scholarly journals Effect of Chylomicrons on the Fibrinolytic Activity of Normal Human Plasma in Vitro

1959 ◽  
Vol 7 (2) ◽  
pp. 205-209 ◽  
Author(s):  
THOMAS C. MERIGAN ◽  
JOHN W. FARQUHAR ◽  
JAMES H. WILLIAMS ◽  
MAURICE SOKOLOW
Keyword(s):  
Human Plasma ◽  
Fibrinolytic Activity ◽  
Normal Human Plasma ◽  
Normal Human

Presence and Possible Origin of ɛ(γ-Glutamyl)Lysine Isodipeptide in Human Plasma

1992 ◽  
Vol 67 (01) ◽  
pp. 060-062 ◽  
Author(s):  
J Harsfalvi ◽  
E Tarcsa ◽  
M Udvardy ◽  
G Zajka ◽  
T Szarvas ◽  
...  
Keyword(s):  
Human Plasma ◽  
Fibrinolytic Therapy ◽  
Normal Human Plasma ◽  
Normal Human ◽  
Hplc Technique ◽  
Significant Change

Summaryɛ(γ-glutamyl)lysine isodipeptide has been detected in normal human plasma by a sensitive HPLC technique in a concentration of 1.9-3.6 μmol/1. Incubation of in vitro clotted plasma at 37° C for 12 h resulted in an increased amount of isodipeptide, and there was no further significant change when streptokinase was also present. Increased in vivo isodipeptide concentrations were also observed in hypercoagulable states and during fibrinolytic therapy.

In Vitro Effects of the Acylated StreptokinasePlasminogen Activator Complex BRL 33 575 Incubated with Normal Human Plasma

1984 ◽  
Vol 51 (03) ◽  
pp. 403-405 ◽  
Author(s):  
B Lämmle ◽  
G Noll ◽  
T H Tran ◽  
A Lohri ◽  
F Duckert
Keyword(s):  
Human Plasma ◽  
Clinical Studies ◽  
Systemic Effects ◽  
In Vitro Experiments ◽  
Normal Human Plasma ◽  
Normal Human ◽  
Repeated Doses ◽  
In Vitro Effects ◽  
Activator Complex

SummaryThrombolysis with acylated streptokinase-plasminogen complexes is aimed to achieve fibrinolysis without systemic fibrinogenolysis. The p-aminobenzoyl-streptokinase-(Lys)-plasminogen-complex (BRL 33 575) should be particularly useful due to its slow deacylation rate. Unexpectedly, repeated doses of 10 mg of BRL 33 575 (corresponding to 310'000 streptokinase equivalent units) induced systemic effects in patients though less than streptokinase alone. In vitro incubation of normal human plasma with BRL 33 575 at concentrations used in patients resulted in nearly complete consumption of α2-antiplasmin and plasminogen and significant fibrinogenolysis within 3 hr. This demonstrates that - despite of slow deacylation of BRL 33 575 - the small amounts of activator generated are highly efficacious in activating plasma plasminogen under conditions in which no physiological clearance of the free activator takes place. Simulating the calculated activator release from BRL 33 575 by infusing equivalent amounts of streptokinase into plasma resulted in less pronounced effects. This is probably explained by anti-streptokinase antibodies which will neutralize the initially infused streptokinase but will be bound by BRL 33 575.Our in vitro experiments indicate that further clinical studies should be done with lower doses of BRL 33 575 or prolonged dosage intervals.

AN INSULIN ASSAY BASED ON THE INCORPORATION OF LABELLED GLYCINE INTO PROTEIN OF ISOLATED RAT DIAPHRAGM

1959 ◽  
Vol 19 (3) ◽  
pp. 259-262 ◽  
Author(s):  
K. L. MANCHESTER ◽  
P. J. RANDLE ◽  
F. G. YOUNG
Keyword(s):  
Human Plasma ◽  
Plasma Samples ◽  
Rat Diaphragm ◽  
Normal Human Plasma ◽  
Advantages And Disadvantages ◽  
Straight Line ◽  
Normal Human ◽  
Isolated Rat

