bluuebunny/crossref_metadata_2025 huggingface dataset:
Factors Involved In the Plasminogen Activation System in Human Breast Tumours
Authors: Damjanovich László , Turzó Csaba , Ádány Róza
Published: July 2018

The plasminogen activation system is a delicately balanced assembly of enzymes which seems to have primary influence on tumour progression. The conversion of plasminogen into serine protease plasmin with fibrinolytic activity depends on the actual balance between plasminogen activators (urokinase type; u-PA and tissue type; t-PA) and their inhibitors (type 1 and 2 plasminogen activator inhibitors; PAI-1 and PAI-2). The purpose of this study was to determine the exact histological localization of all the major factors involved in plasminogen activation, and activation inhibition (plasmin system) in benign and malignant breast tumour samples. Our results show that factors of the plasmin system are present both in benign and malignant tumours. Cancer cells strongly labelled for both u-PA and t-PA, but epithelial cells of fibroadenoma samples were also stained for plasminogen activators at least as intensively as tumour cells in cancerous tissues. In fibroadenomas, all the epithelial cells were labelled for PAM. Staining became sporadic in malignant tumours, cells located at the periphery of tumour cell clusters regularly did not show reaction for PAI-1. In the benign tumour samples the perialveolar connective tissue stroma contained a lot of PAI-1 positive cells, showing characteristics of fibroblasts; but their number was strongly decreased in the stroma of malignant tumours. These findings indicate that the higher level of u-PA antigen, detected in malignant breast tumour samples by biochemical techniques, does not necessarily indicate increased u-PA production by tumour cells but it might be owing to the increased number of cells producing u-PA as well. In malignant tumours PAI-1 seems to be decreased in the frontage of malignant cell invasion; i.e. malignant cells at the host/tumour interface do not express PAI-1 in morphologically detectable quantity and in the peritumoural connective tissue the number of fibroblasts containing PAI-1 is also decreased.
Thrombelastography during and after Elective Abdominal Surgery
Authors: Butler J M
Published: July 2018

Thrombelastography has been performed on recalcified whole blood from 50 patients before, during and after elective abdominal surgery. The characteristic changes of the thrombelastographic indices r, k and mA are described.

During operation r and k shortened, but no change in mA was observed. This response was in part associated with an increase in factor VIII activity. Following operation, while r time was somewhat shortened, much more marked changes in k and mA were evident. Increasing fibrinogen concentration was the dominant factor in determining the post-operative changes in the thrombelastograph.
Measurement of Thrombin Generation in Whole Blood - The Effect of Heparin and Aspirin
Authors: Kessels Han , Béguin Suzette , Andree Harry , Coenraad Hemker H
Published: July 2018

A technique has been developed to monitor the development of thrombin in freshly collected whole blood in the absence of anticoagulants. It is based on the centrifugal separation of the cellular components from subsamples of blood drawn from non-anticoagulated clotting whole blood which are diluted in buffer containing a chromogenic substrate.

It is shown that the burst of thrombin generation after triggering coagulation with trace amounts of tissue thromboplastin occurs sooner in non-anticoagulated whole blood than in citrated whole blood. Heparin is shown to prolong the lag-time of thrombin generation more in native blood than in recalcified citrated blood.

It is also demonstrated that intake of 500 mg of aspirin significantly delays and inhibits thrombin generation in non-anticoagulated, thromboplastin triggered whole blood, whereas it has no effect on the coagulation in citrated plasma. The effect of aspirin intake on thrombin generation in blood is roughly equal to that of 0.03 U/ml of unfractionated heparin. This demonstrates that platelet reactions and the coagulation system are closely linked processes. It further lends support to the hypothesis that inhibition of thrombin generation is a common denominator of antithrombotic therapy.
Pulmonary Embolism in Patients with Upper Extremity DVT Associated to Venous Central Lines - A Prospective Study
Authors: Monreal Manuel , Raventos Antoni , Lerma Rosa , Ruiz Joan , Lafoz Elena , Alastrue Antoni , Llamazares Jaume F
Published: July 2018

We performed a prospective study in 86 consecutive patients with central vein catheter-related deep venous thrombosis (DVT) of the upper extremity, to evaluate the prevalence of pulmonary embolism (PE), and to identify clinical variables that would increase the likelihood of developing PE in an individual patient.

Since upper-extremity DVT was established, all patients received intravenous heparin therapy. Then, a ventilation-perfusion lung scan was obtained within 24 h of DVT diagnosis, whether respiratory symptoms were present or not. Six points of clinical information were recorded on entering in the study, and then compared with the scintigraphic findings: age, sex, the underlying disease, the catheter material, the character of the infusate, and the duration of cannulation.

Thirteen patients were considered to have PE. Sixty-six patients were finally classified as having a normal lung scan, and 7 patients were excluded from the study (because of indeterminate lung scan 6; because of femoropopliteal thrombosis simultaneously present 1). Two out of the 13 patients with PE subsequently died because of recurrent, massive embolism, despite adequate heparin therapy. PE was more commonly present in patients with polyvinyle chloride or polyethylene catheters (10/38,26%) as compared to patients with either polyurethane or siliconized catheters (3/41, 7%; p <0.05, Chi-Square test; Odds Ratio = 4.52,95% Cl 1.01-23.07).

