bluuebunny/crossref_metadata_2025 huggingface dataset:
FIBRONECTIN-FIBRIN-COMPLEXES : AN IMMUNOASSAY TO EVALUATE THEIR FUNCTION IN COAGULATION DISORDERS AND TUMOR SITUATION
Authors: Wilhelm O , Hafter R , Graeff H
Published: August 2018

Fibronectin and fibronectin degradation products interact with fibrin(ogen) by non-covalent and by factor XIII catalyzed covalent binding. In certain physiological or pathophysiological situations these complexes may be of considerable importance. It was the aim of this study to accumulate more information of the function of this proteininteraction. Immunoadsorption in combination with Westernblotting with different monoclonal and polyclonal antibodies demonstrate several fibronectin-fibrin(ogen)-complexes in tumor ascites. For further information we established an immunoassay to measure these complexes. As the capture antibody we used a polyclonal fibronectin antibody and as the tag antibody a monoclonal fibrin(ogen) antibody. In this immunoassay we observed a decrease of these complexes from very high values during the therapy of disseminated intra-vascular coagulation besides also a decrease of high molecular weight fibrin derivatives and fibronectin split products. We also found elevated levels in ascitic fluid and plasma of patients with advanced ovarian cancer (x=202,3± SD 45.1 ng/ml fibrinogen equivalent) in comparison to a control group with benign gynecological diseases (X= 102,9± SD 14,9 ng/ml fibrinogen equivalent). In conclusion we suggest that during intravascular coagulation fibronectin binds fibrin derivatives for elimination from plasma. In the tumor situation these complexes may stem from the fibronectin-fibrin-gel-matrix surrounding the tumor by proteolytical degradation and subsequent release into the ascites, (supported by Deutsche Forschungsgemeinschaft, SFB207, A2).
INFRACLINIC ACTIVATION OF PLATELETS AND FIBRIN FORMATION IN CANCER PATIENTS
Authors: David J L , Lambrichts M , Closon M T
Published: August 2018

Thromboembolism has been frequently reported in cancer patients, mainly in cases with solid tumors. Besides in several animal models, fibrin deposition around the tumor and platelet aggre gates appear to be involved in invasion and metastasis. This study was aimed at evaluating the extent of in vivo platelet activation and fibrin formation in several kinds of human cancer. We excluded from this study patients whose blood was sampled with difficulty as well as those having clinical evidence of thrombosis or embolism, those with thrombocytopenia, increased fibrinogen degradation products or biological pattern of disseminated intravascular coagulation. Fibrinopeptide A (fpA) and β-thrombo-globulin (β-tg) were determined by RIA. Free platelet count ratio (PCR) was determined on whole blood samples as an index of circulating aggregates. Usual coagulation tests, antithrombin III activity, protein C plasma level, F VIII related antigen (F VIII RAG), F VIII Ristocetin cofactor (F VIII RCF) and F VIII procoagulant activity (F VIII C) were also determined.

It was found that in more than fifty percent of patients, fpA was significantly increased above the upper reference limit. Cases with increased B-tg were less frequent. Separate increases in β-tg or fpA levels were often observed. PCR remained within the reference values in almost all patients. F VIII RAG, RCF and C were usually above 150 % of the reference mean.

We conclude that platelet release and fibrinoformation frequently occur in cancer patients showing no sign of thrombotic process. Increased level of fpA with normal plasma β-tg level suggests that thrombin generation occurs only in the extravascular compart ment, probably next to the tumoral tissues. Increased levels of plasma β-tg with normal fpA levels may result from platelet activation by other stimuli than thrombin. It must be emphasized that normal PCR does not exclude the presence of fibrinous circulating aggregates which cannot be dispersed by EDTA. High F VIII activities may be due to the release of the von Willebrand factor from tumoral vessels.
ALPHA-2-ANTIPLASMIN (α2AP) IN ACUTE NONLYMPHOBLASTIC LEUKAEMIA AS A MARKER OF DISEASE ACTIVITY
Authors: Cofrancesco E , Poliani E , Salvatore M , Boschetti C , Moreo G , Cortellaro M
Published: August 2018

α2AP (Coatest Antiplasmin Kit, Ortho Diagnostic Systems), antithrombin III (Atlll, Coatest Antithrombin Kit, Ortho Diagnostic Systems) and plasminogen (S2251) were assayed in 21 patients with acute nonlymphoblastic leukaemia (ANLL: 5M2 5M4 5M5) before and after chemotherapy and during bone marrow cellularity recovery. The aim of the work was to investigated disturbances of coagulation and fibrinolysis with special reference to proteases inhibitors and to evaluate the prognostic value of changes in these parameters in ANLL patients. Low α2AP levels were observed in the initial phase of the disease: Ul.5 ± 2k.79 SD in 10 DIC patients versus 6U.10 !± 20.70 SD in patients without DIC (p < 0.05). α2AP normalized only in the 11 patients who achieved haematological remission (71.63 ± 13.13 SD) and remained low in those who did not respond to chemotherapy (6l.88 ± 17.86 SD, p < 0.01). No significant modification of Atlll and plasminogen levels were observed during the course of the illness.

It may be postulated that proteolytic cleavage of α2AP by granulocyte proteases contributes to the low levels of the inhibitor in ANLL, and suggested that α2AP may represent a marker of leu-kaemic disease activity. In fact the mean α2AP level of all patients at diagnosis plus those who did not respond to chemotherapy was significantly lower than that of patients in haematological remission (55.93 ± 23.13 versus 71.63 ± 13.13, p < 0.05).
FACTOR II ACTIVATING ACTIVITY IN EXTRACTS OF TUMORAL NECROSIS FROM TWO MURINE BREAST ADENOCARCINOMAS
Authors: Blanco A , Bonfil R , Bustoabad O , Lazzari M
Published: August 2018

Increased deposition and lysis of fibrin, associated with malignant tissue, has led to look for activators of both the coagulation and fibrinolytic systems produced by tumor cells. We report the evidences of a procoagblant activity (PA) in the extracts of intratumoral necrosis from two experimental breast adenocarcinomas in murine model (BALB/c). The tumors have different metastatic capacity (MC). M3 without MC and MM3 with high MC.

