bluuebunny/crossref_metadata_2025 huggingface dataset:
Blood Viscosity Variations During Raynaud’s Phenomenon
Authors: Forconi S.
Published: May 2019

Since several authors have found abnormal blood viscosity in patients suffering from Raynaud’s disease but did not study them at the moment when the phenomenon appeared, we want to find out whether haemorheological changes might be provoked in the patients during a cold-induced phenomenon. The study was conducted on ten selected patients suffering from Raynaud’s phenomenon only in one hand when exposed to cold. A relevant and statistically very significant increase of the viscosity was noted in the blood coming from the hand during the cold-induced ischemia. When the ischemia had disappeared, blood viscosity levels returned to those recorded before the experience. No variations were evident in either plasma and serum viscosity, or in packed red cell volume and in plasma fibrinogen concentration. In the other arm of the same patients, a much smaller increase in blood viscosity was noted. No variations were found in any of the parameters observed in six control subjects. These results seems to suggest that blood viscosity changes specifically in relation to the disease, that this change may be related to the behaviour of the red cells (increased aggregability or decreased deformability) as the consequence of the ischemia, and that this hyperviscosity may potentiate the hindrance to the flow at the microcirculatory level.
In Vitro Determination of Thromboembolic States: Cellular, Molecular and Genetic Aspects
Authors: Dintenfass L.
Published: May 2019

Techniques of the in vitro formation of artificial thrombi under controlled conditions of shear rates, and the principles of their formation and morphology, were established by Dintenrass and Rozenberg between 1963 and 1967, and Dintenfass and Stewart in 1968. Aggregation of platelets was found to be a direct function of shear rate. It has been later established that that an elevation of the apparent viscosity of artificial (red-white, and white) thrombi is a significant feature of renal failure, hypertension, myocardial infarction, and peripheral vascular disease. In diabetes mellitus, diabetic and nondiahetic retinopathy, severe anxiety and cigarette smoking there is a serious elevation of the blood viscosity factors. The apparent viscosity of artificial thrombi and of the other blood viscosity factorsis correlated with the fibrinogen level, and albumin/fibrinogen ratios. Significance limits of these correlations vary in different disorders, and the correlation coefficients may exhibit both the negative and the positive signs, mainly depending on the ABO blood groups. Abnormality of the blood viscosity factors can be used probably for detection of the silent states of various diseases, as well as for prediction of impeding cardiovascular episodes.
Interaction Between Low Molecular Weight Dextran Sulphate and Bentonite adsorbed Plasma
Authors: Matsuda T. , Ogawara M. , Seki T. , Miura R. , Yokouchi M. , Murakami M.
Published: May 2019

Effects of low molecular weight dextran sulphate (D.S.; mean molecular weight = 6,0006,500; S content = 17-19%) and/or bentonite (Wako Pure Chemical Industries, Osaka, Japan) adsorbed plasma (supernatant from plasma mixed with 300 mg/ml of bentonite for 10 minutes at 37°C), which contains no fibrinogen nor antithrombin III but α2 macro-globulin, α1-antitrypsin and α2-plasmin inhibitor, on thrombin time or batroxobin (purified fraction of venom of Bothrops atrox) time of fibrinogen solution were investigated. Addition of D.S. or bentonite adsorbed plasma to fibrinogen solution resulted in prolongations of thrombin time and batroxobin time. These results indicate that both D.S. and bentonite adsorbed plasma inhibit conversion of fibrinogen to fibrin by thrombin or batroxobin. However, when appropriately diluted bentonite adsorbed plasma and D.S. were added to fibrinogen solution simultaneously, thrombin times and batroxobin times were shorter compared to those when either bentonite adsorbed plasma or D.S. but not both was added. From these results, it is concluded that D.S. and bentonite adsorbed plasma interact to inhibit each other.
The Effect of Low Dose Heparin on the Reticulo-Endothelial System in Patients Undergoing Major Surgery
Authors: Bentley P. G. , Kakkar V. V.
Published: May 2019

The ability of the reticulo-endothelial system (RES) to remove activated procoagulants from the circulation may provide an important protective mechanism against thrombosis . In 11 patients undergoing major surgery, the phagocytic and catabolic activity of RES was measured using I-labelled microaggregated human serum albumin in a dose of 10μci of 125I and 5mg/kg of aggregates per test. RES function tests were performed before surgery and again exactly 24 hours postoper atively. Plasma samples were also taken for At III, fibrinogen and FOP levels. The phagocytic index and catabolic rate was determined using a computer programme. The phagocytic rate of the RES was not affected either by surgery or by low doses of heparin. However, the catabolic rate increased postoperatively in the control patients (p < 0 .00 5) and low-dose heparin prevented such an increase. The mechanism for such an effect is obscure, but it is suggested that heparin is taken up and stored by RES cells without prior degradation and that this up-take and storage slows the metabolic activity significantly. Further studies are needed to determine the importance of intra-cellular storage and release of heparin.
Autologous 131I-Antithrombin III Turnover Studies in Man
Authors: Reeve E. B. , Leonard B. , Zimet G. , Carlson T. H.
Published: May 2019