We conclude that PE is not a rare event in these patients, and it may be life-threatening even despite adequate heparin therapy. Since the more recently available soft catheters seem to carry a lower risk of developing PE, there seems to be no reason to continue to use polyvinyl chloride or polyethylene catheters.
Effect of Oestrogen Dose on Whole Blood Platelet Activation in Women Taking New Low Dose Oral Contraceptives
Authors: Norris Lucy A , Bonnar John
Published: July 2018

Oral contraceptive use is known to cause changes in the haemostatic system. These changes are thought to be related to oestrogen dose and to provide a possible link between the increased risk of thromboembolic disease known to occur in women taking oestrogen containing oral contraceptives. This study measured whole blood platelet activation, serially, in women taking oral contraceptives containing 20 μg and 30 μg ethinyloestradiol combined with desogestrel. Increased levels of ADP and arachidonic acid induced aggregation were observed in women taking the 30 μg ethinyloestradiol combination. Platelet release of β-thromboglobulin (βTG) was also significantly increased. Increased collagen induced aggregation was observed but this failed to reach statistical significance for the individual treatment groups. In women taking the 20 μg ethinyloestradiol combination, a significant increase was only observed when platelets were stimulated with arachidonic acid. Platelet factor 4 (PF4) levels were unchanged in both groups. Significantly higher levels of βTG were observed in women taking the 30 μg ethinyloestradiol combination compared with women taking the 20 μg ethinyloestradiol combination. These results show that oral contraceptive use is associated with platelet activation. Women taking the 20 μg ethinyloestradiol combination show less changes in platelet activation than women taking the 30 μg ethinyloestradiol combination. This lower dose pill may therefore be particularly suitable for high risk women wishing to use oral contraception.
Association of Antibodies to Heat-Shock Protein-65 with Percutaneous Transluminal Coronary Angioplasty and Subsequent Restenosis
Authors: Mukherjee Manjari , De Benedictis Christine , Jewitt David , Kakkar Vijay V
Published: July 2018

Heat-shock protein (HSP)-65 of mycobacterial origin has been implicated in the mediation of atherosclerosis by immune mechanisms. Any role of HSP-65 in mediating restenosis is however not clear. We determined the anti-HSP-65 antibodies in 28 patients, 25 male and 3 female, aged 35 to 78 years, with coronary artery disease (CAD) and undergoing percutaneous transluminal coronary angioplasty (PTCA). Of the 28 patients, 12 suffered restenosis five to seven months later.

The serum levels of antibody were measured at baseline, immediately after PTCA, before discharge from the hospital, and at 6 weeks, 3 and 6 months after the performance of PTCA. The antibody levels were expressed in OD U/log titre, as explained in the text, and termed estimated OD or EOD. The control group consisted of 29 healthy volunteers, 16 male and 13 female, aged 24 to 59 years. The mean Eod of the patients at baseline was higher than that of the controls (0.60 ± 0.12, SD; 95% Cl 0.56-0.64 compared with 0.53 ± 0.13; 0.48-0.58; p < 0.01). There was no correlation between age and EOD of patients, controls or both taken together, ruling out the influence of age on the EOD changes in the given age range. The mean antibody levels of the patients with CAD were similar whether or not they suffered subsequent restenosis (0.61 ± 0.15; 0.53-0.69 in the patency group, and 0.60 ± 0.10; 0.54-0.66 in the restenosis group). However, the patients who did not develop restenosis had a drop in their antibody levels immediately after PTCA (0.51 ± 0.14; 0.48-0.55; 2-tailed p = 0.029), and at discharge from the hospital (0.52 ± 0.15; 0.44-0.60; p = 0.036) as compared with the baseline. This decrease of antibodies was not observed in the restenosis group. Furthermore, the anti-HSP-65 antibody levels remained slightly low throughout the 6-month follow-up in the patients with patent coronaries as compared with patients who reste-nosed, but the decrease was statistically not significant at 5% level at any stage. Besides the anti-HSP-65 antibodies, the levels of anticardio-lipin (ACL) antibody were also measured in all the patients. The levels of the ACL antibody were found to be within the normal range before and at all stages after PTCA, ruling out the PTCA-associated change in the anti-HSP-65 antibody as a non-specific occurrence. Thus, a drop in the level of antibody against HSP-65 after PTCA seemed to be associated with a favourable outcome, and may serve as a useful prognostic marker of coronary angioplasty
The Production of Heparin Cofactor li Is Not Regulated by Inflammatory Cytokines in Human Hepatoma Cells: Comparison with Plasminogen Activator Inhibitor Type-1
Authors: Kolke Chisato , Hayakawa Yumiko , Niiya Kenji , Sakuragawa Nobuo , Sasaki Hideo
Published: July 2018

Using the Northern blot technique, we screened 6 human hepatoma cell lines to investigate the regulation mechanism of heparin cofactor II (HCII) biosynthesis. We found that HuH-7 and Hep G2 cells constitutively expressed the HC II gene. In conditioned medium, HuH-7 cells constantly produced HC II that was functionally active and formed a complex with thrombin in the presence of dermatan sulfate. HC II is thought be an acute phase reactant, and, therefore, we examined the effects of the major inflammatory cytokines, IL-6, IL-1β, and TNF-α, on the regulation of HC II production in HuH-7 and Hep G2 cells. In HuH-7 cells, the antigen and mRNA levels of plasminogen activator inhibitor type-1 (PAI-1), an acute phase protein produced by hepa-tocytes, were increased in response to stimulation with either IL-6 or IL-1 (3 or both, but HC II antigen and mRNA levels were not changed by the same stimulation. Even when Hep G2 cells were treated with a combination of three cytokines, IL-6, IL-1β, and TNF-α, HC II antigen and mRNA levels were not changed; however, PAI-1 antigen and mRNA levels were clearly increased. These results suggest that the production of HC II in hepatoma cells is not regulated by the major inflammatory mediators, IL-6, IL-iβ, and TNF-α.
The Risk of Abortion and Stillbirth in Antithrombin-, Protein C-, and Protein S-deficient Women
Authors: Jan Sanson Bernd , Friedrich Philip W , Simioni Paolo , Zanardi Sandra , Huisman Menno V , Girolami Antonio , Wouter ten Cate Jan , Prins Martin H
Published: July 2018

Hereditary deficiencies of the naturally occurring anticoagulants are well recognized conditions predisposing to recurrent venous thromboembolism. Since thrombotic phenomena have been implied as a cause of abortion and stillbirth, we hypothesized that these deficiencies increase the risk of fetal demise.