The addition of the extracts to: 1- Normal Plasma, 2- Deficient substrates in coagulation factors, 3- Purified, fibrinogen (I), showed: 1- Shortening of the plasma recalcification time (PRT) and APTT, without ;modification on prothrombin time (PT), 2- Reduction of the PRT on deficient substrates in factors: VIII; VII; VII and X; V; V, VII and X; without modification on II deficient substrate, 3- No PA on I. Table:

C: Control, s: seconds, m: minutes. The PA was not affected by heparin. The results suggest that the PA is independent of the presence of either factor VIII or factor VII (intrinsic or extrinsic pathway respectively), as well as presence of either factor V or factor X. Any effect was observed either on factor II deficient substrate or on I, so, there was no evidence of thrombin activity The PA could be act directly on factor II, suggesting that fibrin formation could be induced by a “non-classical” activation pathway. No significant differences (p>0.5) in PA were observed between both tumoral necrosis extracts. The necrotic area in M3 (37%) is bigger than in MM3 (18%). So, much more PA could be present in MM3 and this could play a role in the MC of this tumor.
EXPERIMENTAL RESEARCHES OF EFFECTS OF POLYACTIN A ON HEMOPOIETIC STEM CELL OF MARROW
Authors: Xiang-yuan Zeng , Ying Zhang
Published: August 2018

Polyactin A is a new immunological enhancement agent which was developped in China for the first time1 It is a polysaccharide extracted from cultured a-hemolytic streptococcus No.33 the mouth.Clinical observation suggests that the drug has marked inhibiting effects on some tumors and can increase the number of leucocytes, enhance immunity of the organism.Polyactin. A has especially good effect on aplastic anemia and it is a new drug in treatment of aplastic anemia.However, it mechanism isn’t understand.

Effects of Polyactin A on hemopoietic stem cells of marrow are studied. The hemopoietic stem cells (CFU-s) were measured by method forming colonies in the spleen after mice were irradiated. Experimental observations showed that spleen weight, the number of CFU-s and spleen index in the group of Polyactin A were notably more than those in the control group. Electron microscopic studies on the colonies of spleen were also presented. The results suggest that Polyactin A can make effects on hemopoietic stem cells of marrow and has good influence to stimulate hemopoiesis.

This study provided experimental evidence for use which Polyactin A can treat aplastic anemia in clinic.
EVALUATION OF CLOTTING PARAMETERS AND PHYSIOLOGICAL INHIBITORS IN PH /+ CHRONIC MYELOID LEUKEMIA PATIENTS UNDERGOING ALLOGENEIC BONE MARROW TRANSPLANTATION
Authors: Tirindelli M C , Arcese W , Mariani G , Papa G , Fossati C , Iacopino G
Published: August 2018

The aim of this study is to evaluate blood coagulation changes in Ph positive chronic myeloid leukemia (Ph + CML) patients undergoing allogeneic bone marrow transplantation (BMT), T-depleted with the monoclonal antibody Campath 1, and to find a possible correlation between changes of procoagulant proteins, physiological inhibitors and venoclusive disease (VOD). VOD is a major complication in, the early period following BMT.

Out of 13 patients, two of them developed VOD. Von Willebrand Factor Antigen (vWF:Ag), Factor VII antigen (F.VII:Ag), Plasminogen antigen (PLG:Ag), Factor V activity (F.V:C), Antithrombin III activity, Protein C antigen (PC:Ag) and fibrinogen activity (FG) were evaluated in all patients following sampling times: day 10 (before conditioning regimen), day 0 (day of BMT), day + 10 (after BMT), day + 28 (median day of engraftment) and day + 60 (after BMT).

PLG:Ag, F. VII:Ag and PC:Ag levels decreased significantly (p values < .05, < .025, < .001 respectively). In particular, in 5 patients whose PC:Ag levels dropped below the limit considered at risk for thrombosis (< 60 U/dl), two of them developed VOD. vWF:Ag, F. V:C and FG increased significantly (p values < .04, < .05, < .05 respectively). Antithrombin III activity did not change significantly throughout the period of observation.

In conclusion, the reduction of PC, PLG and the contemporary rise of vWF and FG can increase the risk of developing thromboembolism in the early stage following BMT.
DEEP VENOUS THROMBOSIS PR0FYLAXIS WITH A LMW HEPARIN (KABI 2165) IN STROKE PATIENTS
Authors: Prins M H , den Ottolander G J H , Gelsema R , van Woerkom T C M , Sing A K , Heller I
Published: August 2018

In a group of 60 patients entering our hospital for completed stroke, within 72 hours after onset of symptoms, treatment with Kabi 2165 2x 2500 anti-Xa U s.c. was compared to placebo 2x s.c. in a double blind trial to test the assumption that Kabi 2165 could prevent DVT, without causing cerebral bleeding in the ischaemic area. The diagnosis of stroke was made on clinical grounds. A.CT-scan of the head was performed before entering the trial to exclude cerebral bleeding or tumor. Follow-up during a trial period of 14 days included a Fibrinogen scan - if positive followed by flebografy. After the trial period or when clinical deterioration occurred a CT-scan of the head was repeated. Before and during the trial period haematologic and coagulation data were obtained and will be reported. Obduction was obtained whenever possible.

The patient groups were comparable, except for a slight preponderance of disturbed consciousness and atrial fibrillation in the Kabi 2165-treated group. This difference did not reach statistic significance. In the Kabi 2165 group there were 6 cases of DVT compared to 15 in the placebo group (p=0,05). In the Kabi 2165-treated group there were slightly more cases of cerebral bleeding and death during trial, respectively 4 versus 2 and 9 versus 4 (both NS). Cerebral bleeding occurred only in patients with a bloodpressure above 150/90 mmHg on entering.

