bluuebunny/crossref_metadata_2025 huggingface dataset:
Triage bei Kopfschmerz in der Hausarztpraxis: Wann einweisen?
Authors: Carmienke Solveig , Holle-Lee Dagny
Published: May 2019

Headache is one of patients’ most common reasons to consult their general practitioner and covers about 2 – 5 % of the consultations in primary care. Often, the general practitioner is the first to be contacted by patients with headache. Mostly, headaches are primary and only 2 % of the patients have secondary headaches. The distinction between primary and secondary headache is the most important step in the management of patients with headache in primary care. Therefore, this article shows important elements of anamnesis and examination of headache patients in primary care. Furthermore, this article focuses on identification of red flags and yellow flags in the consultation of patients with headache and suggests recommendations for referral to emergency department, hospital care or specialist treatment.
Polymikrobielle Leberabszesse und Pleuraempyem bei einem 40-jährigen Mann nach Zahnextraktion und geschlossener Parodontitis-Behandlung: Ein Fallbericht
Authors: Schimmel Thomas , Trawinski Henning , Karlas Thomas , Wendt Sebastian , Lübbert Christoph
Published: May 2019

Wir berichten über einen 40-jährigen Patienten, der sich mit Fieber, Druckschmerz im rechten Oberbauch, atemabhängigen rechtsseitigen Thoraxschmerzen und akuter Dyspnoe vorstellte. Bildgebend wurden größere Leberabszesse sowie ein ausgedehntes rechtsseitiges Pleuraempyem nachgewiesen. 16 Wochen zuvor hatte sich der Patient einer Zahnextraktion aller 3. Molaren (18, 28, 38, 48) und eines 1. Molaren (46) sowie einer systematischen geschlossenen Parodontitis-Behandlung unterzogen. Im Pleura- bzw. Leberabszesspunktat wurden vier verschiedene Spezies der physiologischen Mikrobiota der Mundhöhle nachgewiesen (Streptococcus anginosus, Streptococcus constellatus, Actinomyces odontolyticus, Prevotella denticola). Ein zugrundeliegender Immundefekt war nicht nachweisbar. Unter sonografisch gesteuerter Leberabszessdrainage, operativer Versorgung des Pleuraempyems mittels Video-assistierter Thorakoskopie (VATS) und Einlage von Thorax-Saug-Drainagen sowie gezielter antibiotischer Therapie kam es innerhalb von 6 Wochen zur vollständigen Ausheilung. Der Fallbericht veranschaulicht relevante Infektionsrisiken größerer zahnärztlicher Interventionen und diskutiert die derzeitigen Empfehlungen zur Durchführung einer periinterventionellen antimikrobiellen Therapie in den zahnärztlichen Leitlinien. Wichtig erscheint vor allem eine gute klinische Nachbeobachtung betroffener Patienten.
Evolution of HCV patient characteristics and DAA regimens in the German Hepatitis C Registry (DHC-R) in 2014 and 2015
Authors: Sarrazin Christoph , Buggisch Peter , Mauss Stefan , Müller Tobias , Zimmermann Tim , Klinker Hartwig , Pathil-Warth Anita , Schlag Michael , Nalpas Catherine , Wegner Sven , Lonjon-Domanec Isabelle , Simon Karl-Georg
Published: May 2019

Background The urgent need in HCV-infected patients with liver disease mandated the rapid implementation of IFN-free DAA combination therapies following their regulatory approval in 2014 and 2015 without full knowledge of the optimal combinations and regimens. Investigating the evolution of the DAA utilization patterns and treatment outcomes could provide learnings for future situations.

Methods This was an analysis of a prospective observational database from the German Hepatitis C Registry (DHC-R) covering a period from May 2014 to September 2015. Adult patients had evidence of chronic HCV GT1 or GT4 infection and were treated with an IFN-free combination regimen of simeprevir (SMV) + sofosbuvir (SOF) or other IFN-free regimens: daclatasvir + sofosbuvir (DCV + SOF), ledipasvir/sofosbuvir (SOF/LDV), paritaprevir/r + ombitasvir ± dasabuvir (PrOD), with or without ribavirine (R).

Results A total of 5496 subjects were followed during the period. During this period, clinical recommendations and treatment patterns evolved rapidly in response to new evidence from clinical trials and clinical routine and regulatory approval of additional regimens. High SVR12 rates were seen in this cohort, even in hard-to-treat patient subgroups. In the multivariate analysis, gender, age, advanced cirrhosis, and intensified treatment for cirrhotics were associated with treatment outcome.

Conclusion Despite limited knowledge of the optimal utilization of the newly approved DAA combinations and treatment durations as well as their comparative efficacy and safety profiles, high SVR rates were achieved regardless of the DAA combination. These outcomes were facilitated by the rapid adaptation of clinical recommendations. Future situations with high unmet medical need may follow a similar approach.
Natrium carbonicum und der Pfeffer
Authors: Fischer Ulrich
Published: May 2019

Ein klinisches Symptom ist richtungsweisend bei der Behandlung eines Patienten, der seit 30 Jahren an einer Dermatitis leidet. Die wesentlichen Symptome wurden anhand von Bönninghausens Therapeutischem Taschenbuch analysiert, repertorisiert und anschließend erfolgreich mit Natrium carbonicum behandelt. In den weiteren Ausführungen wird einerseits verdeutlicht, wie bedeutsam eine klare Trennung zwischen verifizierten Prüfungssymptomen und klinischen Symptomen bzw. wie sinnvoll eine Kennzeichnung der klinischen Symptome ist und andererseits in welcher Hinsicht klinische Symptome unsere Arbeit bereichern können.
Bradykinin Generation In Clinical Blood Samples: An Index Of Intrinsic Coagulation Activation
Authors: van Rosevelt R F , Philipse-Hoorweq E M , Armstrong A L , van Aken W G , Mourik J A van
Published: May 2019