A simple one-step heparin sepharose preparation of better than 95% pure human antithrombin III (AT3) has been developed along with a modification of the iodine monochloride method that allows consistent iodination of it. Iodination at a ratio of ~0.7 I atom/1.0 AT3 molecule results in loss of ca. 5% total thrombin neutralizing activity but negligible reduction in rate of reaction with thrombin. After iodination with 131I unreached 131I-products are removed by washing the 131I-AT3 on a second heparin sepharose column. Before injection the *I-AT3 is sterilized by millipore filtration. By this method autologous *I-AT3 can be prepared and reinjected into the patient in 36-48 titor less if desired. Serial measurements are made of plasma 131I-AT3, plasma TCA-soluble 131I, and whole body 131I, (using a whole body counter). Since human AT3 is ~ the same molecular weight as albumin if allowance is made for its increased catabolic rate its kinetic behavior should be predictable from results of our earlier studies with human autologous albumin. This is not found to be so and human *I-AT3 shows the same delay in catabolismos dog *I-AT3. Current studies indicate that this anomalous behavior is unlikely to invalidate estimates of AT3 catabolism but may result in considerable errors in estimates of total Interstitial AT3. As with our dog studies these suggest a protective role for AT3 in the interstitial fluids
Plasma Concentrations of Platelet Specific Proteins Correlated with Platelet Survival
Authors: Doyle D.J. , Chesterman C.N. , Cade J.F. , Morgan F.J.
Published: May 2019

Relationships between platelet survival (51Cr) and plasma concentrations (radioimmunoassay) of β-thromboglobulin (βTC) and platelet factor 4 (PF4) were analysed in 91 studies of patients prior to and after coronary artery bypass surgery. Platelet lifespans were calculated using the linear, exponential, weighted mean (WM) and multiple hit (MH) models. The values obtained approximated normal distributions and all the indices correlated, r values ranging from 0.69 to 6 (p<0,001 in all cases).

βTC had significant negative correlations with all indices of platelet survival, the most significant being with MH (r = -0.39, p<0.001). Of interest was the correlation of βTG with the number of hits (n) in MH (r = -0.29, p<0.01). PF4 correlated with all indices of platelet survival except n, the most significant correlation was with WM (r = 0.33, p<0.01). BTC and PF4 were highly correlated (r = 0.62, p<0.001), however no benefit was obtained by combining measurements of the two proteins in any regression with platelet lifespan.

The results suggest that shortened platelet survival in vascular disease is associated with platelet release of BTG and PF4 and that these platelet specific proteins are comparable as markers of platelet activation in vivo.
Diminished Prostacyclin Generation in Human and Experimentally Induced (Streptozotocin, Alloxan) Diabetes Mellitus
Authors: Silberbauer K. , Schernthaner G. , Sinzinger H. , Clopath P. , Piza-Katzer H. , Winter Maya
Published: May 2019

Platelet dysfunction may play an important role in diabetic microvascular disease. We therefore studied prostacyclin (PGI2) production in the forearm veins of patients with juvenile-onset diabetes (JOD; n = 15) and in 15 healthy subjects. In 10 male Wistar rats diabetes was induced by a single i. v, dose of streptozotooin and in a further 10 male Wistar rats diabetes was induced an alloxan diabetes. From these animals, as well as from 10 control rats abdominal aortic tissue was removed for PGI2-estimation. PGI2-formation was studied using Moncada’s bioassay based upon its potent platelet aggregation inhibiting effect (the standard was kindly supplied by Dr. John E. Pike, The Upjohn Company, Kalamazoo, Michigan, U.S.A.). Prostacyclin generation of the tissue was significant (2p < 0.02) diminished in the JOD-patients as well as in experimental diabetes compared to the respective control groups. The decrease in PGI2-synthesis might play an important role in initiating and/or promoting the vascular complications of the disease
Release Reaction from Human Platelets Submitted to Viscometric and Tube Flow
Authors: Rieger H. , Forst R. , Schmid-Schönbein H. , Krings W.
Published: May 2019

A reliable sign of stimulation of human platelets is the release reaction (RR) as measured by the amount of cellular H-C-serotonin set free in the medium. Until now only chemical mediators were known as being able to trigger the RR. According to our experiments flow forces can also induce the RR in the absence of any aggregating substance. In vis-cometric flow we find a strong correlation (r=0.982) between the amount of H-C-serotonin set free in the plasma and the extent of occuring pla telet aggregation. A similiar result we obtained in platelet rich plasma submitted to a stationary laminar tube flow. A positive correlation can be established between constant wall shear stresses and the 14-C-seroto nin release. In counter-distinction to this, the lactate-dehydrogenase of the platelet cytogel is not released by shearing. These data suggest that the RR of human platelets can be induced mechanically and that the extent Of this reaction is proportionalto the intensity and time of shearing.
Generation of Prostacyclin (PGI2) Activity by Lewis Lung Carcinoma (3LL) Cells
Authors: Poggi A. , Dall’Olio A. , Balconi G. , Delaini F. , de Gaetano G. , Donati M.B.
Published: May 2019

The role of prostaglandins in tumour growth is still a natter of discussion. We have inve stigatcd the capacity of tissues and cultured cells from a murine metastasizing tumour (3LL) to generate PGI2 activity. Intact fragments and homogenatcs from 51.1. tissues, collected at different intervals after i.m. implantation of 2 χ 105 cells into a hind leg of CS7B1/6J mice, were incubated in Tris buffer at pH 7.4 for 5-15 min or at pH 9.2 for. 45 min. The supernatant showed strong inhibitory activity against ADP-induced human platelet aggregation with many characteristics in common with PGI2. It was progressively released into the medium, was labile at acidic pH, was inhibited by indomethacin both in vitro and ex vivo, was potentiated by theophylline and, unlike PGD2, it inhibited aggregation of rat platelets. Muscular tissues from the contralateral leg had very low inhibitory activity in comparison. Metastatic lungs showed 10-20 times the activity of the primary tumour tissue, similar(however, to the inhibitory activity of normal lungs. Isolated metastatic tissue had lover activity than surrounding lung tissue, but still higher than the primary tumour. 3LL cells harvested from primary cultures of the tumour showed strong PGI2 actiyity. These data suggest that in 3LL tissues PGI2 activity may derive not only from their vascular net but also from the cancer cells themselvcs. [Supported by a grant from Italian CNR).
Platelet Aggregability and Thrombogenesis During Chemical Carcinogenesis
Authors: Görög P. , Kovacs I.B. , Born G.V.R.
Published: May 2019

