bluuebunny/crossref_metadata_2025_split huggingface dataset:
DEVELOPMENT OF A SPECIFIC RADIODMMUNOASSAY FOR DETERMINATION OF PEPTIDES DERIVED FROM HUMAN LEUKOCYTE ELASTASE DEGRADATION OF HUMAN FIBRIN (OGEN)
Authors: Wallin R , Saldeen T 
Published: August 2018

This paper describes a RIA for determination of the vasoactive peptide BB 30-43 and related peptides derived from leukocyte elastase degradaticn of human fibrin(ogen). The peptide was synthesized and could easily be labelled with 125I.Rabbits were iirmunized with BB 30-43 conjugated to bovine albumin. The antibody was found to bind about 5C% of the tracer in absence of BB 30-43 in a ˜1/800 diluticn. The RIA can detect peptide concentrations between 50 - 25000 pmol/L. The crossreaction with fibrinogen is very low (<0.001%) and with plas-min derived fibrin(ogen) peptides Bβ 1-42 and BB 15-42 also low (<0.2%). Plasma samples can be analyzed without any pretreatment. In an in vitro study fibrin and fibrincgen was degraded with plasmin or leukocyte elastase. Plasmin degradaticn of fibrin and fibrincgen did not release peptides which cross-reacted with our antibody, whereas leukocyte elastase degradation released peptides from both fibrin and fibrinogen which crossreact whith the antibody.The imnunolqgical activity was not changed after degrading peptide Bβ 30-43 with a) trypsin, b) plasmin, c) batraxobin, d) thrombin, e) elastase, at +37°,1 h, in a molar ratio of 1:100. Even degradaticn by elastase (1:3.5) +37°, 1 h, did not destroy the iirmunological activity.The imriunolcgical stability of peptide B< 30-43 in EDTA-plasma (+37°) seems to be very good. In citrated and heparinized plasma the activity of this peptide seems to vanish quite fast. In spite of these results we have detected high levels of iirmunolcgical activitiy in citrated or heparinized patient plasma. The molecular distribution of the peptides detected in plasma by our RIA corresponded to a fragnent containing about 25 amino acids. This fragnent seemes to be rather stable in plasma. When this fragnent was degraded with elastase in vitro a peptide with a molecular size resembling BB 30-43 was obtained. Over 300 patient samples have been studied. About 20 per cent were positive and the highest levels were found in patients with ARDS, septicaemia, severe renal failure, pulmonary embolism, pneumonia and pulmonary congestion.
ASSOCIATION OF VINCULIN TO THE PLATELET CYTOSKELET0N DURING RELEASE REACTION
Authors: Horvåth A , Asyee G M , Sturk A , Muszbek L 
Published: August 2018

Vinculin is an Mr 130 kD protein, which in various cell types is located at the membrane attachment site of microfilaments and has been implicated in membrane-cytoskeleton interaction. Though we have previously verified its presence in platelets and showed that in resting platelets it is localized submembra-neously and around the ±-granules, its relation to the cytos-keleton is still to be elucidated. It has been also revealed by biochemical studies that in resting bovine gel filtered platelets vinculin is not cytoskeletal component, however about 50% of the total vinculin content became incorporated into the cytoskeleton during thrombin activation. To establish if its association to the cytoskeleton occured during pseudopode formation, aggregation or release reaction, cytoskeletons were prepared from platelets activated in various conditions and the time course of vinculin incorporation was analyzed by immuno-blotting. When pseudopode formation was inhibited by cytochalasin B pretreatment but neither aggregation nor release reaction induced by thrombin was prevented, the amount of vinculin in the Triton insoluble residue even increased. Phorbol myristate acetate (PMA), which induces pseudopode but not contractile gel formation, is a strong stimulus for aggregation, but induces only a slight release of the granule content. In parallel, only a low amount of vinculin was recovered in the cytoskeletal fraction following PMA induced aggregation. In contrast, when release reaction elicited by thrombin occured in the absence of aggregation the association of vinculin did not diminished.The results clearly demonstrate, that platelet shape change and pseudopode formation are not prerequisites of the incorporation of vinculin into the cytoskeleton, which occurs parallel to the release reaction.
HUMAN PLATELET GLYCOPROTEIN Ic-IIa IS AN ACTIVATION-INDEPENDENT FIBRONECTIN RECEPTOR
Authors: Piotrowicz Randolph S , Yamada Kenneth M , Kunicki Kunicki J 
Published: August 2018