A group of 129 female subjects who had been pregnant at least once and who had a family member with documented venous thrombosis associated with a deficiency of AT, PC or PS were studied. We first assessed the obstetric history and subsequently determined the deficiency status. In the 60 deficient subjects 42 (22.3%) of the 188 pregnancies resulted in miscarriage or stillbirth as compared to 23 (11.4%) of the 202 pregnancies in the 69 non-deficient subjects. The relative risk of abortion and stillbirth per pregnancy for deficient women as compared to non-deficient women was 2.0 (95% C.I. 1.2-3.3).
Hemorheological Changes in Elderly Subjects-Effect of Pentosan Polysulfate and Possible Role of Leucocyte Arachidonic Acid Metabolism
Authors: Freyburger G , Larrue F , Manciet G , Lorient-Roudaut M F , Larrue J , Boisseau M R
Published: July 2018

The effects of pentosan polysulfate (PPS) on various hemorheological parameters were studied in a group of very elderly subjects in good general health. Alterations in blood viscosity and filterability were detected in these patients, without any concomitant changes in factors which are known to affect these parameters: notably hematocrit, fibrinogen and plasma lipid levels. The hemorheological abnormalities were considerably improved by twice daily treatment with 50 mg of PPS (i.m.).

Apart from its anticoagulant activity, PPS has been shown to have an anti-inflammatory action. We were interested to investigate its effects on metabolism of exogenous arachidonic acid (AA) by both platelets and leucocytes. It is becoming increasingly recognized that metabolites of AA via the 5 LO pathway appear to play a role in inflammatory processes. In this study, PPS was found to inhibit leucocyte 5 LO activity. Reduction in the levels of these metabolites may therefore have an effect on whole blood rheology.
The Balance Between Thrombin And Plasmin Action On Fibrinogen And The Occurrence Of Venous Thrombosis
Authors: Owen J , Kvam D , Nossel H L , Kaplan K L
Published: July 2018

We have studied activation of coagulation, fibrinolysis and platelets in association with venous thrombosis in patients undergoing craniotomy. Serial 125I-fibriongen leg scans were used to detect the presence or onset of venous thrombosis and were confirmed by venography when positive. Serial measurements were made of plasma levels of fibrino- peptide A (FPA), thrombin-increasable fibrinopeptide B (TIFPB), platelet factor 4 (PF4) and beta thromboglobulin (βTG) as indices of thrombin or plasmin proteolysis of fibrinogen and of platelet α-granule release respectively. 17 patients had positive leg scans and 14 had negative leg scans. In normal individuals the FPA level is less than the TIFPB level with a ratio of FPA to TIFPB which is less than 0.5. In 16 of 17 patients with thrombosis FPA levels were elevated and the FPA:TIFPB ratio was markedly increased immediately preceding and overlapping the change of leg scan from negative to positive or with the initial detection of a positive leg scan. In 9 patients the time of onset of thrombosis was documented by the change of the leg scan from negative to positive. In 8 of these patients the FPA level rose and exceeded the TIFPB level preceding and/or overlapping the change in the leg scan. Similar changes in FPA and TIFPB were present in the other 8 patients but since the initial leg scan was positive the time of onset of thrombosis could not be documented. Prior1 to thrombosis $TG but not PF4 levels were elevated. The marked changes in the FPA/TIFPB ratio prior to thrombosis exhibited a significant positive predictive value for thrombosis. The data document a close association between venous thrombosis and imbalance between thrombin and plasmin action on fibrinogen as reflected by plasma levels of FPA and TIFPB.
The Time Relation Between The Haemostatic Aggregation Of Platelets And Their Release Reaction In Vivo
Authors: Görög P , Born G V R
Published: July 2018

The release reaction of platelets has been assumed to subserve a positive feedback mechanism responsible for their aggregation in haemostasis and thrombosis. This assumption is based mainly on in vitro experiments. Considerable uncertainty remains about the contribution of the release reaction to the initiation of haemostasis in vivo. The rapidity of the process and the presence of other tissues makes it impossible to follow the reaction quantitatively in vivo by methods which permit this in vitro. We have therefore applied quantitative electron microscopy to find out how quickly the concentration of dense bodies decreases in platelets during their haemostatic aggregation. In mice, platelets were enriched in dense bodies by pretreatment with serotonin.

Mesenteric arteries were incised with a sharp blade. Bleeding was stopped by a micromanipulator-operated device about 15 sec and 60 sec after the cut. The cut segments were immediately fixed in situ with glutaraldehyde and postfixed. Serial sections were made for electron microscopy. Platelets isolated from peripheral blood of the same animal were prepared similarly. Electron micrographs were projected on to a television screen and numbers of dense bodies and total platelet areas were determined by an image analysing computer. After 15 sec there were no significant differences in numbers of dense bodies in platelets from different parts of the haemostatic plugs (8.31±0.57/100 μ2 mean ± s.e.m.) and in platelets from the blood (8.93±0.38). On the other hand, after 60 sec the parts furthest from the cut contained fewer dense bodies than the nearer parts and the overall dense body number (5.86±0.05) was considerably smaller (p<0.001) than that of platelets from the blood (14.45±0.09).

