in platelets remains unexplained.
DETECTION OF LUPUS ANTICOAGULANTS BY THE “INSIDE-OUT MEMBRANE ADSORPTION TEST”: aPTT IS SHORTENED AFTER INCUBATION OF THE PLASMA WITH THE ADSORBENT
Authors: von Felten A
Published: June 2019

Prolongation of the aPTT in a mixture of a patient’s plasma with normal plasma is characteristic for the presence of a coagulation inhibitor. Since inhibitors directed against individual blood coagulation factors differ in their clinical significance from a lupus anticoagulant (LA), a fast and reliable distinction from each-other would be highly desirable. The test presented which is based on the absorption of antiphospholipid antibodies by use of a stable phospholipid adsorbent largely fullfills this demand.

Method 1. Preparation of the LA-adsorbent (modified acc. to Steck et al., Science 168:255, 1970): Fresh human RBC (bl. gr. 0, in 0.5% EDTA) are washed (NaCl, 0.15m / phosphate buffer,0.005m, pH8.0) and lysed in phosphate buffer alone. The membranes are washed with the same buffer (centrifugations: 40’000g, lOmin, 4°C) and incubated 60 min on ice in phosphate buffer, 0.0005m. After centrifugation, the sediment is resuspended in an equal volume of buffer (0.0005m, with merthiolate 0.1%), homogenized and centrifuged in portions of lml; the supernatant is carefully removed. The adsorbent, stored at 4°C in capped tubes, is stable for at least three months.

2. Performance of test: To 600ul of patient’s plasma mixed with 600ul normal plasma (citrated), 12ul of heparin (501U/ml) are added. 600ul of the mixture are given to the LA-adsorbent, vorte-xed, incubated (room temp., 30min) and centrifuged (40’000g). From the supernatant as well as the not adsorbed portion of the plasma mixture, heparin is eliminated by adding ECTEOLA-cellulose. After centrifugation, aPTT is determined in both supernatant samples. Results The test is positive, i.e. LA is present, if aPTT is shortened by at least 5 sec after treatment with LA-adsorbent.

*not diluted; other samples are mixed 1:1 with normal plasma
HIGH INCIDENCE OF CARDIOLIPIN-BINDING ANTIBODIES IN PATIENTS TAKING ORAL ANTICOAGULANTS
Authors: Koutts K , Exner T
Published: June 2019

It is known that patients with thrombosis in autoimmune disease often suffer recurrent thrombotic episodes on withdrawl of oral anticoagulants, frequently have raised ACA and may require long term therapy. In view of the association between lupus inhibitors, phospholipid binding antibodies and thrombotic episodes we investigated a broad group of patients (n=140) taking oral anticoagulants for anti-cardiolipin antibodies (ACA).

The incidence of raised ACA (defined here as more than 5 S. D. above normal) was 10%, in comparison with less than 2% in both healthy volunteers and randomized hospital patients. Analysis of patients with raised ACA indicated an approximately equal distribution among those with artifical valves and patients treated for thrombotic episodes. ANF and DNA binding studies were negative in most cases and a lupus inhibitor (KCT mixing test) was detectable only in 1/15. APTT/PT ratios were not significantly high in comparison with other clinic patients. The-ACA were predominantly IgG (12/15) in contrast to those fpund in females with recurrent abortions which are frequently IgM's. The binding antibodies appeared to be much weaker in the presence of calcium than in phosphate buffered saline.

The patients having high ACA had no excess thrombotic or bleeding complications during their treatment compared with others in this group. This suggests that thrombotic or occlusive events associated with ACA may be adequately treated with oral anti-coagulants.
INCIDENCE OF PROCAINAMIDE-INDUCED LUPUS ANTICOAGULANT
Authors: Day H J , Cherrey R , O'Hara D , Carabello J
Published: June 2019

Isolated cases of lupus anticoagulants (LA) in association with procainamide have been reported. This study was done to estimate the frequency of LA in patients taking procainamide. Two groups of patients were evaluated: Group A: 110 hospitalized patients (84 males, 26 females, ages 51-78, mean age 74.3) and Group B: 80 ambulatory patients (54 males, 26 females, ages 36-89, mean 67.5 years). The latter group of patients was on this drug for a period of two-five years, while the former group hadbeen on drug at least two full days. All patients were screened with baseline laboratory data including activated partial thromboplastin time (APTT) and prothrombin time (PT) which were performed using Auto APTT® and Simplastin® on a Coagulamate X2® (General Diagnostics/Organon Teknika). Patients taking drugs known to alter the APTT and PT were excluded. All patients were followed with daily (hospital patients) or weekly (ambulatory patients) APTT and PT. Prolongation of the APTT of 5 sec or PT of 3 sec over baseline was considered as a positive LA screening test. Patients with a positive screening test were further evaluated with tissue thromboplastin inhibitor assay (TTI), platelet neutralization procedure (PNP, anti-nuclear antibodies (ANA) and blood serology (RPR). In Group A. 11 out of 110 (1096) developed prolonged APTT while on procainamide . Of these, 9 developed abnormal TTI and 2 had positive PNP. The ANA was positive (titers of 1:320-1:2560) in 10 patients with the only positive RPR test, being in the patient with the highest ANA titer. In Group B, 12 out of 80 (1596) developed prolonged APTT, 11 had positive TTI. The ANA titer was elevated in all positive cases, although one patient's titer was 1:30. The PNP was positive in 1/12. All blood serologies were negative in this group. The difference in incidence between Groups A and B may reflect longer exposure to drug in the latter group. This difference is not statistically significant. This study indicates that the incidence of procainamide-induced lupus antocoagulant is between 10-1596 when the APTT (or PT) is used as a screening test. The TTI and ANA seem to have equal sensitivity in this syndrome when taking this drug. The failure of the PNP to be sensitive to LA may be due to the minimal prolongation of the APTT arbitrarily chosen as representing a positive screening test.