Although the patient group is still small, we like to conclude that in normotensive stroke patients Kabi 2165 2x 2500 anti-Xa U s.c. per 24 hours, is a safe method of DVT profylaxis.
THROMBOSIS PROPHYLAXIS WITH LOW MOLECULAR WEIGHT HEPARIN (KABI 2165) AND CALCIUM HEPARIN IN PATIENTS WITH TOTAL HIP REPLACEMENT
Authors: Potron G , Barre J , Droulle C , Baudrillard J C , Barbie P , Kher A
Published: August 2018

In a prospective controlled randomized trial efficacy and safety of a low molecular weight heparin (Kabi 2165) and calcium heparin were compared in 80 patients with total hip replacement. 40 patients were given Kabi 2165 : 2,500 anti-Xa Units S.C. 2 hours before operation and then every 12 hours for 10 days. 40 patients received calcium heparin : 5,000 iu S.C. 2 hours before operation, then every 8 hours during the first post operative day and then heparin doses were adjusted according to the activated partial thromboplastin time and the thrombin time for 10 days. Bilateral venography was performed routinely on all patients between the ninth and tenth post operative day. The two groups were well matched for risk factors which could predispose to risk of developing venous thrombosis. 7 patients in Kabi 2165 group (17,5 96) and 4 patients in calcium heparin group (10 %) developed D.V.T. (p = 0.33). In 3 patients (7,5 96) of Kabi 2165 group and in 2 patients (5 96) of calcium heparin group, the thrombi extented to the popliteal-femoral vein. The incidence is not significantly different (p = 0,50). There were no significant differences in postoperative mean estimated blood losses, and mean blood units transfused. Mean hemoglobin levels and mean hematocrit values at the day before operation, the 1st, 5th, and 10th day after operation were :

These differences were not statistically significant. Incidence of wound hematomas was similar in both groups No thrombocytopenia was reported in this study. In conclusion, Kabi 2165 2,500 anti Xa Units twice daily seems as efficient as adjusted-dose of calcium heparin thrice daily.
USE OF A LOW MOLECULAR WEIGHT HEPARIN (LMWH) KABI 2165 DURING HAEMODIALYSIS : DETERMINATION OF LOWEST EFFECTIVE DOSE-MEMBRANE DEPENDANT DOSAGE REGIMEN
Authors: Potron G , Toupance O , Droulle C , Chanard J , Kher A , Barbier P
Published: August 2018

The L.M.W.H., Kabi 2165 was compared to standard heparin in stable chronic haemodialysis patients in order to determine the lowest effective dose.

An initial dose of 1,000 to 2,500 anti-Xa units followed by an additional dose of 750 to 1,500 anti-Xa units, 1 or 2 hours after the start of the c’alysis was associated with clotting and bleeding complications on 156 dialysis performed in 8 patients: 33 partial coagulations of extracorporeal circulation and 16 minor bleeding events. This dosage regimen corresponds to half of that of standard heparin needed in the same patients to prevent clinical problems.

In 9 patients treated with haemodialysis for 4 hours or less using cuprophane membrane, a single dose of 80 anti-Xa units/kg given at the start of dialysis achieved a sufficient antithrombotic effect on 195 dialysis, but minor bleeding complications were observed in 4 cases.

On 270 dialysis perfomed in 12 patients using polyacrylonitrile membrane, it was necessary to increase by 1,000 anti-Xa units this previous dosage to obtain satisfactory effectiveness.

The biological activities measured in this study indicates that : rises in fibrinopeptide A and platelet factor 4 were associated with clotting in the extracorporeal circulation.

Increased levels of factors VIII : RAg and decreased levels of prekal1ierein in the predialysis blood sanples were corrected by Kabi 2165.

In conclusion, Kabi 2165 would appear to be an effective and easier antithrombotic agent in patients on chronic haemodialysis.
LOW MOLECULAR WEIGHT HEPARIN (FRAGMIN) PROPHYLAXIS IN GYNECOLOGIC SURGERY
Authors: Briel R C , Hermann P C , Doller P
Published: August 2018

In a prospective, randomized study patients undergoing hysterectomy were treated either by the low molecular weight heparin Fragmin or by the combination of unfractionated sodium heparin + dihydroergotamin (HDHE). The dosage in the Fragmin group was 2× 2500 anti Xa-U on day 1 = day of surgery, from day 2-8: 1× 5000 anti Xa-U, in the HDHE-group from day 1-8: 2× 5000 IU heparin + 0.5 mg DHE. 99 patients were randomly allocated to prophylaxis with Fragmin, 101 to HDHE prophylaxis. 95 and 96 respectively were evaluated, the others excluded for different reasons. The 2 groups were comparable for general data and risk factors. Duration of surgery, intraoperative blood loss, transfusion rates and postoperative hemoglobin levels were identical. Blood volumes in subcutaneous and subfascial drainages were slightly but not significantly higher in the Fragmin group. In patients with an additional Marshall-Marchetti-operation, blood volumes in the drainages of the spatium retzii were significantly higher in patients on Fragmin. No differences were observed in the incidence of minor and major wound hematoma. Painful injections and sugillations at the injection sites were more frequently observed in the HDHE-group. The thermographic DeVeTherm test, which was carried out daily for diagnosis of DVT, gave positiv results (= temperature difference 1°C) on one day only in 14 patients of each group. The test was positive on 2 or more consecutive days in 4 patients on Fragmin and 2 patients on HDHE. Phlebography, which was carried out in the latter patients, gave a positive result in 1 patient of each group. Localization of DVT was mainly the lower limb. Plasma anti-Xa activity (S-2222) 4 hrs. after injection of 5000 anti-Xa IU Fragmin was 0.45 IU/ml being 10 fold higher than after HDHE. aPTT was slightly prolonged in both groups, thrombin time and thrombelastogramm gave even more pronounced changes in the Fragmin group. The present data indicate that Fragmin dosage should be further decreased to avoid bleeding complications.
EFFICACY AND SAFETY OF A LOW MOLECULAR WEIGHT HEPARIN (LMW-HEPARIN SANDOZ) IN PATIENTS WITH DEEP VEIN THROMBOSIS
Authors: Vogel G , Machulik M
Published: August 2018

