bluuebunny/crossref_metadata_2025 huggingface dataset:
Studies On PIVKA-VII
Authors: Mariani G , Avvisati G , Romoli D , Mazzucconi M G , Mandelli F
Published: July 2018

The factor VII related antigen (Pivka-VII) in 25 random patients under long term treatment with oral anticoagulants was studied. All the patients had Thrombotest levels well within the therapeutic range (3 to 11%) and had been receiving the drugs for at least 4 months. Factor VII activity (VII: C) mean levels were 8.7 u/dl(sem 0.78) and factor VII antigen (VII: Ag) mean levels were 30.7 u/dl(sem 1.99), (ratio Pivka/Activity 4.04). Barium chloride adsorbed the same mean amount of factor VII: C and factor VII: Ag (8.7 u/dl and 8.0 u/dl respectively). After exposure to cold in 4/25 subjects the “cold activation” phenomenon became evident, characterised by an increase of factor VII: C levels and immodified VII : Ag levels. In the remaining 21 patients, mean VII : C as well as VII: Ag levels did not show modifications (8.5 u/dl and 30.7 u/dl respectively). Moreover, three normal volunteers received three different oral anticoagulants ( Acenocoumarin, Coumarin and Ethyl-Biscomacetate). A sharp increase of factor VII:C levels was observed in all the three subjects (from 30 to 50%), a few hours after the beginning of the treatments, followed by consensual decrease of the two properties related to factor VII that reached the nadir after 48 hours. The vitamin K administration was followed by a rapid and consenual increase of the two parameters.
Generation Of Coagulation Factors By The Isolated Rat Liver Perfused With A Synthetic Blood Substitute
Authors: Owen C A , Bowie E J W
Published: July 2018

Measuring the release of small amounts of a clotting factor from an isolated perfused rat liver is difficult if the perfusate already contains some of the factor. Further, platelet-containing perfusates generate a coagulant activity that may invalidate clotting assays.

We have successfully employed a completely synthetic blood substitute for rat liver perfusions. The perfusate is “Fluosol-43” generously furnished by Alpha Therapeutic Corporation. The oxygen-carrying perfluorochemical is FC-43 (perfluorotributylamine) and the substitute for albumin is hydroxyethyl starch. Using the Brauer perfusion technique, we found that rat livers in 5 hours released an average of 2.3% of the normal plasma concentration of prothrombin, 8.4% factor V, 16.2% factor VII, 7.0% factor IX, 3.7% factor X, 28.3% factor XI and 12.3% factor XII. Antithrombin III and plasminogen were also generated.

Only minute amounts of factor VIII were released unless serum, cryoprecipitate or cryoprecipitate-free plasma was added; then the yield was 8.8% on average. The more “venom factor” (platelet aggregability with Bothrops alternata venom) added to the synthetic perfusate, the more factor VIII was released.
Platelet Function Changes In Acute Myocardial Infarction
Authors: Strano A , Raineri A , Novo S , Piraino G L , Mazzola A , Traina M , Davi G
Published: July 2018

Whether the thrombotic component of myocardial infarction is primary or secondary in a given patient, platelet function alterations can influence many mechanism from which depends if the thrombotic lesion grows or sends platelet emboli to the smaller myocardial vessels. Recently in some cases of infarction, coronary artery spasm has been demonstrated angiographically; thromboxanes, vasoconstrictive and platelet-aggregating substances, are released by platelets during myocardial ischemia. The local release of these substances may modify the myocardial cell viability and regional blood flow.

The aim of the present study was to investigate changes in platelet function in relation to the time interval after the beginning of the chest pain in 8 patients suffering from acu te myocardial infarction.

The following parameters were estimated: plasmatic PF4 and BTG levels, thromboxane B2 formation by platelets stimulated with thrombin, plasmatic levels of thromboxane B2 MDA formation by platelets stimulated with thrombin, platelet sensi tivity to exogenous prostacyclin; factor XIII activity was also determined.

The tests showed, in the first three days, an augmented re lease of platelet constituents together with a “platelet exhaustion”, demonstrated by a reduced formation of MDA and thromboxane B2; in the following days the platelets changed to a state of hyperactivity. The platelet sensitivity to pro stacyclin was reduced during the whole period after the onset of the acute myocardial infarction; this provides an additional mechanism involved in increased platelet aggregation.
Platelet Activation During Treadmill Exercise In Patients With Chronic Peripheral Arterial Disease
Authors: Baele G , Bogaerts H , Clement D L , Pannier R , Barbier F
Published: July 2018

β-Thromboglobulin (β-TG) and platelet factor 4 (PF-4) are specific platelet proteins released during in vivo platelet activation. An increase in PF-4 after exercise- induced myocardial ischemia was reported by Green, L.H. et al.. This observation prompted us to measure β-TG and PF-4 in patients with chronic occlusive arterial disease of the lower limbs and to look for increments during treadmill exercise.

β-TG was measured using the Amersham test kit and PF-4 using the radioimmunoassay kit of Abbott Diagnostics Division. Plasma levels in 28 normal individuals ranged for β-TG from 7 to 39 ng/ml with a mean value of 21.0 ng/ml, for PF-4 from 1 to 19 ng/ml with a mean value of 6.0 ng/ml.