Fibrinopeptide A (FPA) is frequently elevated in thromboembolism and DIC. As to the question which of the coagulation pathways (intrinsic vs extrinsic) is triggered under these conditions, measurement of FPA alone permits no answer. We measured both kallikrein (reflecting contact activation) and FPA in patients with classical DIC, established deep venous thrombosis (DVT) and pulmonary embolism (PE). Circulating kallikrein was measured as its capacity to generate bradykinin (BK) in whole blood (in the presence of kininase inhibitors) immediately after venipuncture. After incubation for 10 min. at room temp, kallikrein inhibitors were added. The increment of the BK concentration (ΔBK) -measured by radioimmunoassay- was used as marker of circulating kallikrein. The specificity and sensitivity of this method were superior to amidolytic assays for kallikrein.

These data indicate the measurement of ΔBK is a potential tool to discriminate between intrinsic and extrinsic coagulation activation. In 2 patients with thromboembolism FPA normalised upon adequate anticogulant therapy, whereas ΔBK remained accerlerated for several weeks. This observation suggests that in these patients anticoagulant therapy was not adequate in eleminating the cause of coagulation activation. It is known that FPA is also frequently elevated in a variety of disorders not primarily associated with thrombosis or DIC. In 36 patients with chronic inflammatory diseases, FPA was elevated in 20 cases, whereas only in 1 patient ΔBK was accelerated (together with elevated FPA). We therefore conclude that the measurement of ΔBK is a more specific marker of intravascular coagulation activation, than the measurement of FPA.
Isolation And Characterization Of A Cdna Clone Coding For α1-Antitrypsin
Authors: Kurachi K , Chandra K , Woo S L C , Davie E W
Published: May 2019

Poly(A)-RNA enriched for α1-antitrypsin was isolated by specific immunoprecipitation of baboon liver polysomes. Alpha1-antitrypsin consisted of greater than 90% of the cell-free translation products of this mRNA. A doublestranded cDNA was synthesized by using reverse transcriptase and made blunt-ended with nuclease S1. After tailing with dCTP and terminal transferase, the double-stranded cDNA was annealed to pBR322 DNA. The latter DNA had been cleaved previously at the single Pst I site and similarly tailed with dGTP. The resulting plasmids were used to transform Escherichia coli strain RR1. Clones that hybridized to 32P-labeled cDNA synthesized from the α1-antitrypsin-enriched mRNA were then identified. The recombinants containing baboon cDNA inserts were further screened by a solution hybridization assay with [3H]cDNA synthesized from the enriched mRNA. The cDNA inserts from the positive clones were then sequenced to identify clones containing α1-antitrypsin. One insert, designated pBaαA1, was found to code for the carboxyl-terminal region of α1-antitrypsin. It also contained a noncoding region of 76 base pairs and a poly(A) tail of 60 base pairs.
Mechanism Of Proteinase-Binding To α2-Macroglobulin
Authors: Sottrup-Jensen L , Hansen H F , Mortensen S B , Petersen T E , Magnusson S , Jörnvall H
Published: May 2019

Peptide-bond cleavage in the bait-region of the α2M polypeptide chains, associated with proteinase-α2M complex formation was shown to occur in the sequence: GLRVGFYESDVMGRG HARLVH (part of the 298-residue “middle” segment) such that plasmin, thrombin and trypsin cleave the Arg-Leu (RL) bond, elastase mainly the Val-Gly (VG) and partly the Gly-Phe (GF) bonds. Thus, in the first step of complex formation the proteinase active site binds to the bait region of α2M.

J.B. Howard found that heat inactivation of α2M causes cleavage of a Glu-Glx bond, and inactivation with CH3NH2 leads to incorporation of CH3NH2 on the γ-carboxyl of the same Glx-residue. We have found that reaction of α2M with trypsin or elastase but not trypsinogen causes the 2:1 stoichiometric appearance of up to 4 SH-/tetrameric α2M for up to 2 trypsin/2 elastase bound. The -SH was found to be the Cys-SH of thiol-ester-containing PYGCGEZNM sequence. Inactivation of native α2M (by CH3NH2, heating, dissolving in 2 M guanidine or 1.6% SDS at 50°C) also led to the appearance of up to 4 SH for 4 CH3NH2 incorporated per tet- rameric α2M. Competition between trypsin and CH3NH2 for the thiol-ester site proved that trypsin binds to the same Glx- γ-carboxyl that incorporates CH3NH2. Thus, the second step of complex formation involves the thiol-ester site γ-carboxyl of α2M and a proteinase-site other than its active- site (since the complex is active against small substrates). Other compounds with nucleophiles, e.g. putrescine or insulin, when added with trypsin, were also incorporated into α2M. The thiol-ester mechanism may be associated with the rapid elimination of proteinase α2M-complexes, particularly since the recent finding by B.F. Tack that complement factor C3 has an identical thiol-ester-containing sequence, points to at least one common function of these proteins.
Objective Assessment Of Late Results Of Treatment Of Deep Vein Thrombosis
Authors: Kakkar V V , Lawrence D
Published: May 2019

There have been few prospective clinical studies where the haemodynamic status of the deep venous system has been assessed objectively following an episode of deep vein thrombosis (DVT). Recently, a foot plethysmograph has been developed in Sweden which allows haemodynamic changes in the venous system to be measured accurately. The purpose of this prospective study was to investigate the haemodynamic changes that may occur following treatment of DVT using heparin or thrombolytic therapy.