Human platelets express the membrane glycoprotein (GP) heterodimer GPIIb-IIIa, which functions as an activation-dependent fibronectin (Fn) receptor. We have immunopurified the components of an activation-independent Fn receptor (FR) from human platelets employing a well-characterized rabbit polyclonal antibody raised against the beta chain of the chicken embryo fibroblast (CEF) FR (anti-band 3). This antibody crossreacts with antigen(s) expressed on both chicken thrombocytes and human platelets and inhibits the binding of both normal and thrombasthenic platelets (lacking GPIIb-IIIa) to Fn-coated surfaces in the absence of platelet activation.A monoclonal antibody directed against GPIIb-IIIa (AP2) partially inhibits the adhesion of normal platelets to Fn, but the combination of AP2 and anti-band 3 results in a level of inhibition greater than that obtained with either antibody alone. Thus, the presence of the FR alone is sufficient for the observed normal to enhanced binding of thrombasthenic platelets to Fn, whereas adhesion of normal platelets involves the synergistic action of the FR and GPIIb-IIIa. The adhesion of platelets to Fn mediated by the FR is inhibited by the tetrapeptide RGDS.Immunopurified FR appears to be a complex of two proteins: an alpha chain with an apparent molecular weight of 155/130 KD (nonreduced/reduced) and a beta chain with an apparent molecular weight of 125/147 KD. The alpha chain is composed of two subunits, dissociated by reduction, with electrophoretic mobilities identical to platelet glycoproteins previously designated lea and IcB. The beta chain comigrates with that platelet glycoprotein known as GPIIa. In an immunoblot assay, anti-band 3 binds to GPIIa but not to GPIc. The fact that anti-band 3 iramunoprecipitates both GP therefore suggests that they exist in a complex.Our findings establish GPIc-IIa as yet another platelet glycoprotein receptor complex and pave the way for future studies of the relative role of GPIIb-IIIa and GPIc-IIa in the adhesion of platelets to physiologic surfaces.
DEVELOPMENT OF A COAGULATION FACTOR X CONCENTRATE AS A BY-PRODUCT OF COAGULATION FACTOR IX PRODUCTION
Authors: Miekka Shirley I , Clark David B , Menache Doris 
Published: August 2018

The American Red Cross is developing a Coagu.lation Factor X (FX) concentrate to provide a safer alternative for replacement therapy in Factor X deficient patients, who can experience thromboembolic complications with current treatments. Based on a survey of hemophilia treatment centers, we estimate the frequency of the homozygous disorder to be approximately 1/150th that of hemophilia A, or about 65 patients in the USA. We have devised a method for producing FX as a by-product of our Coagulation Factor IX concentrate (FIX). The method starts with adsorption of cryoprecipitate supernatant plasma with DEAE-Sephadex resin followed by elution of Vitamin K-dependent coagulation factors. This material is adsorbed to sulfated dejctran resin and Factors II and X are eluted by increasing the salt concentration. At 0.45 M NaCl, FII elutes quickly while FX is retarded and can be recovered essentially free of FIX by collecting the slower eluting material. FIX is then recovered at still higher ionic strength. The pooled FX is concentrated, diafiltered and treated to inactivate viruses. Approximately 30% of plasma FX was recovered in pilot scale experiments (600 liters plasma). Specific activity was > 51 FX units / mg protein corresponding to a purity of around 50% and 3000-fold purification over plasma. The ratios of Factors "X : II : IX : Protein C were 1.0 : <0.03 : <0.03 : 0.2. The major contaminant, conprising nearly 50% of the protein, was found to be inter-alpha trypsin inhibitor (IaI), a serine protease inhibitor whose function in plasma has not yet been determined. This inhibitor is also present in the DEAE-Sephadex eluate and. in the FIX concentrate. However, Western blot and HPLC analyses have shown that IaI is present in two different forms.In FX it behaves as expected for the IaI monomer (Mr = 160 kDa), while in the DEAE-eluate and in FIX it exists also in r higher molecular weight form (≥400 kDa) corresponding either to aggregates, complexes or larger native species not previously described. The nature of the possible interaction of Ial w.vdi these coagulation factors is unknown and is currently boinn evaluated.
THROMBOSIS IN PATIENTS WITH HEPARIN-INDUCED THROMBOCYTOPENIA
Authors: Chesney C M , Lands R H 
Published: August 2018