β-TG and PF-4 were measured in 59 patients with chronic peripheral arterial disease before and 5 min. after treadmill exercising till occurrence of claudication. Plasma levels of β-TG before treadmill exercising ranged from 24 to 260 ng/ml with a mean of 77.9 ng/ml, PF-4 levels ranged from 2 to 240 ng/ml with a mean of 30.4 ng/ml. These levels were significantly higher than those measured in normal individuals.

After treadmill exercise β-TG levels showed a statistically significant increase to a mean value of 87.3 ng/ml but PF-4 did not rise significantly (mean value : 32.4 ng/ml). The supplementary increase of already elevated β-TG levels may be explained by enhanced in vivo platelet activation during treadmill exercising till occurrence of claudication. As the clearance of PF-4 from human plasma has been shown to be much faster than the clearance of β-TG increases in PF-4 levels may be more difficult to detect during dynamic explorations of the vascular system.
Correlation Of Platelet With Clinical Studies In Patients With Coronary Artery Disease Pre And Post Coronary Artery Surgery
Authors: Doyle D J , Chesterman C N , Cade J F , Morgan F J
Published: July 2018

80 patients with angina were studied preoperatively with measurements of 51Cr platelet survival (PS), plasma PF4 and βTG (RIA), platelet aggregates and platelet count. Clinical data was recorded prospectively. 58 patients were restudied a mean of 34 weeks postoperatively, 44 of whom entered a double blind randomized trial of sulphinpyrazone.

Mean preoperative PF4 and βTG were elevated above normal values (PF4 14 vs 8ng/ml, βTG 54 vs 32ng/ml) and the mean PS shortened (150 vs 174hrs). These tests returned towards normal postoperatively (PF4 10ng/ml, βTG 39ng/ml and PS 186hrs). This trend is partly due to a significant change in βTG, PF4 and PS in patients treated with sulphinpyrazone.

A significant negative correlation was detected between platelet count and angina grade (r = -0.30, p< 0.001), number of previous transmural myocardial infarctions (r = -0.36, p< 0.001) and angiographic grading of severity of coronary artery disease (r = -0.28, p<0.05). No other significant correlation of platelet studies with coronary artery disease (CAD) status was detected. There was however an association between elevated PF4 and βTG and shortened PS in the 4 postoperative patients with peripheral vascular disease (PVD) compared to those without PVD (mean PF4 21.8 vs 7.3ng/ml, βTG 98.1 vs 34.2ng/ml). In addition βTG was negatively correlated with postoperative (not preoperative)triglyceride concentrations (r = -0.73, p< 0.05) which were also correlated with PS (r = 0.72, p< 0.05).

Abnormalities in platelet parameters are present in patients with CAD but they do not appear to be sensitive indicators of CAD severity and may well be affected by other factors for example extent of atheroma elsewhere, serum lipids and preoperative stress.
Platelet Survival Time In Men With Post Myocardial Infarction And Normal Coronary Arteriogram
Authors: Casals F , Castillo R , Alfonso L , Vidal J L , Paré J C , Betriu A
Published: July 2018

Some patients fail to show any evidence of arte riographic coronary obstruction following an episode of myocardial infarction. This type of subjects would all present a shortened mean platelet survival.

Over a year’s period, platelet survival time (Chromium 51 labelling)was measured in 8 subjects who, one month after suffering an acute transmural infarction, were found to have a normal coronary arteriogram (NC). Survival times were determined between 3 and 12 months after the infarction. At this moment, all the patients were asymptomatic.

As control, and simultaneously, platelet survival time was evaluated at random in 8 patients who had had a myocardial infarction with arteriographic evidence of coronary obstruction (AC), and in 11 normal subjects (NS).

Platelet life span was measured by fitting the gamma fonction at 11 days of survival experimental points and the goodness of fitting surveyed by a graphic terminal (Tektroniks 4051). Since a failure of fitting with this system was found in 30% of cases, the mean weight method of computting platelet life span had to be employed.

The results of the mean platelet survival time in the three groups of individuals show no significant differences : NS 7,76 ± 0,89; NC 7,48 ±1,68 AC 7,64 ± 0,87 (days ± 1 SD). Only two patients with normal coronary arteriogram had values lower than the inferior range of normal subjects.

These results do not agree with others.
Abnormal Platelet Serotonin Uptake And Binding Sites In Myeloproliferative Disorders
Authors: Caranobe C , Sié P , Nouvel C , Laurent G , Pris J , Boneu B
Published: July 2018

We previously shown that the platelets of patients suffering from myeloproliferative disorders (MD) present not only a reduced capacity to store serotonin (5 HT) in their dense granules but also a dramatic reduction in the initial velocity (Vi) of 5 HT uptake ; this suggests that the abnormalities are not restricted to the dense granules but involve the transport mechanism accross the platelet membrane.

The present study concerns the 5 HT binding sites of MD platelets which present such a reduction of the Vi Max (Li- neweaver-Burke plot) of the 5 HT uptake. Binding assays were performed according to the method of Schik et al. (Biochem. Pharmacol. 1979, 28, 2667). Schatchard plot analysis of the binding data revealed two binding sites both in normals and patients : site A with a high affinity and a low capacity and site B with a low affinity and a high capacity.