113 Patients were included in this prospective study. None of these had suffered previous DVT or had signs and symptoms of the post-phlebitic limb at the time of initial treatment. In each case the diagnosis was established by ascending phlebography. In 79 of these patients, thrombus (>5 cm) was confined to the calf veins only. Initial treatment in these patients consisted of heparin in therapeutic anticoagulant dosage (adjusted according to APTT) for 7 days. Oral anticoagulation with Warfarin was commenced on the 5th day of treatment and continued for a minimum of 3 months. The remaining 34 patients had extensive thrombosis involving femoral, popliteal and iliac veins; in 32 of these calf veins were also involved. They received streptokinase therapy for a period of 5 days; the loading dose of 500,000 IU of SK was followed by a maintenance dose of 100,000 units per hour. Thrombolytic therapy was followed by heparin for 3 days and concurrent anticoagulation with Warfarin, the latter being given for a minimum of 6 months. In both groups, ascending phlebography was repeated on the 6th or 7th day to assess the extent of lysis or propagation of the thrombus. Haemodynamic assessment by foot volumetry was performed at 6,12 and 24 months following the initial episode of DVT. 77% Of 34 patients who sustained major DVT and received SK and 16% of 79 patients with calf vein thrombosis treated with heparin had evidence of severe haemodynamic impairment equivalent to that seen in the established post phlebitic syndro.
Haemostatic And Eaemorheological Abnormalities In Type II Hyperlipoproteinaemia
Authors: McArdle B M , Lowe G D O , Drummond M M , Whigham K A , Stromberg P , Lorimer A R , Forbes C D , Prentice C R M
Published: May 2019

Thirty subjects with type II hyperlipoprotein- aemia had significantly higher levels of the three major determinants of blood viscosity (haematocrit, fibrinogen and alpha-2-macroglobulin), the two major inhibitors of plasmin (fast antiplasmin activity and alpha-2-macroglobulin), and plasminogen, compared to 30 controls matched for age, sex and smoking habit. Fibrinogen, fast antiplasmin activity and plasminogen correlated significantly with total and LDL (low density lipoprotein) cholesterol. In vitro, purified LDL did not affect these assays. Levels of factor VIII coagulant activity and related antigen, anti-thrombin III, platelet count, platelet aggregation to ADP and adrenaline, and red cell deformability did not differ significantly in the two groups. Haematological variables were unrelated to HDL cholesterol, VLDL cholesterol, or triglyceride. We suggest that these abnormalities might promote arterial disease in type II hyperlipoproteinaemia by three mechanisms - increased blood viscosity, inhibition of fibrinolysis, and arterial wall infiltration by macromolecules (fibrinogen and alpha-2-macroglobulin as well at LDL).
Platelet Antiaggregation And Hemodynamic Effects Of Adenosine And Prostaglandin E1
Authors: Masters T N , Born G G V , Robicsek F
Published: May 2019

The efficacy of using adenosine (ADEN) or prostaglandin E1 to prevent platelet loss during cardiopulmonary bypass (CPB) was tested in experiments conducted with a simulated CPB circuit with cardiotomy suction. The hemodynamic effects of the compounds were tested in a canine model to ascertain hemodynamic tolerance of platelet-salvaging concentrations. Platelet concentrations were determined with a Coulter Counter Model S-Plus in 3 separate groups each composed of 10 experiments in which ADEN (1 mM) and PGE1 (0.5 and 1.0 pM) were added to the bypass circuit to determine platelet protection. It was found that PGE1 at both concentrations was effective in salvaging platelets (0.5 μM-70% and 1.0 μM- 95%) and ADEN at 1 mM salvaged approximately 45%. Hemodynamic effects of PGE1 and ADEN were studied in heparinized (3 mg/Kg) mongrel dogs weighing between 20-27 Kg which were artificially ventilated and catheters placed to measure LV and arterial pressures and dP/dT. Via a left thoracotomy a 15 mm Statham Blood Flowmeter Probe placed around the ascending aorta measured Cardiac Output (CO). ADEN, in seven increasing doses (range: 0.45-3.40 mg/Kg/min), was infused for 2 min with 5 min between doses. In two groups of 10 dogs each, PGE1 was infused with two dose ranges: 0.40-3.00 and 2.0-15.0 μg/Kg/min. PGE1 was infused in seven increasing doses in each range and hemodynamic changes recorded during 3 min of infusion. Hemodynamically, ADEN reduced heart rate and systemic resistance progressively as dose increased. The unchanging CO accompanied by reduced dP/dT and LV pressure suggests the myocardium is not directly affected. PGE1 on the other hand, continually reduced CO and dP/dT with increasing doses, but with the smallest dose reduced arterial pressure 20% which remained unchanged with increasing doses. PGE1 therefore appears to directly depress the myocardium with maximal resistance changes occurring with the smallest doses. Platelet salvaging concentrations for ADEN were within acceptable limits, whereas PGE1 was not.
Treatment Of Neonatal Renal Vein Thrombosis (RVT) With Urokinase (UK)
Authors: Pavon A Fernandez , Chacon J L Fernandez , Cuesta M V , Ruza J
Published: May 2019

The renal vein thrombosis represents an etiologic and therapeutic problem in neonatal pathology. In our milieu RVT is more frequent than is reported in current bibliography. This high incidence is accompanied by a high mortality and by the development of chronic renal failure among survivors.

This fact led us to try a thrombolytic treatment that could be effective with respect to life and renal function.

In our study we describe seven newborn patients with radiologically checked RVT. All cases were secondary to hypernatremic dehydration syndrome due to hyperconcentrated feeding. Five patients were submitted to thrombolytic treatment with UK, one patient was treated with heparin and one patient died before any treatment could be tried.