28 patients with deep venous thrombosis (DVT) were randomized to low molecular heparin (LMWH) SANDOZ ( Nurnberg, F.R.G.) 3 OOO U/day per infusio-nem during 10 days or to unfractionated heparin(UFH) 30 000 U/day per infusionem during 10 days. Venography was repeated at day 11. of 14 patients given UFH 5 obtained complete lysis, 6 incomplete lysis and 3 no lysis. The differences were not statistically Significant. No haemorrhagic complications were seen in LMWH grouup but 5 large hamtomas were observed in UFH group. The result suggest that LMWH Sandoz and UFH were equally effecetive on thrombus reduction whereas hemorrgic complications were more common with UFH than with LMWH
PLASMA EXCHANGE WITH A VERY LOW MOLECULAR WEIGHT (VLMW) HEPARIN FRAGMENT CY 222. EVOLUTION OF THE COAGULATION FACTORS
Authors: Crespo B , Assogba U , Bayrou B , Rouffi F , Crespo B , Durande J P , Rottenburg J
Published: August 2018

In a randomized, between-patient study we investigated the effects of a VLMW Heparin fragment (CY 222) versus standard Heparin (SH) in plasma exchanges (n = 8) on coagulation factors (CF): Fibrinogen (FGN), II, V, VII + X, IX, XI, XII, VI11c, VIIIRag.VIIIvwf and ATI11, Prot C, Prot S, Plasminogen.(PGN), APTT, Prothrombin . time (PT), TT, anti factor Xa (AXa), D-Dimer, Platelet Count. Heparin was administered as a bolus and an infusion during the sesr sion, CY 222 as a bolus dose only. 1 to 1.5 plasma volume was exchanged substituted by 5% albumin. The preliminary results (mean, s.d.) at the end of the session (End) and 4 hours later (T4) are summarized below. No differences were found between treatment groups at the baseline (data not shown).

CF are similar in both groups except Factor XII levels at the end of the session, and Factors II, V, XII at T4 (paired t-test: p < 0.05). High APTT levels in all samples seem related to low FGN (p < 0.01). Significant differences in AXa activities were found in each treatment compared with its own standard suggesting different ranges of activities of both drugs. Evolution in D-Dimer levels seems different during the session and at T4 according to the tested drugs and could be related to their mode of administration. Clinical efficiency and tolerance were excellent either with CY 222 or with SH.
A DOUBLE BLIND RANDOMIZED TRIAL OF ORG 10172 LOW MOLECULAR WEIGHT HEPARINOID IN THE PREVENTION OF DEEP VEIN THROMBOSIS IN PATIENTS WITH THROMBOTIC STROKE
Authors: Turple A G G , Levin M N , Hirish J , Carter C J , Jay R M , Powers P J , Andrew M , Magnani H N , Hull R D , Gent M
Published: August 2018

The optimal method of venous thrombosis prophylaxis in patients with stroke is uncertain. ORG 10172 is a low molecular weight heparinoid consisting principally of heparan and dermatan sulphates. In animal studies, ORG 10172 is as effective as unfractionated heparin in preventing venous thrombosis but produces less bleeding. There have been a limited number of descriptive studies on its use in humans, but to date randomized efficacy trials of ORG 10172 in the prevention of venous thrombosis have not been reported. A double blind randomized trial was carried out to compare ORG 10172 with placebo in the prevention of deep vein thrombosis in patients with thrombotic stroke. Seventy-five patients were randomized to receive ORG 10172 (50 patients) in a loading dose of 1,000 anti-Xa units intravenously followed by 750 anti-Xa units subcutaneously 12 hourly or placebo (25 patients). Prophylaxis was commenced within 7 days of stroke onset, continued for 14 days or until discharge from hospital, if earlier. Venous thrombosis surveillance was carried out with 125-1 fibrinogen leg scanning and impedance plethysmography. Venous thrombosis was confirmed by venography which occurred in 2 of 50 (4%) in the ORG 10172 group and 7 of 25 (28%) in the placebo group (p=0.005). The corresponding rates for proximal vein thrombosis were 0% and 16%, respectively (p=0.01). There was one major haemorrhage in the treated group and one minor haemorrhage in the placebo group. The anti-factor Xa levels (units/ml; mean ± SE) gradually rose from 0.18 ± 0.001 and 0.06 ± 0.01 six and 12 hours after injection on the first day to 0.24 ± 0.02 and 0.12 ± 0.01 after 11 days treatment. The results of this study indicate that ORG 10172 heparinoid is effective prophylaxis against deep vein thrombosis in patients with acute thrombotic stroke.
DERMATAN SULPHATE PREVENTS VENOUS THROMBOSIS IN RATS WITHOUT INCREASING BLEEDING
Authors: Maggi A , Abbadini M , Salemi R , Pangrazzi J , Pagella P G , Donati M B
Published: August 2018

Studies have suggested that endogenous proteoglycans, structurally similar to heparin, contribute to the “non-thrombogenicity” of the vascular wall. Our studies concentrated on Dermatan Sulphate (DS-MF 701) Heparin Sulphate (HS) and Standard Heparin (SH), all from Mediolanum Farmaceutici. The antithrombotic potential of these GAGs was evaluated in a well established model of venous thrombosis consisting in the ligature of the vena cava, 15 min after i.v. administration of the drug. A dose response-curve was obtained for DS (0.25-4 mg/kg), HS (1 -5 mg/kg) and SH (0.5-2 mg/kg).