Thus the abnormal 5 HT transport accross the plasmatic membrane is the consequence of a quantitative reduction of the 5 HT binding sites and not of a qualitative defect of these sites. Nevertheless, in spite of the reduction of the number of binding sites, 5 HT-induced platelet aggregation was found normal in these patients.
An Ex Vivo Test Of Platelet Function Correlates With Template Bleeding Times And Is Unaffected By Symptoms Of Patients With Thrombocytopenia
Authors: McGill M , Quiroga L E , Sacher R A
Published: July 2018

The template bleeding time is used extensively for measurements of in vivo platelet function in thrombocytopenic patients and for evaluations of platelet transfusions. Interpretation of results is complicated by technical difficulties of the test procedure and by patient symptoms including sepsis, fever, splenomegaly, alloimmunization and drug therapy. In an attempt to find additional tests which predict function of circulating platelets, ex vivo measurements of platelet reactivity to vessel wall subendothelium, and template bleeding time tests, were performed on normal individuals and a series of hospital patients. Platelet reactivity in controls was 7.2 ± 3.3 um (n=10) and bleeding times were 3.1 ± 0.3 min. Reactivity decreased to 2.6 ± 0.6 um and bleeding times increased to 7.6 ± 3.6 min. in aspirinated controls (n=12). Patients with thrombocytopenia and prolonged bleeding times (n=15) consistently gave reduced reactivity values. Thus, the direct, inverse relationship between in vivo function (bleeding time) and ex vivo function (platelet reactivity) support the idea that the laboratory test is a valid measurement of platelet capabilities for function in vivo. Unlike bleeding time tests, ex vivo measurements of reactivity can be performed apart from patient symptoms which complicate measurement of function in vivo and under controlled conditions.
Itanoxone, An Inhibitor Of Cyclo-Oxygenase, Acts Preferentially On Platelet Than On Vascular Enzyme
Authors: Villa S , DeLhon A , Palmier C , Livio M , Maynardier B , Lauressergues H , de Gaetano G
Published: July 2018

Several attaints have been made to dissociate the inhibitory effects of aspirin on platelet and vascular cells, but no definite results have been obtained. Other drugs, presumably acting on cyclo-oxygenase, are therefore being investigated for their relative inhibitory effect on platelet and vascular prostaglandin synthesis.

The present study was performed in male CD-COBS rats. Itanoxone [ (chloro-2' -diphenyl) -4-oxo-4 methylene 2-butyric acid ], a newly developed, hypolipidemic and hypouricemic compound with moderate anti-inflanmatory activity, showed a short lived, dose dependent (20-200 mg/kg, orally) apparently competitive inhibition of platelet MDA stimulated by either thrombin or arachidonic acid. Repeated doses did not result in any cumulative effect. At doses which completely blocked MDA production, itanoxone also inhibited thrombin-stimulated thromboxane B2 production in platelets but had no measurable effect on vascular prostacyclin generation measured both by a bioassay and a radioimmunoassay of its stable derivative 6-Keto-PGF1α . Pretreatment with itanoxone partially prevented therrhibitory effect of aspirin on both platelet and vascular prostaglandin synthesis. This suggests that itanoxone - like aspirin - acts at the level of cyclo-oxygenase but has much greater selectivity on the platelet enzyme.

This pharmacological activity is of great theoretical interest for potential use of this compound as an antithrombotic drug.
Chemiluminescence In Human Platelets
Authors: Würner P , Abtlg F
Published: July 2018

Chemiluminescence (CL), the formation of malcndialdehyde (MDA) and serotonin release were studied in suspensions of washed platelets in the presence of luminol at 37° C. CL was induced by concanavalin A, A23187, arachidonic acid, thrombin, N-ethylmaleimide (NEM) and thimerosal.

Concanavalin A− and the major part of A23187-induced CL was due to leukocytes contaminating the platelet samples. Leukocytes had no influence on CL by the other stimuli.

Arachidonic acid evoked two separate signals: the first was peaking at 7 s and decayed within 20 s, while the maximum of the second signal occurred 50 s and returned to the background 70-110 s after its addition. Thrombin induced also two signals, however, with low intensity and with a lag phase of 6 s prior to the onset of the first signal. Exposure of platelets to NEM (≥0.5 mM) lead to a single signal after a lag phase, depending on its concentration, between 20 and 50 s. Thimerosal-induced CL consisted also of a single signal peaking after 35-120 s.

ASA and indomethacin prevented CL, formation of MDA and release promoted by NEM or thimerosal. They interfered with MDA-generation and the first signal of CL induced by arachidonic acid or thrombin. ASA and indcmethacin enhanced and NEM (50 μM) suppressed the second siqnal of CL induced by arachidonic acid or thrombin.

It is concluded that CL promoted by NEM and thimerosal as well as the first signal induced by arachidonic acid or thrombin depends on prostaglandin synthesis. The second signals indicate arachidonic acid turnover by lipoxygenase. Sulfhydryl reagents such as NEM interfere with platelet lipoxygenase resulting in only one signal of CL. Measurement of CL offers a useful tool with the advantage to allow monitoring of the time course of prostaglandin synthesis and lipoxygenase pathway in intact platelets.
Immunochemical Studies Of Fibrinogen Products Produced During Thrombosis
Authors: Canfield R E
Published: July 2018