Among the five patients treated with UK, four showed a complete recanalization of the RVT, with recovery of their renal function without sequelae. One of the patients, in whom an intracraneal haemorrhage was clinically suspected, died during treatment.

The dosage of UK during treatment was 1000 U/Kg/hour.

The duration of treatment was individualized according to renal evolution.

The description of treated cases will be done individually.
Clotting Abnormalities During The Course Of Immune Type Glomerulonephritis Induced By HgCl2 In The Brown Norway(BN) Rat
Authors: Michaud A , Sapin C , Aiach M , Druet P , Forestier F
Published: May 2019

This work was undertaken in order to study the clotting process during the course of a biphasic immune type glomerulonephritis(GN) induced by HgCl2 in the Brown Norway (BN) rat. This toxic agent induced, in a first stage, the formation of antiglomerular basement membrane (GBM) antibodies and in a second stage an immune complex type GN.

Antithrombin III (AT III), fibrinogen degradation products (FDP) were evaluated by electroimmunodiffusion using specific antisera. The other classical methods were performed using commercial reagents. The albumin platelet content was evaluated by a radioimmunoassay. The soluble fibrin monomer complexes (FM) were searched for by a chromatographic technic.

Transient clotting proteins variations related to nephrotic syndrome were observed (low AT III level, low factor XII activity, increased factor V activity, and increased fibrinogen level). Disseminated intravascular coagulation (DIC) was suggested by low platelet counts, rise in FDP, and presence of FM. Fibrin thrombi in the glomerular capillaries were observed by immunofluorescence in the rats presenting evidence for DIC and in rats which died during the first phase of the disease. The animals presenting DIC syndrome were the most severy ill.

The hypothesis of an activation of the haemostasis system was supported by a low albumin platelet content and by the existence of high molecular weight fibrinogen derivatives. Complement activation, immune complexes, endothelial lesion might trigger DIC : the mechanism responsable for initiation of DIC are under study.

According to this results, it can be concluded that DIC which is responsable for the death of several animals, plays a major pathogenic role in this experimental autoimmune disease.
Reversible And Irreversible Changes In Platelet CytoSkeleton, Energy Charge And Secretion By Exposure To pH 5.3
Authors: Mürer E H , Stewart G J , Tuszynski G
Published: May 2019

The effects of exposure to low pH on platelets isolated from plasma by saline-EDTA washing or by gel filtration were compared. Platelets isolated by the two methods responded similarly with respect to nucleotide metabolism, secretion and leakage. Both showed decrease in adenylate energy charge and metabolic ATP level after exposure to pH 5.3 and reversal to almost normal levels after return to pH 7.6, while the pH 5.3-induced secretion (about 50% of preabsorbed serotonin) was continued after the return to higher pH and reached about 80%. The response of the cytoskeleton of EDTA-washed and gel-filtered platelets to low pH differed quantitatively. Analysis of transmission electron micrographs (TEM):TEM of Triton-insoluble residue showed similar structures and SDS-PAGE showed primarily actin (42,000d) and likely myosin (200,000d). Conclusion: The method of isolation did not affect the response of platelets to low pH when studied by platelet nucleotide metabolism, secretion or leakage but did affect the response when studied by the organization of the cytoskeleton (actin filaments).
Cyclic Nucleotides And PlateLet-Leukocyte Aggregates In Patients With Occlusive Arterial Disease
Authors: Kiersnowska-Rogowska B , Bielawiec M , Rogowski F , Bodzenta A , Giedrojć J
Published: May 2019

The aim of the work was to study plasma cyclic nucleotides level /cyclic-AMP and cyclic-GMP/ and platelet-leukocyte aggregates in patients with obliterative arteriosclerosis of the lower limbs. The following parameters were investigated: cyclic-AMP and GMP radioimmunochemically by the use of a cyclic-AMP and cyclic-GMP assay kit, platelet leukocyte aggregates by the method of Silbergleit in our modification.

The remainder were hospital deaths, five within the first 48 hours, 8 between 2-14 days, and 4 between 14 and 94 days. Five cases showed subendocardial infarction (SI). Four of these occuring out of hospital. The remaining 18 cases were transmural infarctions (TI). Sixteen of 17 hospital deaths exhibited TI.

Significant decrease in cyclic-AMP level was found in arteriosclerotic patients in comparison to the control group. No significant changes in cyclic-GMP level were observed. The number of platelet-leukocyte aggregates was significantly higher in these patients.

The influence of Hydroxy-ethyl-rutosides /HR/ on investigated parameters was also studie.

The significant decrease in the level of cyclic-GMP in plasma was observed in patients after intravenously injection of 1000 mg of HR whereas no changes were found in cyclic-AMP level. The significant decrease in the number of platelet-leukocytes aggregates in the blood obtained after HR injection was also observed.