The results suggest that haemostatic aggregation of platelets does not initially depend on their release reaction.
Injury And Inflammation Induced In The Microcirculation Of The Rabbit By Antigen-Antibody Complexes
Authors: Crawford J P , Movat H Z , Ranadive N S , Hay J B
Published: July 2018

An experimental model was used to study the morphogenesis, quantitation and kinetics of the inflammatory response in the dermis of rabbits. New Zealand White rabbits were immunized with ovalbumin, bled, the antibody precipitated by ammonium sulphate and redissolved in saline. Circulating antibody was determined by the quantitative precipitin test. The reversed passive Arthus reaction was elicited by injecting normal rabbits intravenously with antigen [1.0 gm] and intradermally with 0.9 mg of antibody, in triplicate sites. The lesions were allowed to develop for 1-8, 18 and 24 hours post-injection. To assess the inflammatory response, 5 parameters were examined:
1) Exudation of plasma using 125I-serum albumin;
2) Infiltration of leukocytes with 51Cr-labelled cells;
3) Hemorrhage with 59Fe-erythrocytes; 4) Formation of microthrombi with 111In-platelets; and 5) Blood flow with 57Co-microspheres. The animals were killed, the lesions punched out with a cork borer and counted in a gamma spectrometer. Skin lesions from rabbits not injected with tracers were fixed in glutaraldehyde-formalin and prepared for microscopic examination. With the aid of these techniques, increase of vascular permeability and the formation of microthrombi peaked in 2-hour old lesions. Increased blood flow peaked in leasions 3-4 hours postinjection. Infiltration of polymorphonuclear leukocytes reached a maximum in lesions 2-3 hours old, followed by hemorrhage, which reached a maximum and plateaued after 4 hours. Immune complexes may induce a hemorrhagic inflammation, the kinetics of which can be ascertained. Preliminary studies indicate that these techniques are applicable for the examination of experimentally-induced hypersensitivity pneumonitides and disorders of the pulmonary microcirculation.
Suppression Of Exercise-Induced Plasminogen Activator And Plasma Renin Activity Following Aspirin Ingestion
Authors: Osmond D H , Hedlin A M , Loh A Y , Sakac D , Topp R R
Published: July 2018

The influence of aspirin on the fibrinolytic and renin systems was studied in 10 healthy young men, who served as their own controls. Basal blood samples were obtained after a 15-minute rest, immediately after exercise, and following a 15-min post-exercise rest. Later, (7-14 days) 1250 mg aspirin was ingested 1 hr. before test samples were obtained, as before. Exercise on a bicycle ergometer induced doubling of the resting heart rate within 2 min., and was continued for 3 min. longer. Aspirin induced a significant reduction in the level of plasminogen activator, before and after exercise, and also some decrease in hemoglobin and hematocrit levels. Exercise almost doubled the baspl plasma renin activity (PRA, angiotensin I, ng/ml plasma/hr., by radioimmunoassay) in the control and aspirin experiments. Aspirin did not significantly blunt this exercise-induced rise in PRA. However, after 15-minute rest, the control PRA remained elevated, while the postaspirin PRA was 37% below it (p < 0.025). After cold activation of the plasmas (-4°C, 72 hr.), the PRA increment over corresponding non-activated samples was used to estimate “prorenin”. Prorenin as % of active renin before aspirin was 347.2, 210.5, and 210.8, for the basal and 2 post-exercise periods respectively; after aspirin the values were 460.5, 354.9, and 353.4. Clearly, exercise depresses the proportion of prorenin, suggesting an exercise-induced enhancement of prorenin conversion, possibly related to the exercise-induced stimulation of the fibrinolytic system. After aspirin, all 3 prorenin percentages are higher, implying that it interferes with prorenin conversion. Thus, exercise increases both fibrinolysis and PRA. The increase in PRA could be due in part to fibrinolytic enzyme conversion of prorenin (plasmin activates prorenin in vitro). Aspirin suppresses plasminogen activator, and thus prorenin activation.
Oral Administration Of Concentrated Factor VIII Or Factor IX Preparations
Authors: Sakuragawa N , Takahashi K , Horikoshi I
Published: July 2018

In current treatment and prophylaxis of bleeding of the patient of hemophiliacs, purified preparation of coagulation factors is administered intravenously, though some hemophiliacs developed hepatitis and antibody to factor VIII. If we can replenish coagulation factors orally, it will be of great benefit for hemophiliacs. Materials and methods: (1) Concentrated factor VIII( Con- coeight, Green-Cross, Osaka ) and factor IX( Konyne, Cutter ) were used. (2) Coagulation factors were enclosed in liposomes with addition of aprotinin, and the liposomes were further encapsulated in enteric capsules or suspended in milk to administer orally. (3) Coagulation studies were performed by assaying a-PTT, PT and coagulation assay methods.

Results: (1) A concentrated factor IX preparation ( 400 units ) was given to dogs orally to find raised concentration of factors IX, X, VII and II from 100% to 150%. Konyne contained these factors mentioned above. When 1200 units of Konyne was given in the same way, intravascular coagulation was ocurred. These results showed factor IX including factors X, VII and II were absorbed from the intestine. (2) A concentrated factor VIII preparation ( 100 units ) was administered orally to hemophiliacs as volanteers to find raised concentrations of several percentage by biological activity and about 30% by immunological activity. (3) It is absolutely necessary to add aprotinin to prevent proteolysis by the proteases in the gastrointestinal tract in the intestine capsules. One of the absorption mechanism may be due to the fusion of the upper layer membrane of liposome with the gastrointestinal cell.
Combination Therapy Of Sulphinpyrazone And Fibrinolytic Enhancing Agents In The Management Of Recurrent Thrombophlebitis In Behcet’S Disease
Authors: Afifi A M
Published: July 2018