SUMMARY Insulin increases the incorporation of [14C]glycine into the protein of isolated rat diaphragm in vitro. This effect of the hormone can be used as the basis of a straight-line assay for insulin. The advantages and disadvantages of such a method of assay are discussed. Normal human plasma also stimulates incorporation of glycine into diaphragm protein, and the activities of two plasma samples were equivalent to 2 and 10 mu. insulin/ml. plasma, respectively.

scholarly journals PRESERVATION OF NORMAL HUMAN PLASMA IN THE LIQUID STATE. II. COMPARATIVE IN VITRO STUDIES ON THE PHYSIOLOGIC ACTIVITY OF LABILE CONSTITUENTS OF LIQUID AND FROZEN PLASMA 12

1944 ◽  
Vol 23 (3) ◽  
pp. 351-356 ◽  
Author(s):  
F. H. L. Taylor ◽  
Eugene L. Lozner ◽  
C. S. Davidson ◽  
H. J. Tagnon ◽  
Lloyd R. Newhouser ◽  
...  
Keyword(s):  
Human Plasma ◽  
Liquid State ◽  
In Vitro Studies ◽  
Normal Human Plasma ◽  
Normal Human ◽  
Physiologic Activity ◽  
Frozen Plasma

Effects in vitro of bovine and human antihemophilic globulins on substrate plasmas

1964 ◽  
Vol 206 (1) ◽  
pp. 74-78
Author(s):  
Philip H. Geisler ◽  
Mary F. Eichman ◽  
Leandro M. Tocantins
Keyword(s):  
Human Plasma ◽  
Normal Saline ◽  
Normal Human Plasma ◽  
Bovine Plasma ◽  
Normal Human ◽  
Species Specific

Greater clot-promoting activity has been claimed for antihemophilic globulin ( AHG) preparations from bovine plasma than for those from normal human plasma. This has been attributed to an actual higher amount of the globulin in bovine plasma. A comparison was made of the effects of bovine and human antihemophilic globulins, collected and prepared in the same manner, on the rate of clotting of not only normal and hemophilic human plasmas but on bovine plasma as well. AHG was prepared from citrated plasma by dilution, acidification, and solution of the washed precipitate in normal saline. While bovine AHG is more effective than human AHG in accelerating the clotting of normal and hemophilic human plasmas, human AHG has a greater effect on bovine plasma than does bovine AHG. Moderate dilution of the plasma substrate enhances the clot-accelerating action of the homologous AHG. The response of a plasma substrate to AHG preparations suggests the presence in plasma of species-specific antagonists which reduce the clot-promoting activity of the homologous AHG, but which are less effective against preparations from heterologous sources.

An assay of the capacity of biological fluids to stimulate renal glucose-6-phosphate dehydrogenase activity in vitro as a marker of their ability to inhibit sodium potassium-dependent adenosine triphosphatase activity

1982 ◽  
Vol 94 (1) ◽  
pp. 99-NP ◽  
Author(s):  
S. Fenton ◽  
E. Clarkson ◽  
G. MacGregor ◽  
J. Alaghband-Zadeh ◽  
H. E. de Wardener
Keyword(s):  
Human Plasma ◽  
Adenosine Triphosphatase ◽  
Sodium Intake ◽  
Active Material ◽  
G6pd Activity ◽  
Normal Human Plasma ◽  
Sodium Potassium ◽  
Normal Human ◽  
Glucose 6 Phosphate Dehydrogenase

A highly sensitive cytochemical method for the assay of the ability of plasma and extracts of human urine to stimulate renal glucose-6-phosphate dehydrogenase (G6PD) activity in vitro is described. In the proximal convoluted tubules there was a linear increase of G6PD activity with the logarithm of concentration of a highly purified natriuretic extract from normal human urine (0·384–384 ng active material/l) which was used as a standard. The stimulation of G6PD obtained with dilutions of normal human plasma was parallel to that produced by the standard. The sensitivity of the assay permitted the measurement of as little as 0·384 ng active material/l of the natriuretic extract (0·001 units/ml) and dilutions of 1/10 000 could be detected using normal human plasma. The mean ± s.e.m. index of precision was 0·068± 0·003 (n = 9). It is known that inhibition of sodium potassium-dependent adenosine triphosphatase (Na+-K+-ATPase) is associated with a rise in G6PD activity. We have confirmed this observation by demonstrating that ouabain, a potent inhibitor of Na+ -K+-ATPase, stimulates renal G6PD activity in our assay and that natriuretic extract, human plasma and ouabain stimulated renal G6PD activity in vitro and simultaneously inhibited renal Na+-K+-ATPase activity in vitro. The plasma from 12 normal subjects (five of whom were previously shown to inhibit renal Na+-K+-ATPase activity in vitro in a manner related to sodium intake) stimulated renal G6PD activity in vitro, and this activity was also directly related to sodium intake. It is suggested that the change in the capacity of plasma to stimulate renal G6PD activity in vitro is a marker of the concentration of a circulating sodium transport inhibitor.