Our results have shown the disturbances of plasma cyclic nucleotides balance and function of platelets and leukocyte in arteriosclerotic patients. This study also suggests that HR may be a modulating agent of these parameters.
Role of Cyclic Amp in Platelet Function : Evidence from Inhibition of Adenylate Cyclase in Intact Platelets by Adenosine Analogues
Authors: Haslam R. J. , Davidson M. M. L. , Desjardins J. V.
Published: May 2019

Adenosine exerts independent stimulatory and inhibitory effects on the adenylate cyclase activity of platelet particulate fractions (Haslam & Lynham, 1972). Two adenosine analogues, 9-(tetrahydro-2-furyl) adenine (SQ 22536) and 2′, 5′-dideoxyadenosine (DDA) have now been found to show marked non-competitive inhibitory activities only. Basal and PGE1-stimulated adenylate cyclase activities were inhibited ~50% and ~70% respectively by 100 μM SQ 22536 and ~60% and ~80% respectively by 100 μM DDA. Both compounds also inhibited adenylate cyclase in intact platelets, when this was measured as the increase in cyclic [3H]AMP in platelets labelled with [3H] adenine and then incubated with papaverine. At the concentrations tested (10-500 μM), neither SQ 22536 nor DDA induced platelet aggregation or potentiated aggregation and release of [14C] 5-HT induced by suboptimal concentrations of ADP, Arg8-vasopressin, arachidonic acid or collagen added to heparinized or citrated platelet-rich plasma. However, both compounds partially blocked the inhibition by PGE1 or papaverine of aggregation induced by ADP or Arg8-vasopressin. From the concentrations exerting equal effects, DDA was ~3 times as potent in this regard as SQ 22536. Above 100 μM, the anti-inhibitory effects of both compounds decreased. The actions of these compounds in overcoming inhibition of aggregation by PGE1 were correlated with decreases in platelet cyclic [3H]AMP in platelets labelled with [3H] adenine. The results show that cyclic AMP plays no role in the responses of platelets to aggregating agents unless the platelet cyclic AMP level is elevated above the resting level and confirm that the effects of PGE1 on platelet function are mediated by cyclic AMP.
Studies of Superoxide Production in Human Platelets
Authors: Marcus A. J. , Silk S. T. , Safier L. B. , Ullman H. L. , Broekman M. J.
Published: May 2019

Superoxide radicals are measurable in platelet suspensions by two different assay systems (SOD-inhibitable cytochrome c and nitroblue tetrazolium reduction). The

-producing mechanism is detectable in platelet supernatants and the rate of

generation is proportional to platelet concentration. Prior ingestion of aspirin does not inhibit

production by platelets. Following stimulation by collagen and thrombin platelets develop a marked increase in their capacity for cytochrome c and NBT reduction, but in contrast to leukocytes stimulation does not result in an increase in superoxide production. Sodium fluoride, a powerful stimulus for

production in leukocytes, and also a platelet aggregating agent, is under study. A concentration of 20 mM sodium fluoride increases platelet superoxide production approximately twofold over baseline values. This level of fluoride induces 88% 5-HT release in the absence of LDH leakage. The increase in superoxide generation does not correlate directly with 5-HT release, and the fluoride effect on

may not be related to its influence on secretion. Electrophoretic studies indicated that fluoride had no effect on platelet superoxide dismutase. Experiments with ionophore A23187 indicated that the action of this agent was not comparable to fluoride but was similar to collagen and thrombin in that

production did not exceed baseline levels. In some instances the ionophore appeared to inhibit

production. Superoxide radicals may play an important role in platelet-platelet or platelet-blood vessel interactions.
Ca2+ Regulation of Bovine Thrombosthenin
Authors: Muszbek L. , Kuźnicki J. , Drabikowski W.
Published: May 2019

In order to reveal the type of its Ca2+-regulation bovine thrombosthenin - natural platelet actomyosin - was investigated by competitive actin and myosin binding assay and by urea gel electrophoresis. The Ca2+-sensitivity of low ionic strength Mg-ATPase activity of platelet actomyosin was not influenced by the addition of excess skeletal muscle actin free of regulatory proteins. In contrast, the replacement of platelet myosin by skeletal muscle myosin resulted in a hybrid actomyosin insensitive to Ca2+. If actomyosin was reconstructed from crude platelet actin and skeletal muscle myosin again no regulatory effect of Ca2+ could be observed. In the presence of EGTA a fast moving band with the mobility corresponding to muscle troponin C (TN-C) was detected by alkaline urea Polyacrylamide gel electrophoresis. However, if Ca2+ was added this protein, unlike TN-C, neither changed its mobility nor formed a complex even if muscle troponin I was included into the system. The experimental results indicate that our Ca2+ sensitive bovine thrombosthenin preparation does not contain TN-C like component and is not regulated by an actin -, but rather by a myosin-connected system.
Significant Role of Fragment 1·2 Plus Light Chain of Bovine High Molecular Weight Kininogen in Contact Mediated Coagulation
Authors: Waldmann R. , Scicli A. G. , Scicli G. M. , Guimaraes J. A. , Carretero O. A. , Kato H. , Han Y. N. , Iwanaga S.
Published: May 2019

We have previously reported that high molecular weight kininogen (HMWK) partially corrects the partial thromboplastin time (aPTT) of Fitzgerald Trait plasma (HMWK deficiency), and in addition, we have also reported that kinin free HMWK which is also Fragment 1·2 free has decreased correcting capability. Presently, we have studied the effect of lysyl-bradykinin-free HMWK with Fragment 1·2 intact, obtained by hydrolysis of HMWK with urinary kallikrein. In addition, the effect of light chain with and without attached Fragment 1·2 were studied. When the correcting capacity of HMWK was compared, on a molar basis with the above-mentioned fragments, the following results were obtained. Kinin and Fragment 1.2 free HMWK has a significantly decreased correcting capacity on the aPTT of Fitzgerald trait plasma while lysyl-bradykinin free HMWK shows no decrease in correcting capability. Light chain with Fragment 1·2 attached also has intact correcting capability while light chain alone has a significantly decreased correcting activity. The above data suggest that the active site of HMWK resides in the light chain with Fragment 1·2 attached. Since Fragment 1·2 alone has inhibitory capacity, it may serve as a binding site and the light chain as the active site.
Activation of Hageman Factor by Factor XIA-HMW Kininogen
Authors: Meier Henry L. , Thompson Russell E. , Kaplan Allen P.
Published: May 2019