The efficacy of combined sulphinpyrazone and fibrinolytic enhancing drugs in preventing recurrences of superficial and deep thrombophlebitis in 15 patients with Behcet’s disease was compared with that of fibrinolytic enhancing agents and placebo in a double blind crossover trial. Patients in the trial developed recurrent superficial and deep thrombophlebitis while on anticoagulants and immunosuppresive drugs. Analysis of the treatment results revealed that both protocols were virtually equipotent in controlling superficial thrombophlebitis as evidenced by reduction in the number of these episodes to 14% or less by either protocols. This contrasts with the wide difference in the response rate of the deep vein thrombosis in the two protocols, where 10 patients (66%) developed deep thrombotic episodes during therapy with placebo and fibrinolytic enhancing agents while only 3 (20%) developed these episodes during therapy with sulphinpyrazone and fibrinolytic enhancing agents. This statistically significant difference in response rate, (p<0.01) suggests that sulphinpyrazone is particularly effective in preventing recurrences of deep thrombophlebitis in Behcet’s disease.
Cation Contributions To Potentiometric Dye Responses In Human Platelets
Authors: Miller Jonathan L , Mazer Theodore M
Published: July 2018

The potentiometric dye, Di-S-C3(5) (0.9μM) was incubated with stirred suspensions of-3washed human platelets (5-6 × 104μl) in buffer containing 137mM NaCl, 2.7mM KCl, 0.2% dextrose, and 25mM Tris-HCl, pH 7.4 in an Ami neo-Bowman fluorometer, with excitation λ 620nm and emmision λ 680nm. Equilibrium levels of fluorescence (F) nearly doubled as external potassium concentration (K °) was increased from 2.7mM to 105.6mM. Addition of°2mM CaCl2 always produced immediate transient increases in F, regardless of K0 , in contrast to the increases or decreases produced by the potassium ionophore, valino- mycin (VAL) (5μM) in adherance to the electrochemical gradient determined by the choice of K°. Equilibrium F was also relatively insensitive to external calcium (Ca°) at either high or low levels of K . However, add?tion of the calcium ionophore, A23187 (10μM) produced immediate increases in F, with peak values (F-A23peak) increasing sharply with increasing Ca°. At a given Ca the F-A23peak was insensitve to K ° however, with the prior addition of VAL, F-A23peak became sensitive to K , particularly so at the lowest values of K° , where the F-A23peak was significantly lower in tne presence than in the absence of VAL. Replacement of sodium by choline had no significant effect on equilibrium F or on responses to VAL or to A23187- Platelet agglutination induced by ristocetin plus cryoprecipitate was not accompanied by similar increases in F.

These findings suggest that Di – S–C3 F reflects a strong contribution of potassium permeability in the resting state, with relatively little contribution by calcium. Production of an activated state in platelets by A23187, in contrast, results in an increase in F that appears to reflect a major contribution by calcium.
The Existence Of T Helper (Tμ) And T Suppressor (Tγ) Cells For The Generation Of Monocyte Thromboplastin Activity By Lipopolysaccharides
Authors: Levy G A , Schwartz B S , Edqinqton T S
Published: July 2018

Human peripheral blood mononuclear cells (PBM) in response to LPS stimulation generate increased quantities of thromboplastin activity. Monocytes are the cellular source of this activity and direct lymphocyte collaboration is required for its expression. PBM were separated by adherence into monocyte and lymphocyte fractions. Lymphocytes were further fractionated into T and non-T cells by rosetting with neuraminidase treated SRBC. 1 × 105 monocytes had a basal activity of 250 mU which increased to a maximum 2850 mU when monocytes were stimulated by 10 ug LPS for 6 hrs at a T cell: monocyte ratio of 4:1. No increase in thromboplastin activity was observed when monocytes were stimulated by LPS either alone or in the presence of non-T cells. Moretta et al. have described a system in which T cells are segregated into helper and suppressor subsets according to their ability to mediate immunoglobulin synthesis in response to pokeweed mitogen (PWM) stimulation. Using this system, T cells were further subfractionated into helper (Tμ), suppressor (Tγ ) and T null cells by cytoadherence to IgM or IgG coated ox RBC. 1 × 105 monocytes when incubated with increasing numbers of Tμ cells generated a maximal 4150 mU thromboplastin activity as the ratio of Tμ: monocytes approached 4:1. No increase in monocyte thromboplastin activity was observed above basal levels of 160 mU when monocytes were stimulated by LPS in the presence of either Tγ or T null cells. Tγ cells were observed to suppress Tμ helper cell function with a decrease in monocyte thromboplastin activity from 4150 mU to 1100 mU as the Tγ: Tμ ratio increased from 0:1 to 4:1. Thus, at least two populations within the T lymphocyte series the T μ (helper) and Tγ (suppressor) fractions modulate the expression of thromboplastin activity by monocytes.
Requirement Of Fibrinogen And Calcium For Inter-Platelet Bridges In Platelet Aggregation: A Hypothesis
Authors: Kornecki Elizabeth , Niewiarowski Stefan
Published: July 2018