Immunochemical measurement of the products of fibrinogen proteolysis has provided a method to study the terminal events of coagulation that are initiated by thrombin as well as those events associated with the fibrinolytic actions of plasmin. Thrombin releases fibrinopeptide A (FPA) and later fibrinopeptide B (FPB). Immunologic techniques to measure FPA are now well established; determination of FPB is complicated by degradation of the peptide in plasma. Early plasmin cleavage occurs at the NH2-terminal end of the Bβ-chain of fibrinogen yielding Bβ 1-42. This fragment exhibits limited crossreactivity with antisera to FPB. The action of plasmin at this site on fibrin I may play an important role in determining whether thrombin release of FPA ultimately leads to thrombus formation in vivo. Other early plasmin cleavage products arise from the COOH-terminal half of the α-chain. Details concerning the application of these immunochemical measurements to an understanding of the role of thrombin and plasmin-mediated proteolysis of fibrinogen and fibrin will be discussed. In addition, immunochemical attempts to detect the presence of factor XIIIa-catalyzed crosslinks will also be described.
Crossed Immunoelectrophoresis In The Identification Of Platelet Membrane Glycoproteins And Complexes
Authors: Karpatkin S , Shulman S , Howard L , Sadanandan S
Published: July 2018

Human platelet membranes were solubilized in 1% Triton X-100 and subjected to crossed immunoelectrophoresis, employing a rabbit anti-piatelet membrane antibody. Ten different antigens were observed fairly consistently; one could be identified as albumin, the other as fibrinogen. Surface antigens were determined by antibody adsorbtion experiments, and cell surface labeling with 125I-lactoperoxidase. Four surface antigens reacted with concanavalin A, when this was employed as an intermediate spacer gel. A major surface antigen, 10, was present on all preparations and was inversely related to antigens 13 and 18, which moved more cathodally. Membranes from preparations with full 10 antigen peaks had absent or diminished 13 and 18 antigen peaks, whereas preparations with absent to incomplete cathodal curves had increased 13 and 18 antigen peaks. Digestion of intact washed platelets with α chymotrypsin resulted in a decrease in the 10 antigen peak and an increase in 13 and 18, suggesting a structural relationship. Extraction of platelet membranes in EDTA or EGTA resulted in the disappearance of 10 and appearance of 13 and 18 (which react with concanavalin A) and 15 and 16 (which do not). Splitting of the major antigen into 13, 18, 15 and 16 could be prevented by addition of excess Ca++ relative to EGTA (but not by excess Mg++). Similar results were obtained in the absence of chelating agents following dialysis at 60-200 hrs against ‘Ca++-free’ buffer. Five patients with Glanzmann’s thrombasthenia have been studied. All k components of the major membrane antigen are missing. We conclude that the major antigen, 10, is composed of 2 glycoproteins, 13 and 18, and 2 other surface proteins, 15 and 16, which are held together by Ca++. It is conceivable that patients with Glanzmann’s thrombasthenia are lacking a membrane receptor for Ca++ or the platelet membrane (glyco)protein which anchors these A components.
Methods To Evaluate Endothelial Injury
Authors: Bondjers G , Hansson G , Bylock A
Published: July 2018

During the last decade the potential significance of endothelial injury for the development of atherosclerosis has been subject to an increasing interest. Thus, the effects on the arterial tissue of different types of experimental injury have been thoroughly described. Nevertheless, the hypothesis of a relationship between endothelial injury and atherosclerosis seems difficult to accept unless the effects of any of the well established risk factors for atherosclerosis can be attributed to modulations of endothelial integrity. To evaluate the presence of endothelial injury with light microscopy we have used dye exclusion tests and immunohistochemistry to detect intracellular IgG. The distribution of injured endothelium with these methods appears to be the same, and both methods are well suited for quantitative purposes. As a complement to these methods, 3H-thymidine incorporation and autoradiography may be used as an indirect measure of endothelial injury. On the ultrastructural level, scanning electron microscopy or transmission electron microscopy possibly combined with immunohistochemical techniques may be used. In our hands, however, none of these methods appear very well suited for quantitive assessment of endothelial injury. Nevertheless, dye exclusion tests, immunofluorescence, thymidine incorporation, scanning and transmission electron microscopy together constitute a reasonably complete battery of methods to evaluate the possible relationship between different risk factors and endothelial injury.

Using this battery of methods we have observed an increased prevalence of endothelial injury in the vicinity of branching points as well as in the periphery of atherosclerotic lesions induced with hypercholesterolemia. Both these locations show a relationship to the formation and growth of experimental atherosclerotic lesions.
Role Of Endothelial Integrity In Atherosclerosis
Authors: Schwartz Stephen M , Gajdusek Corinne M , Reidy M A
Published: July 2018

The current view of the pathogenesis of atherosclerotic lesions stresses loss of endothelial integrity as a major event leading to the three major events in lesion formation. ..accumulation of smooth muscle cells, accumulation of lipid and accumulation of platelets. Recent advances in endothelial biology, however, suggest that frank denudation may not exist. Our kinetic studies of small wounds to endothelium both in vivo and in vitro demonstrate the ability of the endothelium to rapidly replace desquamating cells. Furthermore, these small areas of mechanical denudation do not stimulate smooth muscle proliferation. Finally, careful studies of hyperlipemic and hypertensive animals have failed to demonstrate any actual denudation.

The possibility remains that denudation occurs repeatedly, with extremely brief periods of subendothelial exposure. Thus lesion formation could represent the cumulative record of multiple, tiny incidents. This hypothesis is, however, difficult to test. Less dramatic forms of injury which may be of interest include alterations of endothelial cell functions in replicating cells as well as alterations in function which may occur independent of any cell loss. This last category receives new emphasis today because of the rapid increase in the number of functions identified in endothelial cells. These include the apparent ability of endothelial cells to synthesize a growth factor which can stimulate smooth muscle proliferation in the absence of the platelet derived growth factor.