Plasma deficient in prekallikrein possesses an abnormality in the kinetics of surface dependent initiation of coagulation and fibrinolysis. This defect appears to be secondary to a diminished rate of Hageman factor activation, however the abnormality is progressively diminished as the time of incubation of the plasma with appropriate surfaces is increased. Since factor XI and prekallikrein circulate bound to HMW kininogen and HMW kininogen is known to augment the activation of factor XI and prekallikrein by activated Hageman factor, the ability of factor XIA to activate Hageman factor was examined. One ug Hageman factor was bound to supercel alone, or in the presence of 1.5 ug HMW kininogen and 0.1 to 1.0 ug factor XIA for varying time periods (0–60 min). The pellet was washed X 3 and bound activated Hageman factor was assayed by its ability to convert prekallikrein to kallikrein. The kallikrein generated was quantified by release of p-nitroaniline from α-benzoyl phe-val-arg-p-nitroanilide. Factor XIA alone was a weak activator of Hageman factor and the quantity of kallikrein generated was augmented when HMW kininogen was included in the incubation mixture. With limited HMW kininogen the Hageman factor activity appeared proportional to the factor XIA added. The same result was obtained with factor XIA isolated from prekallikrein deficient plasma. The data suggest that factor XIA plus HMW kininogen may represent one of the additional feedback mechanisms by which Hageman factor may be activated and thereby contribute to the gradual activation observed in prekallikrein deficient plasma.
Preparation and Applications of 125-I Labelled Heparin
Authors: Pepper D. S. , Dawes Joan
Published: May 2019

Commercial heparin was derivatized with Bolton and Hunter reagent (SHPP, succinimidyl-hydroxyphenylpropionate) and the reaction products chromatographed on protamine agarose. Biologically active heparin eluted at 1.2 – 1.4 M NaCl and was iodinated by the chloramine T method, . giving a product of specific activity 20 uCi/ug. The labelled heparin co-chromatographed with cold, biologically active heparin on protamine agarose. Using 0.4ng labelled tracer and 102 protamine agarose beads diluted in bland gel, it was possible to measure heparin concentrations of 0.001 u/ml by inhibition of binding. Heparin tracer alone eluted from 6% agarose molecular sieve columns earlier than heparin + plasma, indicating that the hydrodynamic radius of heparin is reduced when bound to AT III under physiological conditions. When injected intravenously, the label showed complex kinetics and did not have a single half life, clearance was initially rapid, but slowed later. Between 80–160 minutes following injection, the levels of circulating radioactivity rose slightly before falling again. This effect was reproducible in different normal persons and was independent of added heparin over a 100 fold range of dose.
Experience with an Amidolytic Heparin Assay Method
Authors: Teien A. N. , Abildgaard U. , Gjesdal K. , Holm A. H.
Published: May 2019

The amidolytic method using Xa, purified antithrombin III(At-III) and the chromogenic substrate Bz-Ile-Glu-Gly-Arg-pNA(S-2222 from KABI, Stockholm, Sweden) measures heparin down to a concentration of 0.010 U/ml plasma. The accuracy of the method was evaluated by adding heparin to plasma samples from 10 normals and 10 patients, to concentrations of 0.05 and 0.5 U/ml. The results indicated that the content was assayed more accurately than by existing clotting assays. The precision was 5 and 2 per cent of the mean value at the two heparin concentrations, respectively. The concentrations of platelet factor 4 and At-III in test plasma had some influence on the result at low heparin concentrations. The “error” due to At-III, however, may be corrected by a simple formula. Heparinized plasma(containing less than about 15 000 platelets/μl) could be kept at 0 – 4 °c for 24 hours before assay, kept frozen and thawed repeatedly without significant loss of heparin activity.

In plasma from patients on heparin prophylaxis with 5 000 U twice daily, heparin was detected in 98 per cent of the samples (range 0.010 – 0.241 U/ml).

The present assay method reflects heparin concentration more directly than the activated partial thromboplastin time test and the thrombin clotting time test. In order to investigate to what extent these three tests mirror the clinical effects of heparin (therapeutic response and bleeding complications) a collaborative study involving 14 hospitals has been started.
Unique Significance of Dihomo-γ-Linolenic Acid Mechanisms for the Prevention of Thrombosis
Authors: Willis A. L.
Published: May 2019

Common anti-platelet aggregatory agents act like aspirin to block cyclo-oxygenation of arachidonic acid (AA) to the endoperoxide PGH2. Alternatively, they act like PGE1 to increase platelet cyclic AMP (cAMP). Inhibition of platelet production of thromboxane (TX)A2 may also be desirable. Dihomo-γ-linolenic acid (DHLA) apparently acts through these and other mechanisms. Competition of DHLA with AA for vesicular gland cyclo-oxygenase occurred so that antiaggregatory PGE1 and PGH1 were produced at the expense of PGH2 and PGE2 (which together produce aggregation); O2 uptake was unimpaired. Consistent with such an action in platelets, the antiaggregatory effects of DHLA were prevented by aspirin or other inhibitors of cyclo-oxygenase and were enhanced by Ro 21–1724, a selective inhibitor of cAMP phosphodiesterase. Prior incubation of platelet microsomes with DHLA suppressed conversion of AA to a TXA2-like vasoconstrictor substance and to more stable material behaving like TXB2 on radio-TLC. In addition, DHLA may compete with dietary AA for incorporation into PG precursor pools. The multiple mode of action reported here supports a concept that DHLA is a natural anti-thrombotic factor of key importance.
Assessment of Plasma Fibrinolytic System with Chromogenic Substrate
Authors: Latallo Z. S. , Teisseyre E. , Lopaciuk S.
Published: May 2019