Fibrinogen and calcium are required for the aggregation of platelets stimulated by ADP or pre-treated with proteolytic enzymes. Specific platelet surface fibrinogen binding sites (receptors) are exposed after platelets are stimulated by ADP or pre-treated with Chymotrypsin or pronase. It has previously been shown in our laboratory that an intact, symmetrical fibrinogen molecule is essential for fibrinogen binding and fibrinogen-induced aggregation of both ADP-stimulated and proteolytically-treated platelets. Here we propose that the mechanism by which fibrinogen and calcium aggregate platelets is by forming inter-platelet bridges linking the fibrinogen receptors of adjacent platelets together. In support of this proposition are the following new lines of evidence: 1) The fibrinogen-induced aggregations of ADP-stfiliulated or proteolytically-treated platelets are inhibited by high concentrations of fibrinogen (Ki=2.6 and 8.5 × 10 5M, respectively). The fibrinogen binding sites on adjacent platelets, at these concentrations, would be saturated by fibrinogen and therefore no inter-platelet fibrinogen bridges could be formed to hold the platelets together. 2) ADP-stimulated or chymotrypsin-treated platelets aggregated by fibrinogen are deaggregated by Chymotrypsin or pronase and this deaggregation coincides with the loss of 125I-fibrinogen from the platelet surface. 3) Preincubation of platelets with EDTA results in inhibition of both platelet aggregation and 125I-fibrinogen binding. Following the aggregations of ADP-stimulated or of chymotrypsin-treated platelets by fibrinogen, the addition of EDTA to the platelet aggregates results in both their deaggregation and their loss of bound 125I-fibrinogen. Thus it appears that divalent cations, especially calcium, are essential for the formation of fibrinogen-linked platelet aggregates.
Platelet Deposition In Extracardiac Conduits In Humans: A Noninvasive Quantification
Authors: Agarwal K C , Dewanjee M K , Fuster V F , Feldt R H , Wahner H W , Robertson J S , Brown J H , Chesebro J H , Kaye M , Puga F J , Danielson G K
Published: July 2018

In right-sided extracardiac conduits inserted for surgical correction of certain congenital heart defects, obstruction is a significant late complication requiring conduit replacement in about 5% of the patients. Our histopathologic study indicates a significant role of platelets in the pathogenesis. This study was designed to evaluate the feasibility of imaging procedure with 111-Indium-labeled platelets to localize and later quantify platelet accumulation in the conduits. Four patients with pulmonary atresia and 1 with truncus arteriosus underwent conduit placement, 2 with and 3 without porcine valve. Autologous platelets collected.from 43 ml of blood drawn prior to surgery were labeled with 111-Indium-oxine in ACD-saline medium (labeling efficiency 50-60%). 400-500 μCi of autologous 111-Indium-labeled platelets were administered intravenously immediately after surgery in 4, and five days after surgery in 1 patient. Imaging was performed at 6 hours and daily for 1-4 days using a portable gamma camera interfaced with a computer and fitted with medium-energy parallel hole collimator. Platelet deposition was noted at the site of the conduit in all 5 patients. Maximum accumulation (0.1-0.3% of dose) of platelets was seen within the first 24 hours. Activity over blood pool and lungs declined and over liver and spleen increased in a predictable manner over the imaging period. This technique has potential use as a noninvasive quantitative tool to study the effect of platelet inhibitor drugs in preventing conduit obstruction.
Regulation Of The Fatty Acid Composition Of Platelet Phospholipids
Authors: McKean M L , Smith J B , Silver M J
Published: July 2018

The fatty acid composition of cell membrane phospholipids does not remain constant after de novo biosynthesis, but undergoes continual remodelling. One of the major routes for remodelling probably includes the deacylation-reacylation steps of the Lands Pathway. This has been shown to be important for the incorporation of long chain, polyunsaturated fatty acids into phospholipids by liver and brain. An understanding of the mechanisms involved in these processes in platelets is especially important in light of the large stores of arachidonic acid (AA) in platelet phospholipids and the role of AA in hemostasis and thrombosis. Previous results from this laboratory have shown that the turnover of radioactive AA, 8,11,14-eicosatrienoic and 5,8,11,14,17-eicosapentaenoic acids in the phospholipids of resting platelets is more rapid than the turnover of radioactive C16 and C18 saturated and unsaturated fatty acids. However, little is known about how fatty acids, especially AA and its homologues, are incorporated into platelet phospholipids during de novo biosynthesis or how they are exchanged during remodelling.

At least three enzymes are involved in the deacylation- reacylation of phospholipids: phospholipase A2; acyl CoA synthetase; and acyl CoA transferase. We have studied acyl CoA transferase and have found considerable activity in human platelet membranes. Experiments are in progress to determine the substrate specificity and other properties of this enzyme.
Development Of A Rapid Immunometric Assay For The Factor VIII Antigens In Plasma
Authors: Gaffney P J , Mahmoud M , Barrowcliffe T W , Kemball-Cook C
Published: July 2018

Immunoradiometric assays (IRMA) for both Factor VIII related antigen (VIII R:Ag) and Factor VIII clotting antigen (VIII C:Ag) have been described, involving the preparation of specific radiolabeled antibodies. The alternative methodology described in this report for F VIII R:Ag involves three steps:1. the immobilization of a crude preparation of F VIII R:Ag (0.05 units/ml) on polyvinyl plates (96-well); 2. the incubation of known amounts (defined by a freeze-dried plasma standard) of F VIII R:Ag with a predetermined excess (1/2000) of a monospecific rabbit antiserum to F VIII R:Ag (called first antibody, ab'); 3. the binding of the various residual amounts of ab' (from step 2) to the immobilized F VIII R:Ag. The residual ab' bound to the coated polyvinyl plates is measured using a radiolabeled (125I) goat IgG to rabbit IgG (called second antibody, ab"). Thus the radioactivity in each well of the plate is inversely related to the amount of F VIII R:Ag in the incubation mixture. A linear standard curve was established for pooled plasma between 0.04-0.002 units of F VIII R:Ag per ml. Test plasmas gave parallel dose-response curves with this standard curve. Thus the F VIII R:Ag levels in a variety of plasmas and concentrates were established and found to correlate with data obtained by Laurell ‘rocket’ immunoelectrophoresis. A similar assay to that described above has been attempted for Factor VIII C:Ag in plasma. The antiserum was obtained from a patient with an elevated level of ‘inhibitor’ (antibody) to F VIII C:Ag. A standard curve has been established using a highly purified preparation of F VIII C:Ag between 0.3-0.01 u of coagulant activity. This form of immunometric assay has the following advantages:1. a source of highly purified antigen is not necessary; 2. no isolation or labelling of the F VIII antibodies is required; 3. the assay is sensitive, technically simple and rapid.
The Effects Of Defibrination Procedures In Plasma At III Assays
Authors: Philo R D , Gaffney P J
Published: July 2018