Kinetics of Human VWF Cleavage by Human ADAMTS13: Comparison of Recombinant and Plasma-Derived VWF

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 4092-4092
Author(s):  
Hanspeter Rottensteiner ◽  
Katalin Varadi ◽  
Susanne Vejda ◽  
Jutta Schreiner ◽  
Herbert Gritsch ◽  
...  
Keyword(s):  
Plasma Concentration ◽  
Human Plasma ◽  
Degradation Kinetics ◽  
Dependent Manner ◽  
Normal Human Plasma ◽  
Normal Plasma ◽  
Normal Human ◽  
Typical Satellite ◽  
Kinetics Of

Abstract A recombinant human CHO-expressed von Willebrand factor (rVWF) which consists of ultra-high molecular weight (UHMW) multimers resembles the VWF that is stored in Weibel-Palade bodies of endothelial cells. Once secreted into plasma, UHMW multimers are rapidly cleaved by ADAMTS13 and therefore are usually missing in VWF purified from plasma. We analyzed in vitro whether the cleavage of rVWF by ADAMTS13 is similar to the cleavage of plasma VWF by ADAMTS13, using a standard assay that depends on denaturing conditions (1.5 M urea) to expose the ADAMTS13 cleavage site of VWF. We explored the kinetics of ADAMTS13-mediated proteolysis of rVWF over time by exposing 1 VWF:Ag IU/ml of rVWF to various concentrations of recombinant and plasma-derived purified ADAMTS13, ranging from 4 mU/ml to 1 U/ml (corresponding to 0.4 to 100% of the normal human plasma concentration), and to ADAMTS13 present in normal human plasma and VWF-deficient plasma. The multimeric structure and function of VWF were analyzed by multiple assays to compare VWF before, during, and after proteolytic cleavage. In addition, the degradation kinetics of recombinant VWF were compared with those of a plasma-derived VWF product. Recombinant VWF was cleaved rapidly and with the same efficiency using recombinant or plasma-derived ADAMTS13. With 0.5 U/ml pADAMTS13 or rADAMTS13, VWF:RCo activity was below the detection limit of 0.17 IU/ml after 15 s. UHMW multimers disappeared within seconds at physiological concentrations of 0.5–1.0 U/ml ADAMTS13 (50–100% of the normal plasma concentration). Furthermore, UHMW were cleaved within 30 minutes with much lower concentrations of 10–30 mU/ml ADAMTS13 (1–3% of normal plasma concentration). The typical satellite bands appeared very early in an ADAMTS13 dose-dependent manner. Although plasma-derived VWF differs substantially from rVWF in its multimeric structure, the decrease in activity was similar for the recombinant and plasma-derived VWF. Specific cleavage of rVWF (3 IU VWF:Ag/ml) by rADAMTS13 (5 U/ml) was also demonstrated without urea under shear stress in the presence of platelets, detected by a monoclonal antibody (N10) that recognizes VWF only when cleaved by ADAMTS13. The combined data show that ADAMTS13 is able to readily cleave human rVWF even at low concentrations and that the UHMW multimeric fraction of human rVWF is removed within minutes by ADAMTS13 in vitro.