There are suggestions that the metastatic spread of malignant tumours depende inter alia on platelets. To find out whether platelets are affected by a carmon technique for experimentally inducing malignancies, rats and mice were injected with 3-methylcholan-tnrene to induce non-metaStasising fibrosarccmata. After 14-21 days, platelet aeeree-atic-n measured in vitro (Born, 1963, Nature, 194, 927) and arteriolar thrcmbogenesis by laser irradiation in vivo (Kovaes et at ., 1973, Microvascular Research, 6, 194) were significantly inhibited. Therafter these inhibitions disaopeared but reaopeared when tumours became palpable, before any change in blood platelet concentration
Vascular Diseases are Essentially a Matter of Haemorheology and of Haemostaseology
Authors: Hess H.
Published: May 2019

Vascular diseases have, in different segments of the vascular tree, a different morphology. There is an arteriosclerosis but not an obliterating arteriosclerosis of thoracic and of proximal abdominal aorta. The obliterating arteriosclerosis exists in the branches and in the continuation of these vessels up to the small arteries. Changes similar to arteriosclerosis don't exist either in the terminal vascular bed nor in veins. The material and structure of obliterations is differently composed in different segments of vessels. All those differences are essentially caused by the different local conditions of haemorheology and haemostaseology. A very important point is - the aggregation and adhesion of platelets is posi tively correlated with the shear rate, whereas the activation of the plasmatic coagulation and the development of aggregates of erythrocytes is negatively correlated with the shear rate. The consequences of this for the different vascular segments are fully discussed in the lecture.
Cancer Cell Procoagulant Activity
Authors: Gralnick H.R.
Published: May 2019

Studies of the procoagulant activity (PA) of human acute leukemia cells (ALC) has demonstrated that acute lymphocytic leukemia (ALL) had approximately 25-50% of the PA of normal human granulocytes, while lymphoid leukemias had increased PA. Acute promyelocyte leukemia (APL) had approximately 4-8 times the activity of normal granulocytes while acute myeloblastic leukemia (AML) cells had the same activity as normal. The PA was characterized as tissue factor (TF). Two correlations of the TF activity in ALC is the incidence of the fibrinogen kinetics and intravascular coagulation. He have done fibrinogen survivals on 15 patients with AL. The results of these kinetic studies have revealed that in 5 patients with ALL the T/2 is 2.94 ± .31 days; fraction catabolic rate (FCR) 29.8 ± 4.3%/day were slightly different from the control of T/2 3.69 ± 0.45 days; FCR 22.1 ± 2.5K/day. In 3 patients with AHL, the T/2 was 1.92 ± 0.79 days; FCR 44.2 ± 20.6%/day. In patients with APL, the fibrinogen survival revealed a T/2 of 0.069 ± 0.25 days; FCR 160.9 ± 62.7%/day. The use of anticoagulants in APL markedly decreases death from hemorrhage. In other investigations we have studied the effect of anticoagulation on the spread of peripheral sarcomas in man. Warfarin anticoagulation was used as an adjunct t o amputation of sarcomas. The results of these studies are quite encouraging in that the patients appear to have a longer remission and to have better overall survival.
Fibrin and Cancer Cell Growth: Problems in the Evaluation of Experimental Models
Authors: Poggi Andreina
Published: May 2019

Studies on the role of fibrin in experimental cancer growth should take into account the following problems: 1) during growth and dissemination of experimental tumours, haemostatic changes may occur which vary depending on the route of inoculation of cancer cells and on the tissue wherethe tumour grows. Thrombocytopenia, haemolytic microangiopathic anaemia and decreased survival of fibrinogen were observed during spontaneous dissemination to the lung of i.m. implanted Lewis Lung Carcinoma cells, not when metastatic growth occurred after surgical removal of the primary tumour or lung nodules developed following i.v. injection of the same cells. 2) Treatment of experimental tumours with drugs active on the haemostatic system may have different effects depending on the stage of growth of the tumour. This observation (which we have made with both warfarin and a defibrinating enzyme in murine metastasizing tumours) could suggest that fibrin may play different roles at different phases of cancer cell growth. 3) The supposed antiturooral activity of drugs active on the haemostatic system may be also influenced by other factors, such as a direct activity on cancer cells or on the host’s immune system or on blood supply to the tumour. As an example, non steroidal antiinflammatory drugs may act not only as antiplatelet agent but also as inhibitors of prostaglandin synthesis by cancer cells and some snake venoms may influence cancer cell growth not only through defibrination, but also with their iinnu-nodepressant properties. (Supported by Italian CNR and NIM. MCI, USA).
Coumarin Anticoagulation and Experimental Metastases
Authors: Hilgard P.
Published: May 2019

Under experimental conditions oral anticoagulants of the coumarin type retard primary tumour growth and drastically reduce the incidence of distant metastases from rodent tumours. Employing different dose schedules it was found that satisfactory therapeutic anticoagulation and long-term treatment are pre-requisites for the anti-tumour activity. The equivocal effects of other anticoagulants, such as snake venoms, heparin and “antiplatelet” drugs on tumour growth and dissemination suggest a unique mechanism of coumarin derivatives. There is experimental evidence that the anti-tumour action is not a direct drug effect on malignant cells but specifically mediated by the vitamin K antagonism of the coumarins. Altered blood coagulability appears not to be the sole mechanism by which oral anticoagulants exert their anti-metastatic effect.