Heparin induced thrombocytopenia was studied in a 2000 bed teaching hospital with 21,000 admissions annually. Approximately 25% (5250 patients) received some form of heparin therapy (lV,SC, catheter flushes). In l6 months 21 patients(<0.4%) developed well-documented thrombocytopenia while receiving therapeutic bovine lung heparin(IV). In 12 cases thrombocytopenia appeared to be directly related to heparin (Group 1). In 9 cases additional disease processes were also present (Group 2). While on heparin 5 patients in Group 1 (42%) developed new thrombotic episodes including pulmonary embolus, transient ischemic attack, stroke, lower extremity embolus, and myocardial infarction. None of these complications resulted in death. All patients were tested by a sensitive and specific lUC-serotonin release method for detecting heparin-induced thrombocytopenia (Blood 67:27, 1986). The test was positive in 10/12 patients (83%) in Group 1 and only 4/9 (44%) in Group 2. All 5 patients with thrombosis had a positive test. In all 21 cases heparin was discontinued. Platelet count began to improve in 1-9 days and eventually returned to normal in all Group 1 patients and the 4 Group 2 patients with positive test. An additional patient not included in Group 1 or 2 received minidose heparin(8000 units SC q8h) as a prophylactic measure following cholcystectomy. Ten days later while still receiving heparin she developed a cold, pulseless right lower extremity. Test for heparin associated platelet antibody was strongly positive despite platelet count of 200,000 per ul. However review of medical record revealed platelet count of 433,000 the preceding day. Therefore the drop represents a decrease in platelet count of 54%. Embolectomy of the right femoral-popliteal system revealed white platelet thrombus. Heparin was discontinued.The results presented here demonstrate that the l4C-serotonin release test is sensitive and specific for heparin induced thrombocytopenia. Well documented heparin induced thrombocytopenia occurs infrequently. However, thrombosis is not rare in these patients and may occur in the absence of absolute thrombocytopenia. Increased awareness of this entity and more accurate diagnosis may result in decreased mortality and morbidity.
CONGENITAL MACROTHROMBOCYTOPENIA, LEUCOCYTE INCLUSIONS, DEAFNESS AND PROTEINURIA: FUNCTIONAL AND ELECTR0NMICROSCOPIC OBSERVATIONS ON PLATELETS AND MEGAKARYOCYTES
Authors: Blockmans D , Heynen M J , Vermylen J , Verwilghen R 
Published: August 2018

We report here a female patient of 33 years with a variant of Alport's syndrome (macrothrombocytopenia, leucocyte inclusions, deafness and proteinuria). The bleeding problems consisted of ecchymoses and menorrhagia, the deafness was of the ^ensorineural type. The platelet count in whole blood was 14.109/I, the mean platelet volume 22.8 μm3 . The template bleeding time exceeded 30 minutes. Ultrastructural studies of the peripheral blood revealed giant spheroid platelets with a high density of organelles, an abundance of vacuoles and an apparently disorganized microtubular system. In addition, unusual granule free areas were observed in the neutrophils of the patient and her mother. Granulocyte function was normal, except for a low myeloperoxidase content.Functional studies of the platelets in platelet rich plasma showed normal aggregation curves related to the low platelet number, although no shape change could be elicited. Platelet aggregation studies in whole blood (impedance method) gave supernormal aggregation curves; this suggests the limited usefulness of this technique in patients with such large platelets.The bone marrow contained numerous dysplastic megakaryocytes. In the mature granular megakaryocytes vacuoles and cysternae were organized in a radiating pattern demarkating elongated platelet territories. The platelet producing megakaryocytes showed fragmentation of the central zone and discharge of platelets through openings of the peripheral zone. These megakaryocytes had an immunological phenotype resembling that of very young cells (TR 14%, GP IIa 17% and GP IIIa 7%). The conversion of the elongated platelet territories into giant spheroid platelets probably results from remodelling within the circulation. The internalisation of plasma membranes would give rise to the extended invaginated canalicular system. Further studies are needed to explain the exact pathogenesis of this syndrome.
THE EFFECT OF ADRENALINE AND ISOPRENALINE ON INTRA-SPLENIC PLATELET KINETICS
Authors: Wadenvik H , Denfors I , Kutti J 
Published: August 2018