A block of tests employing a chromogenic substrate H-D-Val-Leu-Lys-pNA /S-2251, Kabi, Stockholm/ is proposed for a quantitative assessment of the plasma fibrinolytic system. High sensitivity of the substrate to plasmin makes it possible to assay the enzyme directly in plasma. Its activity remains unchanged during storage of plasma apparently due to the fact that the substrate is also sensitive to plasmin-α2 macroglobulin complex. Introduction of conformational changes in plasminogen by acidification of plasma allows to complete plasminogen conversion into plasmin by streptokinase in 5 min. Both immediate and progressive antiplasmin activity is evaluated upon incubation of plasma with a standard plasmin preparation by measuring the residual enzyme activity /modif. method of Gyzander et al. , Symposium “Neue Methoden der Gerinnungsanalyse”, Freiburg, July, 1976/. Activator activity is expressed as plasmin activity resulting from human or bovine plasminogen after incubation with plasma. The whole set of tests can be performed within 30 min. using only 0.4 ml of plasma. Comparison of the results obtained from plasma samples of patients subjected to either continuous or intermittent treatment with streptokinase /alone or in combination with plasminogen/ indicate that this block of tests allows for a rapid and precise analysis of the pattern of the plasma fibrinolytic system in the course of treatment and may serve as a guidance in establishing the regime to be applied.
Amino Acid Sequence Studies on Human Fibrinogen:Arrangement of Interchain Disulfide Bonds Bounding the Regions of Three-Stranded Ropes
Authors: Doolittle R. F. , Goldbaum D. , Doolittle L. R.
Published: May 2019

Human fibrinogen contains 29 disulfide bonds, only three of which are involved in holding the two dimeric halves of the molecule together. Of the remainder, twelve others are arranged in four sets of three bonds each. Thus, each half of the molecule has two of these unusual arrangements separated by a three-stranded rope consisting of approximately 110 residues in each chain. Alignment of the three non-identical chains at the appropriate cysteine residues participating in these assemblies has revealed not only significant homology but also a rhythmic occurrence of polar and nonpolar amino acids consistent with the existence of coiled α-helices. Indeed, other workers had predicated the existence of such coiled-coils on the basis of fiber diffraction studies a generation ago, hypothesizing that they were likely inter-domainal connections holding the distal portions of a Hall and Slayter-type molecule to the central region. Our data indicate that these predictions were essentially correct. To reinforce the point we have constructed a detailed molecular model of the connecting regions, even to the point where the hypothetical atomic coordinates have been recorded. The model is consistent with virtually all physical data and illuminates details of how molecular packing may occur during formation. It also delineates the geography of bond splitting during fibrinolysis, especially with regard to the boundaries of fragments D and E.
Platelet Phospholipid Changes in Atherosclerosis
Authors: Aktulga A. Z. , Ulutin O. N.
Published: May 2019

The platelet phospholipids content and their per cent distribution in atherosclerotic cases and in thromboembolic cases related to atherosclerosis was investigated with respect to normals. In normals the total phospholipid phosphorus value was found to be 10.88±0.24μg/109 platelets whereas this value changed to 14.29±0.39μg/109 platelets in atherosclerosis. The individual phospholipids follow the decreasing order of phosphatidyl choline, phosphatidyl ethanolamine, sphingomyeline, phosphatidyl serine, phosphatidyl inositol. In atherosclerotic cases although no difference was observed in this order, there was an increase in the quantity of certain phospholipids. In these cases the coagulation mechanism, the fibrinolytic activity, the platelet functions, and the serum lipid contents were also investigated in comparison to the above findings.
Effects of Ethanol on Platelet Serotonin Metabolism
Authors: Cowan D. H. , Shook P.
Published: May 2019

The metabolism of serotonin (5-HT) in platelets is altered in several mental disorders and by various neurotransmitters. Since ethanol impairs the function of both nerves and platelets, the effect of ethanol on 5-HT metabolism was studied in normal platelets and in platelets from patients ingesting ethanol. Ingestion of ethanol by 3 subjects produced blood ethanol levels of 65–76 μM. Platelet MAO activity in each decreased by 50% and platelet serotonin levels increased 50–100%. Ethanol, incubated with normal platelets in vitro, competitively inhibited MAO activity. With phenylethylamine as substrate, the Km of MAO was 23 μM and the Ki of ethanol was 33 mM. With tyramine, Km and Ki were 94 μM and 571 mM, respectively.5-HT uptake by platelets increased 50–100% during ethanol ingestion over control values. By contrast in vitro addition of ethanol to normal platelets inhibited initial 5-HT uptake : Km and Ki were 0.0004 mM and 89.6 mM, respectively. The efflux of 5-HT from platelets was 3–5 times control values during ethanol ingestion and with addition of ethanol to normal platelets. Acetaldehyde at a concentration of 2 mM did not alter MAO activity or 5-HT uptake or efflux. The inhibitory effects of ethanol on MAO and 5-HT uptake were not altered by reserpine, aspirin, or tricyclic antidepressants. The results indicate that ethanol substantially alters 5-HT metabolism by human platelets. Inhibition by ethanol of MAO activity and 5-HT transport may contribute to ethanol-induced platelet and neuronal dysfunction.
Evaluation of Cyclo-Oxygenase Pathway in Platelets of the Newborn
Authors: Corby D. G. , Goad W. C. , Barber J. , O’Barr T. P.
Published: May 2019