At the dose of 2 mg/kg DS was able to inhibit thrombotic occurrence by 80%, as compared to heparin 100% and HS 40%. The haemorrhaglc potential of these GAGs was then evaluated using two in vivo tests, the “template” method, which detects any alterations in primary haemostasis, and the tail “transection” method which is not only sensitive to changes in primary haemostasis but also to coagulation and fibrinolytic abnormalities; the tests were carried out 15 min after drug administration. Unlike SH, DS did not induce bleeding at equipotent antithrombotic doses wheareas HS induced bleeding even if not to the same extent as SH. DS did not modify the coagulation tests (APTT, TT and anti-Xa), again unlike SH. In conclusion, DS in the venous thrombosis model used, showed the peculiarity to prevent thrombus formation without inducing bleeding complications.
THE PHARMACOKINETICS OF 125I PENTOSAN POLYSULFATE IN THE RABBIT : EVIDENCE FOR A SATURABLE MECHANISM OF CLEARANCE
Authors: Caranobe C , Houin G , Picard C , Pereillo J M , Boneu B
Published: August 2018

Pentosan polysulfate (PPS) is an antithrombotic sulfated polysaccharide with a MW distribution comparable to commercial low molecular weight heparins (LMWH). Unlike LMWH, but as standard heparin, the half life of the drug after bolus injection is dose-dependent. We demonstrate that PPS disappearance results from a saturable mechanism of clearance. Tyrosine was incorporated onto the xylose chains of the molecule and iodinated with the chloramine T method. The labelled derivative had the same MW distribution than the parent compound.and unchanged biological activities. Five ucies of I-PPS were injected with increasing doses (6-12000 ug/kg) of unlabelled PPS to groups of 2-3 animals each. The CPM curves were broken into 3 exponentials (alpha, beta, gamma) ; the beta exponential was close from the curve of.biological activity disappearance, assessed by IIa- 125I-HC II complexes quantitation. The main pharmacokinetic parameters were calculated using conventional methods. The distribution volume (246 ± 81 ml) was independent of the dose delivered whIIe there were significant correlations between the dose, the clearance (r=0.91) and the half life (r = 0.81). Thus in spite of its LMWH, PPS pharmacokinetics mimics that of SH ; this may be the consequence of the over sulfation of the molecule, and of its strong affinity to endothelial cells.
THE MODE OF ACTION OF PENTOSAN POLYSULPHATE IN PLASMA
Authors: Béguin S , Hemker H C
Published: August 2018

We developed a method which enables as to compute the course of prothrombinase activity in clotting plasma (H.C. Hemker, G.M. Willems, S. Béguin: Thromb. Haemostas. 56, 9-17, 1986) and used this for a study of the effect of pentosan polysulphate (PPS) on thrombin generation.

When added to normal plasma in the concentration range of 0-8 μg/ml PPS induces a linear increase of the pseudo first order decay constant of endogenous thrombin like heparin does, 1 ug of PPS being equivalent to 0.045 Aig of heparin. Contrary to heparin this action is partly (∼ 65%) dependent upon AT III and partly (∼ 35%) upon heparin cofactor II.

In normal plasma PPS causes an inhibition of both extrinsic and intrinsic prothrombinase formation. Only in the intrinsic system an increase of the lag time of prothrombinase appearance is observed. Unlike heparin, PPS does not inhibit factor IXa induced thrombin formation neither does it inhibit prothrombinase formation in the presence of preactivated factor VIII. The prolongation of the lag times must therefore be ascribed to inhibition by PPS of the activation of factor VIII.

The inhibition of extrinsic prothrombinase formation by PPS increases with progressive dilution of thromboplastin and is not seen in haemophilia A or B plasma. This demonstrates the existance of a factor VIII and IX dependent process in extrinsic coagulation that gains in importance when the potency of factgr VII-tissue factor complex decreases, i.e. the Josso pathway.

PPS, but also heparin causes an unexplained increase of prothrombinase action in haemophIIic plasma. The same phenomenon may be expected to exist in normal plasma, be it obscured by a concomitant inhibition. This, together with the incomplete inhibition of factor VIII activation by PPS makes that we cannot use this inhibitor as a means to quantitate the Josso pathway. The best estimate that we can obtain is that, in the presence of 2% thromboplastin, the factor IX dependent activation of factor X contributes more then 20% to prothrombinase generation.
INFLUENCE OF THE SULFATION PATTERN ON CERTAIN BIOLOGICAL PROPERTIES OF GALACTOSAMINOGLYCANS
Authors: Casu B , Marchese L , Naggi A , Torri G , Fareed J , Racanelli A , Walenga J M
Published: August 2018

In order to investigate the influence of charge distribution and chain length on the biological properties of sulfated polysaccharides, additional sulfate groups were introduced into the galactosaminoglycans, chondriotin sulfate and dermatan sulfate. Using a flexible method (with sulfuric acid and chlorosulfonic acid) for concurrent sulfation and controlled depolymerization, numerous products were obtained and characterized by chemical, enzymatic and nuclear magnetic resonance spectroscopic methods. The biologic actions of these products were profiled in both in vitro and in vivo assays for antithrombotic activity. Despite a weaker in vitro anticoagulant activity, low molecular weight over sulfated galactosaminoglycans produced significant dose-dependent antithrombotic actions in animal models which were similar to the actions observed with oversulfated low molecular weight heparins. These results suggest that a significant antithrombotic activity can be elicited through non-specific interactions of polysulfates with cellular and plasma components, and that clusters of sulfate groups such as the 4-6 disulfate group on D-galactosaminoglycan residues may be important for these interactions. Furthermore, these results, also suggest that supersulfation of glycosaminogly-cans results in products with biologic activity distinct from the native material.
PROTHROMBIN TIMES AND CLOTTABLE FIBRINOGEN DETERMINATIONS ON AN AUTOMATED COAGULATION LABORATORY (ACL 810) ANALYZER.
Authors: Farag A M , Brown J M , Sachs R J , Mammen E F
Published: August 2018

The ACL 810 is a fully automated, microcomputer controlled centrifugal analyzer (Instrumentation Laboratory/IL). It performs clot based and chromogenic substrate assays. The clot based assay utilizes laser nephelometry. Prothrombin times (PT) in normal and abnormal plasmas were analyzed on the ACL using different thromboplastins and the data compared with a Fibrometer and a Coag-A-Mate. The correlation between the ACL (II reagents) and the Coag-A-Mate and Fibrometer (Thromboplastin C, Dade) for 100 samples were as follows: ACL vs Coag-A-Mate r = 0.87 (p < 0.001), m (slope) 1.13; ACL vs Fibrometer r = 0.92 (p < 0.001), m = 1.1.