It is well recognized that adrenaline decreases and isoprenaline increases the splenic pooling of platelets. However, the action of these adrenergic drugs on the splenic pooling mechanism is still unclear. The aim of the present study was to investigate the effect of an adrenaline and an isoprenaline infusion on intrasplenic platelet kinetics.Methods. 13 healthy male volunteers were recruited for the study. Autologous platelets were labelled with 111-Indium-oxine. 6 of the subjects received an isoprenaline infusion (0.03 ug/kg/min) over 30 min, 4 an adrenaline infusion (0.2 ug/kg/min) and 3 an adrenaline infusion (0.1 ug/kg/min) over 40 min. The splenic blood flow (SBF), intrasplenic platelet transit time (PTT) and splenic platelet pool size (SPP) were determined twice by compartmental analysis of the equilibration of radiolabelled platelets between circulating blood and splenic platelet pool: at baseline and during the infusions. The initial distribution of injected radiolabelled platelets, between blood and splenic platelet pool, was followed dynamically with a computer assisted gamma camera. Student’s t-test for paired data was employed for comparison of mean values.Comments. The adrenaline infusion diminished the exchangeable splenic platelet pool size by lowering the splenic blood flow. In contrast, the isoprenaline infusion increased the splenic pooling of platelets by increasing the splenic blood flow.
IN VIVO BEHAVIOR OF DETERGENT SOLUBILIZED PURIFIED RABBIT THROMBOMODULIN INJECTED INTO RABBITS
Authors: Ehrlich H J , Bang N U , Esmon N L , Esmon C T 
Published: August 2018

Thrombomodulin (TM) is a thrombin (T) endothelial cell membrane receptor; the TM-T complex readily activates protein C resulting in anticoagulant activity. We investigated the biological effects of detergent solubilized purified rabbit TM upon i.v. injection into rabbits. 125I-labelled TM (lactoperoxidase method) disappeared from the circulation to a t 1/2 of 2.5 h.The administration to rabbits of 2 doses of 50 μg/kg endotoxin 24 h apart did not accelerate the turnover rate of TM. In vitro tests demonstrated that .27 nM TM was required to significantly prolong the activated partial thromboplastin time (APTT) and .54 nM to significantly prolong the thrombin clotting time (TCT). At 3.78 nM TM, both the APTT and TCT prolonged to infinity while the prothrombin time (PT) was minimally affected. When TM was injected into rabbits in doses of 100, 200 and 500 μg/ml APTT and TCT immediately prolonged in a dose dependent fashion whereas no effect on the PT was observed. The disappearance rate of anticoagulant activity (prolongation of the APTT and TCT) closely paralleled the disappearance rate of 125I-labelled TM; t 1/2 for anticoagulant activity being 2.0 h irrespective of the dose administered. Thus, detergent solubilized purified rabbit TM survives in the circulation for appreciable time periods. While in the circulation TM is capable of binding T thereby prolonging the TCT as well as activating protein C thereby causing degradation of factors Va and VIIIa resulting in a prolongation of the APTT. Therefore, TM administered i.v. results in an anticoagulant effect of surprisingly long duration.
NO EVIDENCE OF HEPATITIS OR HIV TRANSMISSION IN VIRGIN HAEMOPHILIC BOYS TREATED WITH BRITISH HEAT TREATED FACTOR VIII CONCENTRATE (8Y)
Authors: Pasi K J , Hill F G H 
Published: August 2018