A possible deficiency of cyclo-oxygenase in platelets of the newborn infant has been considered as an explanation for their impaired aggregation to stimuli which function by promoting the release of ADP. (Corby and Zuck, Thrombos. and Haemostas., USA 36:201–207, 1976). Cyclo-oxygenase activity was evaluated in washed platelets from paired mother and cord blood samples by monitoring the incorporation of radioactivity into metabolites during incubation with (1-14C) arachidonic acid. Platelets from both the mothers and newborns showed normal aggregation to arachidonic acid. Thin layer radiochromatograms of methylated incubation products were essentially identical. Three main peaks of radioactivity, which corresponded to identified arachidonic acid metabolites, were noted (Malmsten et al. Proc. Natl. Acad. Sei., USA, 72:1446–1450, 1975). Platelets from mothers and newborns incorporated similar amounts of radioactivity into 8-(1-hydroxy-3-oxopropy1)-9,12L-dihydroxy-5-10-heptadecadienoic acid (PHD) and 12L-hydroxy-5,8,10-heptadecatrienoic acid (HHT). Since these two compounds are derived from the endoperoxide prostaglandin G2(PGG2), which is believed to initiate the release reaction, the pathway leading from arachidonic acid to PGG2 is probably fully developed in platelets of the newborn infant. It may be speculated that the lack of response by these platelets to external stimuli is related to either the availability or the formation of metabolically available arachidonic acid.
Evaluation of Highly Purified Bovine F. VIII and Bovine Plasma as Reagents to Induce Platelet Aggregation
Authors: Lazzari M. A. , Simonetti C. , Casillas G. , Pavlovsky M.
Published: May 2019

Purified bovine F. VIII is a known reagent for human platelets aggregation used for the study of thrombopaties. As bovine plasma (BP)is easier to prepare and standardize it was studied as an alternative reagent, comparing it with purified bovine F. VIII. Platelet aggregation was studied by a turbidimetric technique(Aggregometer Bryston AG 1). The substrates were platelet rich plasma (PRP)and gel filtered platelets using between 150,000 and 200,000 platelets/mm3 to standardize the platelets surface. 1/10 dilution of bovine plasma was found to be the best for PRP and 1/1 for filtered platelets. Incubation of the substrates with bovine plasma at 37°C for 5 min seems to enhance PAF activity of the platelet aggregation factor (PAF). Primary aggregation was obtained in PRP treated with 2% tetrasodic EDTA, 3. 4% sodium citrate and in plasma with aspirin. No secondary aggregation was observed in EDTA or aspirin plasma. Unlike platelets treated with ADP, their shape did not change since the optical density was not modified. No synergism or competition between PAF and ADP or adrenalin were found. High concentrations of NaCl or urea interfere with PAF, and esposure to 56°C inactivates it. In 50 normal PRP the percentage of total aggregation were: BP 1/10 42% ± 12 ; BP 1/1 79% ± 13 ;purified bovine F. VIII 1/10 86% ± 6 and in 10 batches of filtered platelets bp 1/1 82% ± 7. We consider an advantage to use bovine plasma for platelets aggregation studies.
Localization of Factor VIII Antigen in Platelets and Megakaryocytes of Normal and Von Willebrand Patients.(Electron Microscopy Studies with Peroxidase Labelled Antibodies)
Authors: Jeanneau C. , Sultan Y. , Savariau E. , Caen J.
Published: May 2019

Using peroxidase coupled antibodies to human factor VIII, factor VIII related antigen (F VIIIRA) was localized in platelets and megakaryocytes of normal and Von Willebrand’s patients.

Megakaryocytes and washed platelets were fixed either in glutaraldehyde alone or in a mixture of tannic acid, paraformaldehyde and glutaraldehyde and then incubated with the peroxidase coupled antibodies. In normal platelets, the peroxidase was found in two locations, on the plasma membrane and surface connecting system (S.C.S.) and in the cytoplasm associated with the membrane complex and dense tubular system.

The plasma membrane of the megakaryocyte and the demarcating membrane system were not labelled. Antigen was localized in the perinuclear space, in smooth reticulum and clear vesicles, known to be the future platelet S. C.S.

In homozygotes with severe forms of Von Willebrand’s disease (VWd) no F VIIIRA was detected around or in the platelete either before of after transfusion of normal cryo or after incubation of their washed platelets in normal plasma. In less severe forms, with detectable F VIIIRA in the plasma there is no such clear correlation between plasma and platelet F VIIIRA. Some have antigen in all platelets with a low plasma level. Others have two populations of platelets some with and others totally lacking antigen. In genetic variants of the disease all the platelets are normally labelled.
A New Primary Fibrinolysin from Mammalian Bile
Authors: King J. B.
Published: May 2019

This primary fibrinolysin, termed cholelysin, has been found in the bile of all mammalian species tested so far. 0x bile has been studied in detail. This was freed of bile salts by adsorption on DEAE sephadex or the anion exchange resin Ag 1-X8, precipitated by ammonium sulphate, redissolved, and eluted off a Whatmans DE 32 column. A further stage of purification on sephadex G 75 was sometimes added, followed by freeze drying.

This purified product gave one or two bands following sodium dodecyl Polyacrylamide gel electrophoresis. These bands contained sharply delimited lytic activity when tested on a standard fibrin plate. Molecular weights were estimated at 10 000 and 20 000 respectively, suggesting the presence of two polymers. Fibrin split products differed from those produced by plasmin.

Cholelysin was actively fibrinolytic on a fibrin plate that had been heated to 80° for 30 minutes, caseinolytic, and active against synthetic substrates such as benzoyl arginine methyl ester. It was inhibited by a substance present at the inter-alpha anti-trypsin position in both human and rat serum. Using a rat model originally developed to test the anticoagulant activity of arvin, intravenous cholelysin was shown to have significant anticoagulant activity.