Since the ACL 810 determines PT and fibrinogen simultaneously, we compared fibrinogen levels on the ACL 810 (II Thromboplastin) with those analyzed on a Multistat III Centrifugal Analyzer (MCA) (bovine thrombin, Hemostasis Laboratories) and a Fibrometer (Data-Fi, Dade). The following correlations were obtained: ACL vs Fibrometer n = 50, r = 0.922 (p < 0.001), m 0.72; ACL vs MCA n = 100, r 0.899 (p < 0.001), m 0.7; MCA vs Fibrometer n = 70, r = 0.954 (p < 0.001), m = 0.936. We also compared different manufacturers' thromboplastins on the ACL 810 in the determination of fibrinogen with the following results:

The run-to-run coefficient of variation (CV) for fibrinogen was 4.4% and for PT 0.26%; the intra-run CV was 4.66% and 0.41%. These data reveal excellent correlations for both PT and fibrinogen not only with different instruments but also with different thromboplastins on the same instrument.
THE RELIABILITY OF THE INTERNATIONAL NORMALISED RATIO (INR) DURING SHORT TERM ORAL ANTICOAGULANT TREATMENT
Authors: McKernan A , Thomson J M , Poller L
Published: August 2018

A prospective study has been performed to assess INR values obtained with a variety of thromboplastins during the early days of coumarin treatment. The reagents were BCT/253 (the primary International Reference Preparation), Diagen Activated, Diagen Freeze Dried, Manchester Reagent and Dade Thromboplastin FS. Prothrombin times were performed before the start of treatment and at regular intervals on fifteen patients who were given a slow induction regime. In theory INR should be the same irrespective of the thromboplastin. A wide range of values was however observed with the different thromboplastins on the same plasma samples. The mean deviations of the individual reagents from the mean INR obtained with the primary IRP were: Diagen Freeze-Dried 26%, Diagen Activated 13%, Dade FS 17%, Manchester Reagent 3%.

There are two possible explanations for the discrepant findings. 1) In the induction phase Vitamin K dependent clotting factors are depressed at varying rates and thromboplastins differ in their sensitivity to the depression of these factors. The International Sensitivity Indices from which INR are derived are based on results from long-term stabilised patients. 2) The manufacturers' calibrations may be incorrect as demonstrated by the consistent differences of the results with some reagents from the IRP. The findings indicate therefore that INR values may not be dependable in the early days of oral anticoagulation with some thromboplastin reagents and that manufacturers' calibrations require independent assessment preferably by national control laboratories.
APPLICATIONS OF INTERNATIONAL RECOMMENDATIONS ON THE STANDARDIZATION OF PROTHROMBIN TIME IN ORAL ANTICOAGULANT CONTROL
Authors: Dati F , Becker U , Heimburger N
Published: August 2018

The determination of prothrombin time (PT) in oral anticoagulant control is affected by a broad variation. The responsible factors are: type of thromboplastin incorporated in the PT reagents, procedure for use, clotting factors or heparin inhibitors added to the reagent, method of expression of PT results. Recently, joint recommendations have been issued by International Committees (ICSH/ICTH) taking into account the system of International Thromboplastins and the statistical model for thromboplastin calibration established by WHO. The aim is a standardization of commercial thromboplastins for PT tests in order to allow the use of the international scale of oral anticoagulant intensity (INR: Intern. Normalized Ratio). Following such recommendations we have standardized two new PT tests, based on coagulometric and photometric methods which rely on the same sensitive human placental thromboplastin. The coagulometric PT test (Thromborel®S) is performed with conventional coagulometers. The photometric PT assay (Chromoquick®) uses a new chromogenic substrate specific for thrombin. This method is based on the measurement of the time necessary to reach a fixed increase of absorbance (0.1 A) using a special microprocessor-controlled photometer.

The two PT reagents were calibrated either directly against a reference preparation (BCT) or via an intermediate standard thromboplastin in two multicentric studies. The calibration procedure by the WHO method allows to assign the corresponding ISI (Intern. Sensitivity Index) to the PT reagent used and the transformation of the obtained prothrombin ratio (PR) into INR by the equation INR = PRISI. The calculated ISI values were 1.08 for the coagulometric PT reagent (n = 330) and 1.07 for the photometric reagent (n = 365), respectively.

The reproducibility of the ISI value for the new human placental thromboplastin for 64 different batches amounts to 3.6 %, the mean ISI value being 1.12.

Comparison with the reference thromboplastins in PR values gave a good correlation.

A) Coagul. PT assay (x): r = 0.964; y = 1.03x ™ 0.1;

B) Photom. PT assay (x): r = 0.940; y = 1.02x ™ 0.1.
ANTICOAGULATION IN PREGNANCY IN PATIENTS WITH CARDIAC VALVE PROSTHESIS
Authors: Meschengieser S S , Woods A I , Lazzari M Z
Published: August 2018

The use of oral anticoagulation in pregnancy is controversial due to the risk of embryopathy (1° trimester), intracerebral hemorrhages (2° trimester) and the use of heparin is associated with a high morbidity due to abortion and prematurity. The outcome of 50 pregnancies in 30 patients with heart prosthesis was evaluated. Acenocoumarol and aspirin 500 mg/daily were given in all the patients since their surgery. Anticoagulant doses were controlled with prothrombin time (PT) performed with human brain thromboplastin and APTT. The therapeutic range for the PT was the International Calibrated Ratio (ICR) 2.5 to 3.5. In 12 pregnancies, oral anticoagulation was replaced by subcutaneous heparin in the first trimester; the same policy was followed before delivery except in 4 cases. The foetal loss was 34% with equal distribution along the three trimesters and no correlation with excess of anticoagulation. The incidence of hemorrhage was 6% and the rate of cerebral embolism was also 6% (3 on to 50). Two of the three episodes of embolism appeared while patients were on heparin. A total of 33 normal babies were born (66%). No typical warfarin embryopathy was found and no perinatal mortality was observed. A slight reduction in the anticoagulant doses was necessary in almost half of the cases.