This work was supported by NIH grants: HL-03174. HL-18645, HLC-7312 and a grant from the R.J. Reynolds Industries, Inc. Dr. Schwartz is an Established Investigator of the American Heart Association.
Mechanisms Of Viral-Induced Endothelial Injury
Authors: Friedman H M
Published: July 2018

We examined the role of common human viruses as inducors of injury to human umbilical vein or bovine-thoracic aorta endothelial cells grown In vitro. Indicators of infection included endothelial cytopathology, viral replication and viral antigen detection by immunofluorescence. Herpes simplex, adeno, measles and parainfluenza viruses infected both human venous and bovine aorta cells. Mumps, polio, and echo 9 viruses grew only in human venous cells, while coxsackie B4 replicated only in bovine aorta endothelium. Several viruses (CMV, influenza A, RSV) infected neither type of endothelial cell.

We examined one virus, herpes simplex type 1 (HSV) in some detail and showed that HSV can establish a persistent infection of bovine aorta endothelial cells in vitro. When HSV was inoculated at a multiplicity of infection (MOI) of 0.1, the amount of infections virus produced by the culture fluctuated over a 3 month period. By the third week after inoculation the endothelial monolayer had recovered and no infectious virus was detected for an additional 2 months. However, by the third month, typical herpetic changes reappeared, 103 plaque forming units of intracellular and supernatant virus was detected, and by infectious cell center assays, approximately 1% of cells produced virus.

In separate experiments, human umbilical vein endothelial cells were inoculated with HSV (MOI of 1.0) and examined for induction of surface receptors for the Fc portion of IgG and for C3b. Using 51Cr-labeled sheep erythrocytes coated with IgG or with IgM and purified complement components (C1, C4, C2, C3) an Fc and C3b receptor were detected on the infected cells. These receptors were distinct from one another as demonstrated by blocking experiments using purified Fc fragments, and consisted, at least in part, of viral antigens since antibody to HSV blocked function of both receptors.

These studies demonstrate that viruses can produce acute or persistent infection of endothelial cells and can induce receptors for IgG and complement on the cell’s surface.
Triais And Tribulations In The Prevention Of Myocardial Infarction
Authors: Mitchell J R A
Published: July 2018

“There is no present or future - only the past happening over and over again, now” (Eugene O’Neill).

When coronary thrombosis was thought to be clotting in the wrong place, it was assumed that coumarin anticoagulants would modify it, and would prevent myocardial infarction and death. Early trials did not reoognize that sudden deaths and deaths from pump failure and dysrhythmia soon after infarction were not thrombotic events. Bad design, emotive opposition and therapeutic greed led to disillusionment with anticoagulation.

The next belief was that thrombosis was haemostasis in the wrong place, so the recognition that dipyridamole, sulphinpyrazone and aspirin modified platelet behaviour led to the hope that these agents would be anti-thrombotic. Our present inability to confirm or deny this shows that our inept handling of anticoagulants has not enabled us to avoid similar problems with these first-generation platelet modifying agents and the problems posed by ART, AMIS and PARIS will be discussed.

finally, the newest theoretical concept is that thrombosis is due to TXA2/PGI2 imbalance, so mini-dose aspirin is being suggested as an antithrombotic while we await the arrival of thromboxane synthetase inhibitors, stable PGI, analogues or TXA2 receptor blockers which are being hailed as antithrombotic agents for the future. Unless we show that We have learnt how to test for coronary anti-thrombotic activity we shall be as confused about these new agents in 20 years as we are about the old ones.

Meanwhile, the anticoagulant flame has been kept alight in Holland and the results of their trial in patients over 60 and many years after an initial infarction will be described. If the original idea that thrombosis was clotting in the wrong place is correct shall we soon be saying ‘Come back anticoagulants, all may yet be forgiven.?
Secondary Prevention In Coronary Heart Disease - Beta- Blockade, Surgery, And Control Of Primary Risk Factors
Authors: Julian D G
Published: July 2018

Numerous factors are involved in the progression of coronary atherosclerosis and the development of its complications and it is probable that different sub-sets of patients will respond best to different therapies. Combination therapies are probably most appropriate for some patients.

Randomised studies have been completed with a number of beta-blockers after infarction including alprenolol, practolol, propranolol, oxprenolol and timolol. Although it is questionable if benefits are conferred on the total population treated, it seems probable that one or more of these drugs diminish the risk of death (especially sudden death) in patients who start the drug in the convalescent phase following infarction. They probably do not prolong in those with a good prognosis nor in those whose left ventricular function is severely compromised.

The European Coronary Artery Surgery Trial demonstrated, in a randomised study of a highly significant improvement in prognosis in men under 65 with angina who had undergone coronary artery bypass grafting for three vessel coronary disease. The prognostic value of surgery after myocardial infarction remains to be established.