There is a theoretical role for vitamin K dependent γ-carboxyglutamic acid containing proteins in interactions between tumour and host. Vitamin K deficiency increases tne phagocytic activity of peritoneal macrophages after nonspecific immunostimulation; this might be an important factor in controlling tumour growth. The synthesis of serine proteases with clot promoting activity by tumour cells is partly vitamin K dependent; these proteases could also nave considerable implications for the metastatic potential of blood borne cancer cells.
Platelet Survival Half-Life in Coronary Disease Patients under and over age 50
Authors: Chesebro J.H. , Fuster V. , Byrne J.H. , Olson D.A. , Kazmier F.J.
Published: May 2019

Coronary disease (CAD) is usually severe when t he first symptoms appear. Thus identification of CAD patients (PTS) prior to symptom on set would be desirable for future preventive interventions. Since we found that cigarette smoking ≥10 pack-years (S) and family history of CAD at a young age (FH) are the 2 most important risk factors in CAD, and since platelet deposition to damaged endothelium appears important in the pathogeneais of atherosclerosis, platelet consumption was estimated by 51Cr platelet survival half-life (PS) in 3 groups of subjects:(1) in 49 accarentlv normal individuals without the risk factors (AN, no RF), (2) in 36 apparently normal individuals with the risk factors (AN, with RF), and (3) in 114 symptomatic PTS with angiographically proven CAD.

There Is (1) a probable role of platelets in CAD as suoported bv the shortened PS, (2) a possibility that S or FH when associated with short PS (≤92 hrs) might identify the AN individual ≤ age 50 who is developing CAD, (3) a lower percent of CAD PTS >age 50 with a shortened PS and perhaps lesser probability of identifying asymptomatic CAD PTS >-age 50.
Relationship Between Aggregation and Binding of 125I-Fibrinogen and 45Calcium to Human Platelets
Authors: Peerschke E.I. , Grant R.A. , Zucker M.B.
Published: May 2019

Since calcium and fibrinogen are essential cofactors for ADP-induced aggregation, their mechanisms of action were investigated. Aspirin-treated platelets were filtered through Sepharose 2B equilibrated with cation-poor Tyrode’s solution. After adding the radioactive compounds at 22, platelets were centrifuged through silicone oil. Trapping was assessed in separate samples with 14C-sorbitoi. Calcium binding was maximal at 1 hr and with 200 uH CaCl2. Two binding sites could be demonstrated on normal and thrombasthenic platelets and on platelets which had lost their ability to aggregate (but not to change shape or promote clot retraction) after treatment with EDTA (8 min, 37°, pH 7.8). ADP did not alter calcium binding in the presence or absence of fibrinogen. Fibrinogen, however, bound to normal gel filtered platelets in the presence of ADP and ionized calcium “ but not to thrombasthenic or EDTA-treated platelets or to normal platelets in the presence of EDTA or at pH 6.5. Binding of fibrinogen increased with concentration but saturation was not observed even at physiologic levels. Fibrinogen binding was similar in gel filtered platelets and citrated piatelet-rich plasma. These studies indicate that stimulation of platelets with ADP under conditions favorable to aggregation is associated with binding of fibrinogen but not of additional calcium.
Responses of Afibrinogenemic Platelets to Bacteria
Authors: Clawson C.C. , White J.G.
Published: May 2019

The interaction of platelets from total congenital afibrinogenemia (CA) with bacteria (Staph. aureus 502A) has been examined by aggregometry and electron microscopyIt has oeen shown (Am.J. Path. 65:367, 1971) that normal human platelets in autologous plasma or as albumin-washed platelets (AWP) in saline respond with several strains of bacteria with a full physiologic aggregation reaction. This response is similar to the aggregation reaction with particulate collagen, and results in incorporation of the bacteria into the aggregates between but not within platelets. In contrast, CA platelets responded much more slowly to contact with bacteria, forming very small aggregates. Most striking was the frequent engulfment of bacteria by single CA platelets Lanthanum staining demonstrated that the Bacteria entered the CA platelets via the open canalicular system which remained open to the extracellular media. Replacement of human purified fibrinogen to the CA platelets greatly accelerated the aggregation reaction but did not eliminate the engulfiment phenomenon. This engulfment process could not be duplicated in normal AWP. Comparisons of interactions with bacteria by CA platelets and normal AWP with and without replacement fibrinogen demonstrated that fibrinogen is more important to the platelets response to bacteria than to its response to collagen.
Isolation, Partial Purification and Characterisation of A Plasminogen Activator Produced by Endothelial Cells in Vitro
Authors: Laug W.E.
Published: May 2019

Cloned endothelial cells obtained from the aorta of 1-2 day old calves produced high fibrinolytic activity, which was 90% dependent upon the presence of plasminogen when grown on 125 I fibrin coated dishes. High plasminogen-dependent proteolytic activity was also demonstrated in the cell lysate and in the culture medium of the cells. The production and secretion of this prtitease were found to increase during the log phase of cell growth and to reach a maximum at con fluency. Thereafter they remained constantly high. This protease, partially purified from the culture medium of confluent endothelial cell cultures, is aiginine specific and activates plasminogen by piOteolytic cleavage to plasmin. Its proteolytic activity which is highest in the pH range of 7.5 to 8.0 is irreversibly inhibited by diisopropyl fluorophosphate, suggesting that it is a serine protease. The molecular weight of this protease is approximately S2000.
Human Vascular Plasminogen Activator (VPA) and Fibrinolytic Specificity
Authors: Allen R. , Pepper D.S. , Scotland S.E.
Published: May 2019