Defibrination procedures using heat or ancrod (a coagulant fraction from Agkistrodon rhodostoma venom) are commonly used in heparin co-factor AT III assays which use a clotting time endpoint to measure residual thrombin. The test plasma is compared to a freeze-dried standard plasma which has been defined to contain a particular number of AT III units. This study examines the effects of both heat and ancrod defibrination of fresh and freeze-dried plasmas using a chromogenic-based AT III assay system in which no defibrination is necessary.

The mean potency of the heat (56°C × 15 min) defibrinated freeze-dried standard plasma compared to the untreated standard was 0.51 (mean of 9 assays using a multidose bioassay design) whereas the corresponding figure for the frozen test plasmas was 0.72. Thus a test plasma would give falsely elevated readings for AT III compared to a freeze- dried plasma standard. Assays of ancrod (0.55 iu/ml) defibrinated plasma against untreated plasma gave mean potencies not significantly different from 1.0 for both the freeze-dried and frozen plasmas. Thus this form of defibrination allows a more valid comparison of fresh and freeze- dried plasmas.

Analysis of these samples by crossed immunoelectrophoresis in the presence of heparin confirms that the ancrod defibrinated samples are essentially identical to the untreated ones whereas the heat defibrinated samples are altered, the freeze-dried plasmas markedly so.

The data suggest that, should a defibrination step be used in the assay of AT III, the use of ancrod is preferred to heat. The heparin co-factor AT III assay using the chromogenic substrate, S-2238, to measure residual thrombin would seem the preferred method of assay in order to avoid any defibrination step.
In Vivo Studies on the Inhibition of Coagulation by Fractionated Heparin and by a Heparin Analogue I. Effects of Heparin Fractions
Authors: Schmitz-Huebner U , Balleisen L , Asbeck F , Loo J van de
Published: July 2018

High and low molecular weight heparin fractions obtained by gel filtration chromatography of sodium mucosal heparin were injected subcutaneously into six healthy volunteers and compared with the unfractionated substance in a cross-over trial. Equal doses of 5,000 U were administered twice daily over a period of three days and heparin activity was repeatedly controlled before and 2, 4, 8 hrs after injection by means of the APTT, the anti-Xa clotting test and a chromogenic substrate assay. In addition, the in vivo effect of subcutaneously administered fractionated heparin on platelet function was examined on three of the volunteers. The results show that s.c. injections of the low molecular weight fraction induced markedly higher anti-Xa activity than injections of the other preparations. At the same time, APTT results did not significantly differ. Unfractionated heparin and the high molecular weight fraction enhanced ADP-induced platelet aggregation and collagen-mediated MDA production, while the low molecular weight fraction hardly affected these assays, but potently inhibited thrombin-induced MDA production. All heparin preparations stimulated the release of platelet Factor 4 in plasma. During the three-day treatment periods, no side-effects and no significant changes in the response to heparin injections were detected.
Urokinase Therapy in Neonates with Catheter Related Central Venous Thrombosis
Authors: Wever M L G , Liem K D , Geven W B , Tanke R B
Published: July 2018

The results of fibrinolytic therapy with urokinase were evaluated in 26 neonates with catheter related central venous thrombosis. Complete thrombolysis could be achieved in 13 patients (50%), partial thrombolysis in 3 patients (12%). No effect was seen in 10 patients (38%). Therapy success was influenced by age, size and location of the thrombus. Coincidence of infection occurred in 16 patients (62%). Mild hemorrhagic complications were seen in 2 patients (8%), no other significant side effects were observed. Nine patients with residual thrombosis were treated with oral anticoagulants following urokinase resulting in resolution of the thrombus in 6 patients within 3 months (67%). The incidence of asymptomatic recurrent thrombosis was high (28%). Urokinase might be an effective and safe treatment for central venous thrombosis in neonates. Prophylactic antibiotic therapy during the infusion of urokinase and long-term treatment with oral anticoagulants after thrombosis are advisable. Early detection of thrombosis might enhance the success rate of fibrinolytic therapy. Therefore, we strongly recommend routine echocardiographic screening of central venous catheters.
Lymphocyte Adhesion to Human Endothelial Cells Induces Tissue Factor Expression via a Juxtacrine Pathway
Authors: Schmid Esther , Müller Thomas H , Budzinski Ralph-M , Pfizenmaier Klaus , Binder Klaus
Published: July 2018

To study the effect of lymphocyte adhesion on the procoagulant activity of endothelial cells, we have stimulated HUVECs with interferon-γ to upregulate adhesion molecules. Subsequent addition of lymphocytes induced the expression of tissue factor (TF) by HUVECs. Both CD4+ and CD8+ T-cells promoted this TF synthesis via distinct adhesion molecules (CD4+ T-cells: E-selectin and ICAM-1; CD8+ T-cells: MHC-I molecules). In addition, tumor necrosis factor-α and -β (TNFα, TNFβ) and platelet-activating factor (PAF) were involved in lymphocyte-mediated TF expression on HUVECs. We demonstrate that PAF plays a pivotal role in this process. Adhesion of lymphocytes to endothelial cell surface molecules induced the release of PAF. PAF, in turn, caused the production of T.NFα and TNFβ, both of which are potent stimulators of TF expression.
Measurement of Activated Factor IX in Factor IX Concentrates: Correlation with In Vivo Thrombogenicity
Authors: Gray Elaine , Tubbs Jill , Thomas S , Oates A , Boisclair M , Kemball-Cook G , Barrowcliffe T W
Published: July 2018