No satisfactory randomised controlled trials have established the prognostic value of controlling hyperlipidaemia or hypertension in patients with coronary heart disease, nor have they proven the effectiveness of exercise programmes or the stopping of smoking. However, the cessation of smoking after myocardial infarction is almost certainly beneficial.
Monoclonal Antibodies As Probes Of Factor VIII/Von Willebrand Factor
Authors: Meyer D , Zimmerman T S , Edgington T S
Published: July 2018

Monoclonal antibodies are essential for the isolation and specific assay of the various components of Factor VIII/von Willebrand Factor (F.VIII/vWF) and for the immunochemical analysis of the surface epitopes of this protein. In this study, we have produced and characterized a panel of thirty-eight antibody synthesizing hybridomas. Mice were hyperimmunized with human F.VIII/vWF and three days after a 10-20 jig booster dose, spleen cells were harvested and fused with P3X63Ag8 or SP2/0 myeloma cells. All antibodies reacted by solid phase ELISA with purified F.VIII/ vWF and normal cryoprecipitate and did not bind to cryoprecipitate from severe vWD, fibrinogen, or fibronectin. Antibody titers (20 μl) varied from lO2 to 106. Seven antibody producing hybridomas derived from P3X63Ag8 myeloma cells were cloned, recloned, and were stable in ascites growth. Two of these partially blocked VIIIR:RCo while the mixture of the seven exhibited significant inhibition, suggesting a cooperative effect in steric hindrance or induction of allosteric effects on the VIIIR:RCo function of the molecule. Three out of thirty-one hybridomas derived from SP2/0 myeloma cells blocked Factor VIII activity. A spatial map of the epitopes at the surface of F.VIII/vWF was derived from competitive displacement studies of the monoclonal antibodies on immobilized F.VIII/vWF. Three types of results were observed : no displacement, indicating independence of two epitopes; partial displacement, indicating steric proximity; and complete displacement, indicating epitope identity or extremely close proximity.
Factor VIII, 1981
Authors: Hoyer L W
Published: July 2018

During the past decade there has been intense research interest in hemophilia and von Willebrand’s disease, and in the properties of factor VIII. These studies have provided an evolving understanding of factor VIII structure and function. A central issue during this period has been the nature of plasma factor VIII. The initial concept of a single protein with two distinct biologic functions has proved to be less satisfactory as an explanation for experimental data than the alternative hypothesis that plasma factor VIII is a complex of two distinct proteins. Recent studies indicate that factor VIII procoagulant protein (antihemophilic factor) and factor VUI-related protein (von Willebrand factor) are under separate genetic control, have distinct biochemical and immunologic properties, and have unique and essential physiologic functions. While the nature of their interaction and the details of their biochemical structures remain to be determined, the information now available permits a preliminary understanding of the molecular defects in hemophilia and von Willebrand’s disease. This paper will discuss what is known about the components of the factor VIII complex and it will emphasize the limits of our understanding of how these proteins facilitate plasma coagulation and primary hemostasis.
Interactions Of Triphenyl-Methane Compounds With Fibrinogen And Fibrin Monomers
Authors: Ryckewaert J-J , Doolittle R F
Published: July 2018

A synthetic compound, aurin tricarboxylic acid (ATA), in the concentration range of 10-100 nanomoles/ml exhibits the property of significantly shortening the thrombin clotting time of fibrinogen. Upon binding to fibrinogen or fibrin monomers, obtained either by the action of thrombin or reptil ase on fibrinogen, ATA possesses the ability to accelerate the polymerization formation of fibrin. Polymerization studies of fibrin monomers at different ionic strengths show that ATA greatly enhances the lateral aggregation of fibrin polymers even at high ionic strength. Calcium ions do not influence the effects of ATA. Spectroscopic methods have been used to study and locate the binding sites in the different parts of the fibrinogen molecule. Compounds with structures that are closely related to that of ATA do not affect the polymerization process, with the exception of the parent compound aurin, which has a slight effect. ATA does not influence the binding ability of fibrinogen for the synthetic peptides: Gly-Pro-Arg and Gly-His-Arg, as was shown by equilibrium dialysis experiments with radioactive- labelled peptides. Nevertheless, in the presence of Gly-Pro-Arg or Gly-Pro-Arg-Pro, which are known to interfere with the polymerization of fibrin monomer, ATA becomes a potent inhibitor of the overall process of fibrin formation. This duality of behavior has permitted us to study the different steps of longitudinal and lateral association of fibrin monomers.
The Mechanism Of Inhibition Of Fibrin Assembly By Fragment D
Authors: Williams J , Hantgan R , Knoll D , McDonagh J , Hermans J
Published: July 2018

Measurements of clot rigidity and fiber thickness indicate that fragment D is a potent inhibitor of fibrin assembly. At physiological ionic strength, D concentrations in excess of 2 moles D/mole fibrinogen lead to a large decrease in clot rigidity and fiber thickness. Above 14 moles D/mole fibrinogen, gelation is inhibited. The molecular weight and radius of gyration of the fibers, determined by light scattering, confirm that short oligomers result, composed of 3 fibrin monomer molecules at 120 moles D/mole fibrinogen and 9 monomers at 14.4 moles D/mole fibrinogen. If D is added to a solution of long protofibrils, no inhibition is observed. Apparently the polymerization of monomers to protofibrils is blocked by D, but not lateral association.

Inhibition of protofibril growth was studied in 0.5 M NaCl, pH 7.4, where lateral association is limited by the ionic strength. Stopped-flow light scattering data show a small decrease in the initial rate of polymerization and a slightly prolonged t½ ; the final intensity is less than that for a solution of long protofibrils. This result suggests that fragment D binds to the growing protofibrils with a small effect on the initial polymerization rate, but exerts its inhibitory effect by limiting the later stages of protofibril growth. Measurements of the length of the inhibited protofibrils, calculated from diffusion coefficients obtained by dynamic light scattering, confirm that polymers containing as few as 15 monomers have been obtained. Negative stain electron microscopy also shows a clear limitation of polymer growth under these conditions.