As the rate of foetal loss with heparin is not better than with oral anticoagulants and the incidence of embolism is higher, we are doubtful about the indication of subcutaneous heparin in the first trimester considering our absence of malformations with the acenocoumarin.
EVIDENCE FOR BIIIARY EXCRETION OF PHENPROCOUMON AND ITS METABOLITES IN HUMANS; IDENTIFICATION BY HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY AND GAS CHROMATOGRAPHY-MASS SPECTROMETRY
Authors: de Vries J X , Raedsch R , Stiehl A , Voelker U , Walter-Sack I , Weber E
Published: August 2018

Recently it has been shown that in man the oral couma-rin anticoagulant phenprocoumon is eliminated up to 60-70 % in urine and 30-40 % in faeces; in urine phenprocoumon (PH) and its metabolites 7-hydroxy-(7-OH),6-hydroxy-(6-OH) and 4'-hydroxy-(4'-OH) phenprocoumon are present mainly as conjugates. No data so far were available on the biliary excretion of these compounds.

We examined bile obtained from four in-patients during PH treatment; bile samples were aspirated in the duodenum at the papilla during routine diagnostic endoscopy and immediately deep frozen before analysis. Samples were extracted both untreated as well as after hydrolysis with 6-glucuronidase/aryl sulfatase and separated by reversed phase gradient elution high performance liquid chromatography (HPLC) with fluorescence detection; for confirmation, the same extracts were methylated and analysed by gas chromatography-mass spectrometry (CG-MS) (J.X.de Vries et al J Chromatogr., 338 (1985) 325). PH, 7-OH, 6-OH and 4'-OH were identified by comparison with synthetic authentic samples'''''''
KINETICS OF HYDROLYSIS OF THE CHROMOGENIC SUBSTRATE S2238 BY ALPHA-THROMBIN : INFLUENCE OF THE pH AND THE IONIC STRENGTH
Authors: Verhamme I M A , van Dedem G W K
Published: August 2018

The knowledge of the pH and ionic strength dependence of kcat and Km for the hydrolysis of S2238 (HD-phe-pip-arg-pNA.2HC1) by alpha-thrombin is essential in determining optimal reaction conditions of residual enzyme in systems where also protease inhibitors and glycosaminoglycan catalysts play a role.We studied the kinetic behavior of S2238 in piperazine/glycylglycine/NaOH buffers from pH 6 to 11 and with a calculated ionic strength up to 0.7 M taking into account the pH-dependent concentration of the buffer species. The kinetic parameters of 60 Michaelis-Menten substrate functions were used for the setup of ionic strength and pH profiles. The kcat values are dependent upon the ionic strength, increasing steeply up to about 0.3 M and decreasing again at high ionic strength.The Km however,reflecting the affinity between enzyme and substrate,is nearly unaffected.The Km values at very alkaline pH are markedly elevated,indicating a conformational form which does not readily bind substrate.The pH profiles for kcat and kcat/Km are displaced towards the low pH side with increasing ionic strength.The ascending limb corresponds to the pK of the Asp-His charge relay system,decreasing with increasing ionic strength from 7.2 to 6.6 in the ES complex and from 6.8 to 6.6 in the free enzyme.Apparently substrate binding provokes a pK increase of the active His residue.The descending limb in the kcat profile could be described by a hypothetical pK varying from 11.5 to 10.7 but the activity decrease is probably due to enzyme inactivation.The alkaline limb of the kcat/Km profile is governed by a pK of 9.4 which is rather independent of the ionic strength and could be attributed to the B-chain terminal isoleucine ,forming a salt bridge with Asp 194 and stabilizing the active site conformation as proven for other serine proteases.Data analysis via a modified Debije function with appropriate estimates for the dielectric constant and the radius of the macro-ion can provide information on the charge density of the enzyme.
A NEW DISORDER CHARACTERIZED BY FACTOR V DEFICIENCY AND MOLECULAR ABNORMALITY OF VON willEBRAND FACTOR ANTIGEN
Authors: Akutsu Y , Mori K , Suzuki S , Ishikawa M , Sakai H , Hiwatashi K , Fujimoto H , Endo E , Yasuda H
Published: August 2018

Two patients with a new type of combined factor V and factor Vm deficiency is described. The propositus was 41 year-old woman who had never experienced any bleeding episode. We considered that this case was to be one of the combined factor V/Vin deficiency from the results of prolonged prothrombin time, 23.5 sec. (control 12.6 sec.) and kaolin partial thromboplastin time, 58.9 sec. (control 39.2 sec.) before surgery because of the right postoperative maxillary cyst. Coagulation studies at the time showed that the mean F. V and F. Vm levels were 11.5 % and 27 %, respectively. Von willebrand factor (vWF) which is the biological activity of F. VIII related platelet adhesion and vWF:Ag were also decreased (vWF:Ag 34 %, vwF 36.5 %). Bleeding time, platelet aggregation induced by ADP, collagen and ristocetin were normal. Another coagulation factors were normal and pathological inhibitors were not detected. The antigen levels of protein C (PC) and protein S (PS) were found to be normal. Her younger brother, who had no*past history of spontaneous bleeding, was found to be same abnormality (F. V 13 %, F. VIE 38 %, vwF:Ag 64 %, vWF 58 %). Bleeding time, platelet aggregation, another coagulation factors, PC and PS were normal. Furthermore, decreased intermediate multimers of vWF:Ag in these cases were observed on two-dimensional crossed immunoelectrophoresis.