We have isolated VPA from cadaver limbs and venous occlusion plasma. The enzyme circulates as a complex with soluble high molecular eight derivatives of fibrinogen (SFP) and may be purified as such in the presence of Trnsylol by PEG precipitation, hydroxylapatite, gel filtration and final dissociation on lysineagarose. VPA behaves differently to plasminogen in that it is eluted from a lTBine-agarose column by a higher lysine concentration than plasminogen. By partially digesting fibrinogen (to X+Y+D) and polymerising the product with thrombin plus F XIII it was possible toprepare stable crosslinked SFP which formed strong complexes in solution (and when insolubliaed on agarose) with VPA. A more convenient form of synergist than polymeric fibrin was sought for spectrophotometric use and found in poly-D-lysine (MW144,000) which at 1mg/ml showed a 6 fold enhancement in the rate ot plasmin generation in a VPA-Plg-E2251 assay system. This synergism was not seen with UK, SK or pig heart tissue activator and human melanoma activator showed only a slight enhancement. We propose that simultaneous binding of VP activator (to a polymeric fibrin-specifie site) and plasminogen, together with associated conformational transitions and exclusion of inhibitors is the basis of the specificity or VPA for fibrinolysis over fibrinogenolysis.
On the Origin of Plasma Plasminogen Activator in Man
Authors: Marsh N.A. , Gaffney P. J.
Published: May 2019

In the past, plasma plasminogen activators have been considered a wide group of dlssimiliar substances originating in plasma (the so-called “intrinsic” activators) and outside the vascular compartment (“extrinsic” activators). The “intrinsic” group appear to share the common property of being sensitive to complement Ci-lnactivator A series of simple experiments performed in our laboratories indicate however that these substances may not be quite so diaparate. We have measured “Intrinsic” and “extrinsic” plasminogen activators before and after exercise, a procedure known to elevate systemic levels. Activators were precipitated in the euglobulin fraction by diluting plasma L in 10 and acidifying to pH 5.9. The resuspended precipitate was tested on conventional and rapid fibrin plates using sodium flufenamate (an inhibitor of complement lnactivator) Co distinguish between the two “kinds” of activatorThe results showed a marked increase In “extrinsic” activator liberated by the vascular endothelium in response to exercise, the level of which quickly declined. By contrast theIntrinsic” activator fell immediately after exercise and then rose over the next 90 minutes. We suggest from these results that the two kinds of activator are more intimately related than was previously thought and that the disappearance of post-exercise “extrinsic” activator (formerly ascribed to inactivación by the liver) is due to conversion to the “intrinsic” form commonly assayed in “resting” plasma
Characteristics of Human Milk Plasminogen Activator (Galactokinase)
Authors: Okamoto U. , Horie N. , Yamamoto J. , Nagamatsu Y.
Published: May 2019

Whether milk plasminogen activator is identical with the other activators has been obscure due, to difficulty to obtain good starting materials. Recently, the authors found that human milk, when collected within a week after deliveries, showed remarkable activator activity almost constantly. From these samples, the activator called galactokinase (GK) tentatively was partially purified by chloroform treatment, salting out, gelfiltration and i0n exchange chromatography, and MW (gelfiltration) was ca. 86,000. GK obtained showed basic properties of plasminogen activator. In addition, some different properties were noticed when compared with UK and SK induced activator (SK Act). GK showed high selectivity to D-Phe-Pip-Arg-pNA (originally thrombin substrate), while UK strongly selected Pyroglu-Gly-Arg-pNA and SK Act, D- ValLeu-Lys-pNA. Thus, optimal pH, Km and Vmax of GK were determined. High activity of GK also found in bovine milk collected in early days after deliveries was followed by rapidly decreasing curve of GK activity as seen in human milk.
Demonstration of Complexes Between Factor IX And IGG Antibodies from Hemophilia B Patients
Authors: Örstavik K.H.
Published: May 2019

Plasma from three patients with hemophilia B- and an acquired inhibitor to factor IX (titres: 5 U/ml, 0.73 U/ml and 0.32 U/ml) were incubated with purified factor IX and submitted to crossed Immunoelectrophoresis against a rabbit antiserum to factor IX. A heterogenous precipitin arc was seen instead of the homogenous precipitin arc seen when a control plasma from a patient with hemophilia B- and no acquired inhibitor was used for the incubation. Human IgG antibodies were demonstrated in the cathodal part of the precipitin aros. This was achieved by incubation of the agarose gels, after the electrophoresis, with a peroxidase conjugated rabbit antiserum to human IgG. The IgG antibodies from the two patients with the highest inhibitor titre contained both kappa and lambda light chains, and were thus of a polyclonal nature. It is concluded that IgG antibodies from low-tltred as well as high-titred inhibitor plasmas form complexes with factor IX. The detection of IgG in complex with factor IX may be a sensitive technique for the detection of low-titred inhibitors to factor IX.
Interaction of Thrombocytin with Platelets and Plasma Clotting Factors
Authors: Kirby Edward P. , Niewiarowski Stefan , Stocker Kurt
Published: May 2019