Fragment D interferes with fibrin formation by directly blocking the first assembly step: bimolecular polymerization of activated fibrin monomer molecules to form protofibrils. Fragment D apparently occupies a site normally occupied by a fibrin monomer molecule, thus forming a dead-end complex which cannot undergo further assembly.
Binding Of Plasminogen And Vascular Plasminogen Activator To Fibrin And The Fibrin Alpha-Chain
Authors: Lloyd D A , Cederholm-William S A , Sharp A A
Published: July 2018

Investigation of the mechanism of plasminogen activation in the presence of fibrin has shown that fibrin acts as a regulatory and catalytic surface for both the reaction between activator and fibrin and between activator and plasminogen. Plasminogen (mol wt 84,000) and vascular activator form stable complexes with fibrin. We have carried out studies to determine the location of these binding sites in the fibrin structure.

METHODS: Human plasminogen (mol wt 84,000) was iodinated with 125-I and vascular plasminogen activator was partially purified from venous occlusion plasma by chromatography on lysine-Sepharose. Sepharose coupled fibrinogen, fibrin monomer, fragments E and D and isolated fibrin alpha chain were prepared and the interactions between plasminogen and vascular activator studied.

RESULTS: Plasminogen (mol wt 84,000), plasmin and vascular activator each showed affinity for Sepharosebound fibrin monomer and fibrin alpha chain. In addition the affinity of the vascular activator for fibrinogen and fragment E was increased by treatment with thrombin.

CONCLUSION: The alpha chain of fibrin brings together plasminogen activator and plasminogen in such a way as to produce plasminogen activation resulting in the early degradation steps of fibrin.
Monoclonal Antibodies Against Factor Viii Procoagulant Activity
Authors: Sultan Y , Sola B , Avner Ph , Maisonneuve P , Jeanneau C
Published: July 2018

Monoclonal antibodies against the factor VIII/Von Willebrand molecule were obtained using the hybridoma technique. Splenocytes of mice immunized with a commercial preparation of high purity antihemophilic factor were fiised with a non secreting myeloma cell line SP 2/0. Hybrid cells were cloned twice by limiting dilution. Among 8 antibodies reacting with purified F.VIII/VW factor in a solid phase radio immunoassay, two of them showed an inhibitory activity against the procoagulant activity of the antihemophilic factor. This inhibitory activity was not detected in culture supernatants. Presumably in corelation with subtances of the culture medium interfering with the coagulation process and masking the neutralizing activity. However the anti VIIIc activity was demonstrated in the ascitic fluid obtained from mice injected with hybridoma cells.

The specific anti VIIIc neutralizing activity of F4 115 monoclonal antibody is dependent on Ig concentration, time of incubation, temperature and Ph. In contrast with most of anti VIIIc inhibitors occuring in multitransfured hemophiliac, neutralization was more rapid and highier at 4° than 37°C. This neutralizing activity is absorbed by normal plasma or purified F.VIII preparation, however it is not absorbed by plasmas of a limited number of severe hemophilic and severe Von Willebrand patients. The study of a larger patients population is in progress.

The second antibody F4 415 showed different inhibitory properties. Inhibition of the procoagulant activity is independent of incubation time butdependent on temperature and similarly more active at 4°C.

These monoclonal antibodies provide new tools to investigate the procoagulant piece of the F.VIII molecule
A Simplified Immunoradioactive Assay For Human Factor VIII Coagulation Antigen
Authors: Thomas K B , Howard M A , Koutts J , Firkin B G
Published: July 2018

Normal human factor VIII (FVIII) is a high molecular weight glycoprotein expressing two measurable biological activities, FVIII procoagulant activity (VIII:C) and von Willebrand factor activity (vWf). The corresponding antigenic moieties are referred to respectively as FVIII coagulation antigen (VIII:CAg) and FVIII related antigen (VIII: RAg). Since VIII:C is extremely labile measurement of VIII: CAg is of importance for pre-natal diagnosis of haemophilia where VIII:C is reduced and von Willebrand’s disease (vWd) where all measurable parameters of FVIII are reduced.

Current immunoradiometric assays for VIII:CAg depend on both the availability of high titre human antibodies directed against VIII:C and the possibility of preparing highly specific 125I-IgG from human antiserum

We have developed a solid phase immunoradiometric assay for VIII:CAg. FVIII in a test sample binds to human anti- VIII:C IgG immobilised on the surface of a polystyrene tube. The bound FVIII is detected using rabbit 125I-anti human FVIII-IgG. The standard curve was linear for dilutions of normal pooled plasma (NPP) of 1/5 to 1/500. The lower limit of the assay was 0.002U/ml of NPP. Using this assay the concentration of VIII:CAg correlated well with the VIII:C levels in the plasma from normal individuals (r=0.84, n=15). Homozygote vWd patients had undetectable levels of VIII:CAg, VIII:C, VIII:RAg or vWf activities. Patients with haemophilia A with less than O.OlU/ml VIII:C could be divided into two groups on the basis of the VIII:CAg assay, CRM and CRMf. Markedly reduced levels of VIII:CAg were detected in serum.