Current in vitro tests for thrombogenicity of FIX concentrates used for prothrombin complex concentrates (PCCs), are of little value when applied to high purity FIX (HP FIXs). In the present study, we have developed a chromogenic assay for activated FIX (FIXa) and evaluated its ability to predict in vivo thrombogenic potential of HP FIXs in a modified Wessler stasis model. Among the HP FIXs, only 1 out of 7 products had no detectable FIXa; this product also showed no in vivo thrombogenicity. In the other 6 products, FIXa content ranged from 0.15–1.2 U/1000 iu FIX, and all showed some evidence of in vivo thrombogenicity, with mean thrombus scores ranging from 0.25–4. There was a significant positive correlation (r = 0.55, p <0.02) between FIXa levels and in vivo thrombogenicity of HP FIXs. NAPTT data were not significantly correlated with the in vivo results and the TFCT also showed no direct correlation with the mean thrombus score. These results indicate that HP FIXs may still carry a small residual thrombotic risk and measurement of FIXa content of these products may be a better predictor of thrombogenicity than the current in vitro tests.
Localization of a Vitronectin Binding Region of Plasminogen Activator Inhibitor-1
Authors: Padmanabhan Jaya , Sane David C
Published: July 2018

The PAI-1 binding site for VN was studied using two independent methods. PAI-1 was cleaved by Staph V8 protease, producing 8 fragments, only 2 of which bound to [125I]-VN. These fragments were predicted to overlap between residues 91-130. Since PAI-2 has structural homology to PAI-1, but does not bind to vitronectin, chimeras of PAI-1 and PAI-2 were constructed. Four chimeras, containing PAI-1 residues 1-70,1-105,1-114, and 1-167 were constructed and expressed in vitro. PAI-1, PAI-2, and all of the chimeras retained inhibitory activity for t-PA, but only the chimera containing PAI-1 residues 1-167 formed a complex with VN. Together, these results predict that the VN binding site of PAI-1 is between residues 115-130.
Antithrombotische Wirksamkeit und Verträglichkeit von Embolex®-NM und MonoEmbolex®-NM beim elektiven und traumatischen Hüftgelenksersatz
Authors: Fritsche H. -M. , Lechner F. , Blümel G. , Haas S.
Published: July 2018

Niedermolekulare Heparine gelten derzeit als wirksamste Prophylaxe bei Patienten mit besonders hohem Thromboembolierisiko, wie z.B. nach elektivem oder traumatisch bedingtem Hüftgelenksersatz. Allerdings müssen antithrombotische Wirksamkeit und Verträglichkeit für jedes Präparat gesondert nachgewiesen werden. Diese Anforderungen sind für die zugelassenen Darreichungsformen von 3000 E anti-Xa Embolex®-NM und Mono-Embolex®NM erfüllt. Es konnte in einigen Studien sogar eine den bisher üblichen Prophylaxeregimes überlegene Wirksamkeit nachgewiesen werden.
Ambulante Thromboembolieprophylaxe mit niedermolekularem Heparin
Authors: Biegholdt M. , Haas S.
Published: July 2018

Bei Patienten mit Unverträglichkeitsreaktionen oder Kontraindikationen gegen orale Antikoagulanzien wurden Wirksamkeit und Verträglichkeit des niedermolekularen Heparins Fraxiparin im Rahmen einer ambulanten Langzeit-Thromboembolie-prophylaxe untersucht. Überwiegend handelte es sich hierbei um eine Rezidivprophylaxe nach vorangegangenen Thromboembolien.

Insgesamt 45 Patienten erhielten über 10 bis 683 Tage (im Mittel 164 Tage) eine s.c. Injektion von 36 mg Fraxiparin pro Tag. In 2 Fällen wurden nach frischen Unterschenkelvenenthrombosen im ersten Behandlungsmonat 2 Injektionen pro Tag verabreicht. Die Mehrzahl der Patienten injizierte sich das Präparat selbst.

Innerhalb der Gesamtbehandlungsdauer von 20 Jahren und 3 Monaten traten lediglich ein Thromboserezidiv und ein Rezidiv einer oberflächlichen Thrombophlebitis auf. Unerwünschte Begleiterscheinungen wurden bei 8 Patienten beobachtet und bezogen sich überwiegend auf lokale Hauterscheinungen und Injektionshämatome. Sie waren in allen Fällen reversibel.

Schwere Blutungskomplikationen, Thrombozytopenien, Transaminasenanstieg oder Osteoporose traten im Zusammenhang mit der Behandlung nicht auf. Gerinnungsparameter wie aPTT, Thrombinzeit, Quick-Wert und Antithrombin III wurden unter der Behandlung mit Fraxiparin nicht beeinflußt.

Die nur einmal täglich notwendige Injektion wurde von Patienten und Ärzten als erheblicher Vorteil und Erleichterung im Vergleich zu konventionellem Heparin angesehen und begrüßt. Sofern keine therapeutische Antikoagulation indiziert ist, stellt Fraxiparin somit eine Alternative zum Einsatz von oralen Antikoagulanzien oder niedrig dosiertem Heparin zur Rezidivprophylaxe von Thromboembolien dar.