HIV and hepatitis virus transmission is of major concern with factor VIII therapy. Non-A, non-B hepatitis (NANBH) has a near 100% incidence in patients previously treated with unheated large pool factor VIII concentrates. NHS heated high purity factor VIII concentrate (8Y) undergoes severe protracted heat treatment of the freeze dried concentrate theoretically sufficient to inactivate hepatitis viruses as well as HIV. Infusions of 22 different batches of 8Y have been given to 18 children with haemophilia A (10 virgin patients; 8 who had only received single donor cryoprecipitate) have been treated for up to 18 months. Regular testing for viral antibody seroconversion and biochemical liver enzymes have been made. None had had clinical or biochemical evidence of liver disease prior to the commencement of 8Y therapy. All these boys were immunized with HBVax at the time of the first treatment with 8Y and were HIV antibody negative.Liver function tests were to be performed monthly but due to patient non-compliance this was only achieved in 60% of patients.All patients receiving 8Y have remained anti-HIV seronegative. Only the virgin patients can be considered suitable for evaluation with regard to the transmission of NANBH. These boys by this time have received multiple batches of 8Y (mean 5 batches, range 1 to 14). In only 1 patient has an isolated rise in aspartate transaminase (AST) been noted (AST 27 to 131 IU/1) 6 weeks aftertreatment with a new batch, but no rise in alanine transaminase (ALT) or clinical evidence of liver disease was found. Viral serology was performed. AST returned to normal within 12 days.This batch was received by 3 of the virgin and 2 of the previously cryoprecipitate treated boys. All these 5 boys who were exposed to the suspect batch had normal liver enzyme levels when measured within 4-6 weeks of exposure.Of the 10 virgin patients receiving multiple batches of 8Y a transient rise in AST but with no rise in ALT has only been noted in 1 patient. In the absence of firm biochemical evidence of liver disease NANBH is an unlikely cause. Lack of transaminase rises in other virgin patients strengthens this assunption. We conclude that 8Y reduces the incidence of NANBH and HIV transmission.
INTERACTION OF POLYMORPHONUCLEAR LEUKOCYTES AND ENDOTHELIAL CELLS : FUNCTIONAL CONSEQUENCES
Authors: Williams T J , Rampart M , Nourshargh S , Hellewell P G , Brain S D , Jose P J 
Published: August 2018

The mechanisms involved in the accumulation of polymorphonuclear leukocytes (PMNs) in an inflammatory reaction are complex. A key phase in this process is the attachment of the PMN to the microvascular (venular in most tissues) endothelial cell, initiated by the extravascular generation of a chemical mediator. Experiments in vitro suggest that mediators, such as C5a, may act in vivo by stimulating the increased expression of the CD18 complex on the surface of the PMN within the venule lumen (1), whereas IL-1 may act by causing the expression of an adhesive molecule on the endothelial cell (2). In vitro the former process is rapid whereas the latter is slow in onset. We have measured the local accumulation of intravenously-injected Ulln-PMNs in response to intradermally-injected mediators in the rabbit, in order to investigate possible mechanisms in vivo. PMN accumulation was found to be rapid in onset in response to C5a, the rate of accumulation falling progressively to low levels by 4 hours. In contrast PMN accumulation in response to IL-1 was slow in onset, reaching a peak rate at 3-4 hours. Intradermal injection of the vasodilator prostaglandins PGI2; PGE2 and the neuropeptides VIP and CGRP caused a marked potentiation of the rate of leukocyte accumulation. PMN accumulation induced by C5a was associated with increased microvascular permeability, as indicated by the leakage of intravenously-injected 125I-albumin with a time-course in parallel with the rate of PMN accumulation enhanced by intradermally-injected vasodilators. Depletion of circulating PMNs abolishes these responses to C5a (3). In contrast, leukocyte accumulation induced by IL-1 was associated with little plasma protein leakage, even in the presence of intradermal vasodilators. This observation indicates that PMN emigration itself does not lead to increased microvascular permeability. C5a, but not IL-1, may stimulate emigrating PMNs to secrete an endogenous factor that increases permeability by an action on endothelial cells (3). This factor does not appear to be the phospholipid PAF (4). In contrast to the enhancing effects of local PGI2, intravenously-infused PGI2 inhibited PMN accumulation induced by C5a and IL-1, and plasma protein leakage induced by C5a (5). This effect is probably mediated by elevation of cyclic AMP in intravascular PMNs. We have shown that C5a stimulation of PMNs in contact with endothelial cells in vitro induces endothelial cell PGI2 secretion (6). Thus, PGI2 may be part of a negative feedback system in vivo to control interactions between PMNs and endothelial cells.These observations provide some clues to the intricacies of mechanisms of leukocyte accumulation in vivo.