Thrombocytin (Tcn), a thrombin-like enzyme from Bothops atrox marajoensis venom, has been purified to homogeneity by precipitation with sodium salicylate and chromatography on heparin-agarose. It is a single chain glycoprotein (mw = 35,000) which is a typical serine protease and is readily inhibited by DFP and soybean trypsin inhibitor. It is also inhibited by anti-thrombin III in the presence of heparin, and the formation of a covalent complex can be demonstrated. Ten aggregates platelets, induces the release reaction, and stimulates clot retraction in a manner similar to thrombin, but has only 0.13. as much fibrinogen-clotting activity as thrombin. Preincubation of platelets with prostaglandin E1, or with metabolic inhibitors inhibits aggregation by Ten. Like thrombin. Ten cleaves prothrombin to form Intermediate 1 and Fragment 1, and it activates Factor XIII by cleaving the a chain at approximately the same position as does thrombin. Ten can also degrade the α-chain of fibrinogen. It activates Factor VIII and Factor X, but these activities are very weak. In summary, Ten possesses many of the activities of thrombin, especially in regard to platelet aggregation and release, but has only a very slight fibrinogen-clotting activity. This may make it very useful for studying thrombin-mediated platelet activation in plasma-containing systems without the complications associated with fibrin generation.
Membrane Glycocalicin and Platelet Agglutination by Bovine Factor VIII Related Protein
Authors: Solum N.O. , Hagen I. , Stabæk T. , Filion-Myklebust C.
Published: May 2019

Membrane glycocalicin was extracted from human platelets in 3 M KCl and purified using affinity chromatography on WGA-Sepharose as the most efficient step. Antibodies to the purified material were raised in rabbits. Monospecific antiserum was obtained either directly or after adsorption with soluble fraction from platelets which had been frozen and thawn in ED TA-buff er and therefore contained very little glycocalicin. Such anti serum agglutinated human platelets, and in low amounts it inhibited agglutination by bovine factor VIII. This inhibition could be reversed by adsorption with platelet “ghosts” dependent on their glycocalicin content. Immunoelectrophoretic quantitation (Laurell) gave values of 47-67{U 9 of glycocalicin per 10 platelets corresponding to around 3% of tota plateLet prote in and accounting for most of the neuraminidase-sensitive plateletsialic acid. Whereas glycocalicin was quantitatively removed from whole platelets by 3M KCl, this procedure could remove only 39% of glycocalicin from glycocalicin-containing platelet ghosts prepared in the presence of EDTA. Such KCl treated ghosts could still agglutinate by bovine factor VIII whereas the KCl-treated whoie platelets were not agglutinated.
Increased Ristocetin Induced Binding of Factor VIII to Platelets in Von Willebrand’s Disease (vWd)
Authors: Ruggeri Z.M. , Pareti F.I. , Mannucci P.M. , Zimmerman T.S.
Published: May 2019

Initial reports of ristocetin-induced platelet aggregation (RIPA) demonstrated it to be decreased in some patients with vWd. We now report 20 patients (from five unrelated families) in whom RIP A was increased, apparently as the result of an increased ristocetin-induced binding of Factor VIIIrelated antigen (VIIIR:Ag) to platelets. All the patients had a life-long bleeding tendency, with prolonged bleeding time, and an abnormal two-dimensional crossed immunoelectrophoresis (2DCIE). Increased RIPA was demonstrated by measuring the minimum ristocetin concentration necessary to induce platelet aggregation. This was 0.42 mg/ml á 0.11 SD in the patients, and 0.91 á 0.097 SD in 17 normals (t = 13.83; P < 0.001). VIIIR:Ag binding to platelets occurred at ristocetin concentrations (0.4 mg/mI) which were ineffective in normals (who required >0.6 mg/mI). In contrast, the VIIIR:Ag of other patients with abnormal 2DCIE and markedly decreased RIP A did not bind to platelets at ristocetin concentrations as high as I mg/ml. It has been previously demonstrated that 30% to 60% of normal VIIIR:Ag binds to isolated human platelet membranes in the absence of ristocetin or any other agent, and that binding is restricted to the larger forms of VIIIR:Ag. However, VIIIR:Ag from the patients with increased RIPA, including two with normal ristocetin cofactor activity, showed decreased or undetectable binding as did all other patients with abnormal 2DCIE. This study suggests that ristocetin induced platelet Factor VIII interaction does not accurately reflect the “bleeding time factor” defect in vWd.
Activity Staining in Congenital Factor XIII Deficiency
Authors: Henriksson P. , Nilsson I.M. , Stenberg P.
Published: May 2019

Recently, Stenberg and Stenflo [Analytical Biochemistry in press 1979) introduced an activity staining procedure for transamidating enzymes (transglutaminases; endo-γ-glutamine: ε-lysine transferases! e.g. Factor XIII). This specific technique combines agarose gel electrophoresis with activity staining based on the enzyme catalyzed incorporation of mono-dansylthiacadaverine into casein. The method permits detection of plasma Factor XIII activity down to 1% of the normal adult standard. The procedure was applied to plasma and red blood cell lysate from two patients with tentative congenital plasma Factor XIII deficiency (based on clot solubility). No activity was found in platelet poor as well as in platelet rich plasma which confirmed the diagnosis. The red blood cell lysate displayed normal transglutaminase activity which means separate genetic determinations of the plasma and red blod cell trans-amidases. By Immunoelectrophoresis, the plasma Factor XIII b-subunit was quantitated to 43 and 44 % of the concentration in normal standard plasma.
Calcium-Dependent Cross-Linking Processes in Platelets
Authors: Cohen I. , Glaser T.
Published: May 2019