The advantage of this assay over the previously available assays for VIII:CAg are that smaller volumes of high titre human anti VIII IgG are required and the time consuming step of preparing 125I-human anti VIII-IgG is eliminated. The rabbit 125I-IgG specific for human VIII is the same as used in routine immunoradiometric assays for FVIII:RAg
Detection Of Antihaemophilic Factor By Immunoradiometric Assays In Lymph Node And Lung Extracts
Authors: Exner T , Joshua D E , Rickard K A , Kronenberg H
Published: July 2018

Although there have been many investigations into the site of synthesis of antihaemophilic factor, the organs and cells involved are still not clearly known and a multiorgan synthetic pathway is suspected. Extraction experiments have previously not given satisfactory results because of the presence of interfering procoagulants and the lability of factor VIII coagulant activity. Immunoradiometric assays (IRMA) for factor VIII coagulant antigen (VIII CAG) detect antigens closely related to the active coagulant site of factor VIII and are less affected by extracting media. We applied an IRMA to the detection of VIII CAG in various tissue extracts. Samples of fresh normal tissue taken at autopsy were extracted and the centrifuged supernatants tested for factor VIII CAG and factor VIII antigen by radioimmunoassays using homologous and heterologous antisera respectively. Whereas factor VIII antigen was extracted readily with isotonic saline, factor VIII CAG required 1.5M sodium chloride for optimal extraction. The possibility of VIII CAG within tissues being degraded before extraction and decay in the extracts themselves was investigated with aging and proteolytic inhibitor studies. Surprisingly, higher levels of factor VIII CAG were found in some lymph nodes and lung in comparison to liver and spleen which have been more usually considered to play some part in antihaemophilic factor biosynthesis. Factor VIII CAG in all tissue extracts was reduced by the addition of a trace of a second antibody to antihaemophilic factor different to that utilized in the IRMA.
Correlation Of Haematological Parameters In Deep Vein Thrombosis
Authors: Donaldson D R , Crow M J , Davies G A , Rajah S M , Watson D A
Published: July 2018

We have evaluated the response of certain haema to logical parameters in patients undergoing elective thoracotomy for pulmonary and oesophageal disease. These patients are part of a continuing DVT study in which prophylaxis is randomised to either i) Heparin 5000 iu 8 hourly s.c. (Group A) or ii) Heparin 2500 iu + Dihydroergotamine (DHE) 0.5 mg 8 hourly s.c. (Group B). Prophylaxis was commenced at the time of premedication and continued for 10 days. DVT was diagnosed by I125 Fibrinogen uptake test, with confirmatory venography if positive. The haema to logical parameters investigated were Antithrombin III (AT III), Euglobulin Clot Lysis Time (ECLT), plasma $-Thromboglobulin (β-Tg), platelet Factor 4 (PF4) and Heparin level. All investigations were performed pre-op. and day 6 post-op. (AT III and ECLT were also measured on day 3).

A total of 50 patients have completed the study to date with 25 patients in each treatment group. Eleven patients have developed DVT - 5 in Group A and 6 in Group B. There was no significant difference between the pre-op. values in either group. The post-op. AT III values of patients developing DVT were lower than the two treatment groups. ECLT was significantly prolonged in the two treatment groups post-op. compared to the pre-op. values (p < 0.05 (A) and p < 0.01 (B)), but was not prolonged post-op. in the patients who developed DVT. Plasma β-Tg and PF4 were raised pre-op. with further increases post-op. in all groups. Plasma Heparin concentrations in Group B (0.122 iu/ml) were lower than for Group A (0.182 iu/ml) and the DVT group (0.176 iu/ml).

These findings indicate that none of the haematological investigations were able to delineate ‘at risk’ patients. However the development of DVT was significantly correlated (p < 0.05) with the post-op. AT III and ECLT. The use of DHE in the dose cited above did not apparently inhibit the platelet release reaction
Subcutaneous Versus Intravenous Administration Of Heparin In The Treatment Of Acute Venous Thrombosis
Authors: Johnsson H , Andersson G , Holmgren H , Nilsson E , Ljungberg B , Wilhelmsson S , Zetterquist S
Published: July 2018

Ninety-five patients with acute deep venous thrombosis (DVT) in the leg were randomly allocated to either two daily subcutaneous injections (S.C.) or to continuous intravenous infusion (I.V.) of heparin for at least five days. Before entering the trial the existence of a thrombus was confirmed with phlebography and in about two thirds pletysmography and thermography (DevethermR, infrared temperature scanning) was performed at well. Immediately after heparin treatment phlebography was repeated in one third of the patients and in the others a new pletysmography and thermography was performed. The non-invasive procedures are then repeated again after 1, 3, 6 and 12 months (and once a year). In most patients the thrombi were localized in the popliteal-femoral veins and all patients had local symptoms. The amount of heparin required to maintain a prolongation of the APT-time 1 1/2 to 3 1/2 times was the same in both groups and the daily dosages varied between 25.000 and 50.000 units. In two patients, one of whom on salicylates the S.C. administration was stopped due to local haematoma. Major bleeding complications occurred in totally 4 patients, two on heparin I.V. By phlebography a partial thrombolysis was registered in about 25% of the patients after heparin treatment independent of administration route, and in no patient the thrombus progressed in size. By pletysmography and thermography an improvement was demonstrated in about one third immediately after heparin treatment and these improvements were not less pronounced in the S.C. group. Only one major, non-fatal pulmonary embolus (heparin I.V.) occurred.

Thus, S.C. administration of heparin seems to be as efficient and safe as administration of heparin I.V. in the treatment of patients with acute DVT.