Since first case described by Oeri et al in 1954, approximately 40 combined factor V/Vm deficiency have been reported. In most such cases, vWF:Ag, vwF levels, and the electrophoretic mobility were within the normal range. In our cases, vwF:Ag and vwF were apparently decreased, and appeared to be the same abnormality as the von willebrand disease type-I. From the above mentioned findings, these patients were thought to be a new disorder of combined F. V deficiency and molecular abnormality of F. VIE ( vWF:Ag).
DETERMINATION OF ACTIVATED FACTOR VII BY BOVINE AND RABBIT THROMBOPLASTINS.
Authors: Kitchen S , Malia R G , Greaves M , Preston F E
Published: August 2018

Thromboplastins from different animal species vary in their relative sensitivity to non-activated VII and activated VII (Vila) in plasma and this has formed the basis of Vila determinations using bovine and human thromboplastins. Since the use of human thromboplastin is now being discontinued we have re-evaluated the assessment of Vila using bovine and rabbit thromboplastins (BT and RT respectively). Citrated plasma was prepared from four normal individuals and maintained at 4°C in glass containers for 24 hours to promote the formation of Vila. Each plasma was subsampled at 0, 2, 4, 6 and 24 hours and factor VII assays performed using BT and two different rabbit preparations viz, (a) Manchester Reagent (RTmr, ISI = 1.16) and (b) Diagen phenolised (RT diagenlSI = 1.4). Results were expressed as the ratio BT/RT. With all thromboplastins factor VII activity increased with time. The changes were most marked using BT with much smaller increments occurring using the rabbit preparatrions. RTmr was less sensitive to Vila than RT diagen. The ratio BT/RTmr thus provides an index of Vila.

Using this method we determined Vila in plasma samples from 29 normal subjects. Results obtained were x = 1 ,00;s.d. = 0.126; normal range 0.72 - 1.25 The method was applied to 29 plasma samples from 19 patients with disseminated intravascular coagulation giving BT/RTmr = 1.71 (range 0.92 - 3.38). We conclude that the ratio of Factor VII measured by bovine and rabbit thromboplastins respectively provides an index of Factor VII activation in plasma.
TISSUE FACTOR-FACTOR VII INHIBITION REQUIRES FACTOR Xa AND PLASMA LIPOPROTEINS.
Authors: Hubbard A R , Jennings C A
Published: August 2018

Tissue factor rapidly loses procoagulant activity when incubated with defibrmated normal plasma and calcium ions. Inhibition is apparently directed against the tissue factor-Factor VII complex (TF-EVII) and requires Factor Xa and a component(s) found in A1 (OH)3-adsorbed plasma (AP). We have developed a two stage assay for the inhibitor which involves first, the incubation of a TF-FVII complex with test material in the presence of Factor Xa, followed by the amidolytic assay of residual TF-EVII activity.

Our studies have indicated that the component of AP responsible for this effect is lipoprotein. Incubation of AP with antiserum to apo-lipoprotein B (apo B) reduced the inhibitory activity by 73%, whereas antisera to antithrombin III and a2-macroglobulin had no effect. Inhibition by AP does not appear to be caused by an artefact of adsorption, since the inhibitory-capacity of AP was 59% of normal, defibrinated plasma. This correlated well with the apo B antigen in AP, which was 64% of normal. Moreover, the dose/response lines of AP and normal plasma were parallel, suggesting that the inhibitor assay is not affected by the presence of normal levels of coagulation factors.

Purified lipoprotein-rich fractions prepared from AP using density gradient ultracentrifugation all contained inhibitory activity. Incubation of these fractions with anti-apo B greatly reduced the inhibition by the very low density and low density lipoprotein-rich fractions (VLDL and LDL) but had essentially no effect on the high density lipoprotein-rich fractions (HDL). Incubation of LDL with Factor Xa produced an inhibitory component which eluted together with the apo B antigen during gel filtration. Inhibition appears to require the interaction of Factor Xa with plasma lipoproteins, particularly LDL. The product of this interaction is then able to bind and inhibit the TF EVII complex. The requirement of Factor Xa in order for inhibition to be expressed is indicative of a feedback anticoagulant response which may have physiological significance.
RAT HEPAT0CYTES IN CULTURE INACTIVATE FACTOR Xa.
Authors: Qureshi G D , Sun M , Gervin C , Evans H
Published: August 2018

Plasma contains zymogens of clotting factors, which under various stimuli are activated to serine proteases. Whereas much knowledge has been gained about the activation of clotting factors, relatively little is known about inactivation of these proteases. Antithrombin III has been shown to inactivate some activated clotting factors in plasma. Studies in intact animals have suggested that activated clotting factors are mainly inactivated in the liver. To investigate more fully the role of liver in inactivating the activated factors, we studied the stability of activated factor X(Xa) in hepatocyte cultures. Monolayer cultures on non-proliferating rat hepatocytes were prepared according to the method of Bissell et al. The culture medium was chemically defined and was free from serum or serum products. After the 24 h stabilization period, 0.5 units/ml of 100% activated bovine factor Xa was co-cultured with hepatocytes for 8 h. Samples were collected at 0, ½, 1 2, 4 and 8 h and tested for Xa activity using chromogenic substrate S-2222. At the end of 8 h only 41.07% of the initial Xa activity remained. Xa inactivation was not affected by a commercially prepared unfractionated heparin (1 unit/ml) and estradiol at 12.5, 25, 125 nM, a potentiator and inhibitor of antithrombin III, respectively. Inactivation of Xa in hepatocyte cultures was inhibited by the addition of cycloheximide (10-4M). Our data suggests that factor Xa is inactivated in hepatocyte cultures by one or more hepatic derived factors which do not meet the functional characteristics of antithrombin III.