When platelet cytoplasmic Ca2+ is increased by the ionophore A 23187, there is the coincident appearance of a cross-linked polymer and the partial disappearance of five high molecular weight polypeptide bands (> 145,000). The glycoproteins show a partial disappearance of bands lb, IIb and IV and the total disappearance of hands la and Ilia. The disappearance of the protein bands, possibly contributing to the polymer formation, is prevented by histamine, aminoacetonltrile and cystamlne, which, as pseudodonor amines are known Inhibitors of factor XHIa-catalyzed cross-linking. 14C-histamine, at a tracer concentration, was incorporated into the polymer as well as into myosin, glycoproteins IIb and IIIa (α-actinln), actin and two unidentified low-molecular weight proteins. The polymer formed is also apparent in isolated membranes following the iono-phore-stimulated increase in intracellular Ca2+. These findings are unrelated to a proteolytic activity since the platelet Ca2+-dependent proteases are inhibited by leupep-tin. Ca2+-activation of a platelet cytosol transamidase would explain the data obtained. This platelet transamidase(s) may couple membrane proteins to cytoplasmic contra-tlle proteins. Thus, a new concept is proposed for the stabilization of platelet membranes and platelets as they form the hemostatic plug.
β Thromboglobulin, Platelet Factor 4 and Malondialdehyde Formation in Hyperlipid Emic Patients.
Authors: Zahavi J. , Betteridge D.J. , Jones N.A.G. , Galton D.J , Leyton J. , Kakkar V.V
Published: May 2019

Thromboembolic complications occur frequently in the hyperlipidemic patients. Platelets play an important role in the pathogenesis of such thrombi. Malondialdehyde (MDA) formation, which could indicate prostaglandin synthetase activity (PGSA), was measured in washed platelets stimulated by arachidonic acid in 34 hyperlipidemic (H) type II and V patients. Results were compared to age and sex matched healthy subjects and correlated to plasma β-thromboglobulin (βTG) and to 5 fractions of serum lipid levels. In 10 of the patients platelet factor 4 (PF4) level was also measured by radio-immunoassay. MDA (mean 10.92 nM 108 platelets) βTG (mean 115.7 ng/ml and PF4 (mean 65.4Mg/ml) were all
significantly elevated in the patients (p ⩽ 0.005) exceeding the upper range of control subject in 60% of them. There was a significant correlation between βTG and PF4 as well as between βTG and serum total triglyceride (TG) and VLDL-TG levels. No correlation was found between MDA and βTG or MDA and serum lipids. These results indicate that in-vivo platelet activation and “release reaction” and apparently PGSA are increased in hyperlipidemic patients, the former occurring not only through the prcstaglandin pathway. The abnormal in-vivo platelet activation, presumably induced by the high lipid levels - mostly VLDL-TG, may reflect a pre-thrombotic state.
Evidence that a Fat-Rich, Thrombogenic Diet Enhances Facfor X Activating Activity of Rat Platelets.
Authors: Tremoli E. , Colucci M. , Donati M.B. , Semeraro N.
Published: May 2019

The mechanism by which certain fat-rich diets are able to induce a marked thrombotic tendency in rats is uncertain. Several abnormalities of platelet function have been reported including increased platelet adhesiveness, enhanced platelet aggregability and increased platelet factor 3 activity. We have studied a recently described platelet coagulant activity (factor X activating activity) in rats fed a fat-rich, thrombogenic diet for 1, 2 and 7 weeks as compared to rats fed normal laboratory chow. Whatever the duration of the special feeding period, a highly significant shortening of the special clotting time, devised for measuring this activity, was observed. When platelet coagulant activity of individual “fat-fed“ rats was quantitated by reference to that of the respective randomly paired control animal, a 2-5 fold increase was found as early as after one week of dietary treatment. Partial thromboplastin time, thrombin time and soluble fibrin monomer complexes did not differ in control and treated animals. It seems that platelet coagulant activity, as measured in our test system, is one of the first laboratory parameters to be modified by fat-rich diets. These findings may be relevant to an understanding of the role of platelet coagulant activities other than platelet factor 3 in thrombotic phenomena. Supported by CNR (Italy).
Increased Plasma Fibrinogen and Platelet-Aggregates in Type II Hyperlipoproteinaemia
Authors: Third J.L.H.C. , Lowe G.D.O. , Drummond M.M. , Bremner W.F. , Lawrie T.D.V. , Prentice C.R.M.
Published: May 2019

Plasma-fibrinogen and circulating platelet-aggregates (method of Wu and Hoak1) were measured in 21 patients with Type II hyperlipoproteinaeima and 21 matched control subjects. Patients with hyperlipoproteinaemia had increased levels of fibrinogen (3.5 g/l ± SEM 0.2 vs. 2.5 g/l±0.1, p(0.01) and platelet-aggregates (platelet aggregate ratio 0.71 vs. 0.65, p(0.01). Young patients with hyperlipoproteinaemia had prematurely high fibrinogen levels, and the normal fibrinogen rise durina adult life was abolished. There were no significant correlations between fibrinogen, platelet-aggregates, and plasma lipids (cholesterol, cholesterol fractions, or triglyceride). High librinogen and platelet-aggregate levels may play a part in the development of the premature arterial disease associated with Type II hyperlipoproteinaemia, or may be markers of arterial injury.

1Wu, K.K., Hoak, J.C.Lancet, 1974, ii, 924.
The Possible Residual Initial Polymerisation Site in Fibrin Plasmic Fragment D-Dimer.
Authors: Purves L.R , Lindsey G.G.
Published: May 2019

The stable plasmic fragment of fibrin, D-dimer, again becomes susceptible to plasmin digestion if calcium ions are removed. Pragressive cleavage af the C-terminal end of the γ chain occurs releasing a peptide containing the cross link and any biologically attached (F XIII: transglutaminase) dansyl cadaverine. The cleaved peptide binds tightly to the parent malecule but further plasmin digestion can be inhibited by calcium ions. D-dimer has two and D monomer a single high affinity calcium-specific binding sites (Kd 1χ10-5M). The farmation of a noncavalently-linked D-dimer (NCDD) can be shown by the molecular size on malecular sieving and by the retention of molar volume by the dansyl-cadaverine labelled but cleaved D-dimer using fluorescence polarisation. Some partial dissociation occurs at 5°C but reassaciation occurs on return to 35° C. Antibodies to D-dimer have a 5-fold lesser affinity for fibrinogen and D-monomer with our without fragment E present but NCDD gradually loses affinity with extent of cleavage and in time. The interactions of D-dimer with the E domain on fibrinogen and NDSK are dependent on the intact calcium-stabilised γ chain. The inter-D interaction maintaining NCDD could be the residual initial polymerisation site of fibrinogen.
Existence of Common Antigenic Determinants in the Aα, Bβ and γ Chains of Some Mammalian Fibrinogens.
Authors: Cierniewski C.S.
Published: May 2019

Polypeptide chains Aα, Bβ and γ of porcine fibrinogen were isolated by preparative SDS polyacrylamide gel electrophoresis. Their purity was estimated by electrophoresis in polyacrylamide gel, amino acid composition and N-terminal amino acid analyses. Antisera to the pig polypeptide chains were produced in rabbits and they were employed in immunological comparative studies of porcine, bovine, human and duck fibrinogens. Antisera to the pig Aα chain showed in gel immunodiffusion and passive hemagglutination a strong cross-reaction with porcine, bovine and human fibrinogens. Antisera to the pig βB and γ chains cross-reacted only with porcine and bovine fibrinogens but they did not recognize human fibrinogen, The reaction of antiγ antisera was detectable only by passive hemagglutination test. In all cases antigenic similarity of the analyzed fibrinogens was mainly related to antigenic determinants of the Aα, Bβ and γ chains exposed on the intact fibrinogen molecule. None of analyzed antisera reacted with duck fibrinogen.
The Relevance of Blood Tests in the Management of Thromboembolism
Authors: Nossel Hymie L.
Published: May 2019

Fibrinopeptide A (FPA) levels reflect fibrin I formed by thrombin proteolysis of fibrinogen - a single biochemical event in fibrin formation and thrombosis. Studies in symptomatic patients suspected of venous thromboembolism (TE) demonstrated a good correlation in 40/45 patients between elevated FPA levels and the presence of thrombosis and/or pulmonary embolism indicated by venogram and/or lung scan and in 50/55 patients between normal FPA levels and the absence of TE. Within 15 minutes of intravenous heparin in 30 patients with documented TE, FPA levels had reached the normal range. In 3 of 17 patients studied over 10 days of anticoagulant therapy elevated FPA levels were associated with recurrence or extension of embolism. Thus, plasma FPA measurements may be useful in the diagnosis and management of venous TE. A second biochemical event - fragment X formation by plasmin cleavage of the Aα and N-terminal Bऔ chains of fibrinogen - can be monitored as thrombinincreasable fibrinopeptide B immunoreactivity. Initial studies suggest imbalance between thrombin and plasmin action favoring thrombin during the inception of thrombosis and favoring plasmin during resolution. The use of these tests is providing specific and quantitative information concerning biochemical reactions associated with clinical TE. In correlation with 125I-fibrinogen scanning and conventional diagnostic techniques, these tests will provide basic pathophysiological understanding which will facilitate the development and use of new techniques in diagnosis, prediction and management of venous TE.
Lipoproteins and Fibrinogen in Atherogenesis
Authors: Walton K.W.
Published: May 2019

Previous work from this and other laboratories has shown that material antigenically related to the low-density lipoproteins (LDL) and to fibrinogen is demonstrable in atherosclerotic plaques. When arterial intima is extracted electrophoretically, ‘bound’ and ‘labile’ fractions of these antigens are in each case demonstrable. In the case of the “bound’ fraction of fibrinogen-related antigen (FRA) it has not been clear as to whether the material is in the form of native fibrinogen, as fibrin monomer, or as fibrin. To examine this problem fresh-frozen sections of arteries containing FRA have been examined by immunohistological techniques using antisera specific for fibrinogen, fibrinopeptides, plasmin and fibronectin. The results obtained with these antisera will be compared with one another and with results obtained with antisera to the antigens of LDL.
Chemical and Physical Effects of Factor XIII on Fibrin
Authors: McDonagh Jan , McDonagh R.P.
Published: May 2019

The primary function of Factor XIIIa is to form ε(γ-glutamyl)-lysine crosslinks in fibrin. The result is the formation of covalent linkages between two γ chains and multiple α chains. This reaction has both biochemical and biophysical consequences. Once crosslinking has occurred, the ability of plasmin to lyse the fibrin gel into soluble components is greatly retarded. The inhibition of fibrinolysis in crosslinked fibrin appears to be more closely correlated with α chain crosslinking than with γ chain dimerization. There is no effect of plasmin per se on either the activation or inactivation of factor XIII. The crosslinking of fibrin also results in changes in the mechanical properties of the fibrin gel, so that the network becomes mechanically more elastic. The crosslinking of γ chains at their carboxy termini appears to have the function of cementing the protopolymer in place, while α polymer formation is highly correlated with increasing elasticity. This implies that the physiologically significant step of strengthening the clot is the formation of a polymer.