Zymogen-Like FXa Is An Effective Pro-Hemostatic To Reverse The Anticoagulant Effects Of Direct FXa Inhibitors

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 2383-2383
Author(s):  
Nabil K Thalji ◽  
Sunita Patel-Hett ◽  
Reema Jasuja ◽  
Joachim Fruebis ◽  
Debra Pittman ◽  
...  
Keyword(s):  
Blood Loss ◽  
Human Plasma ◽  
Active Site ◽  
Half Life ◽  
Thrombin Generation ◽  
Fold Increase ◽  
Normal Human Plasma ◽  
Normal Human

Abstract Oral anticoagulants are the mainstay of treatment for prothrombotic disorders. The emerging oral factor Xa (FXa) inhibitors, which include rivaroxaban and apixaban, have been shown to be highly effective anticoagulants in several clinical scenarios, including venous thromboembolism and non-valvular atrial fibrillation. Compared to warfarin, direct FXa inhibitors have less variable pharmacokinetics, may not require routine monitoring of coagulation parameters, and have comparable to a somewhat lower bleeding risk. Despite these advantages, no approved strategy has been developed to reverse the anticoagulant effects of these drugs in the event of life-threatening bleeding or emergent need for surgery. This represents an urgent unmet clinical need. Our group has recently developed a panel of FXa mutants that are more zymogen-like than wild-type (wt)-FXa. These “zymogen-like” FXa variants have lower activity in in vitro assays compared to wt-FXa due to impaired active site maturation. Furthermore, the variants have longer plasma half-lives (>30 minutes) in vitro compared to wt-FXa (1-2 minutes) due to diminished reactivity with antithrombin III (ATIII) and tissue factor pathway inhibitor (TFPI). Remarkably however, binding to FVa rescues the activity of these zymogen-like FXa variants and as a result they are highly effective procoagulants in vivo in the setting of hemophilia (Nat. Biotech; 2011, 29:1028-33). We hypothesized that these variants could also be effective procoagulants to overcome the effects of direct FXa inhibitors. Furthermore, since direct FXa inhibitors bind the FXa active site, we expect them to compete with ATIII and TFPI for FXa binding and prolong their half-lives. We tested both of these hypotheses in in vitro coagulation studies and in vivo hemostasis models. Rivaroxaban dose-dependently inhibited thrombin generation in thrombin generation assays (TGA) when added to normal human plasma. Specifically, 500 nM rivaroxaban, the expected therapeutic steady-state plasma concentration, decreased peak thrombin generation to ∼10% of normal, and addition of 3 nM of the FXa zymogen-like variant FXaI16L restored peak thrombin generation to 105% of normal. Higher concentrations of rivaroxaban (2.5 µM) completely abrogated thrombin generation in this assay, but 10 nM FXaI16L restored thrombin generation to 72% of normal under these conditions. We compared these data to results obtained with other proposed reversal strategies. Gla-domainless, catalytically inactive FXa (GD-FXaS195A), which has been shown to reverse the effects of rivaroxaban by scavenging the inhibitor, restored thrombin generation in the presence of 500 nM rivaroxaban, but required high concentrations (1 µM; >300-fold greater than FXaI16L) to be effective. In addition, activated prothrombin complex concentrates (FEIBA), which have been shown to have some ex vivo efficacy, were ineffective under our assay conditions. In tail-clip hemostasis studies in mice, rivaroxaban dose-dependently increased blood loss, with 50 mg/kg rivaroxaban resulting in 217% of normal blood loss. Addition of FXaI16L (200 mg/kg) reduced rivaroxaban-induced blood loss to 141% of normal. To examine the effect of rivaroxaban on the half-life of FXa, we pre-incubated FXaI16L or wt-FXa with or without rivaroxaban in normal human plasma and then performed TGA experiments after various incubation times. When wt-FXa or FXaI16L were pre-incubated in plasma in the absence of rivaroxaban, their half-lives were 4.6 minutes and 1.37 hours, respectively. Remarkably, when wt-FXa or FXaI16L were incubated in plasma in the presence of 500 nM rivaroxaban, their respective half-lives were prolonged to 9.4 hours (123-fold increase) and 18.1 hours (13.2-fold increase). These results suggest that a zymogen-like FXa variant, FXaI16L, can reverse the effects of rivaroxaban in vitro and in vivo. Furthermore, FXaI16L is a bypassing agent that only requires catalytic amounts of protein, in contrast to scavengers or “true” antidotes like GD-FXaS195A that require stoichiometric concentrations. This indicates that much lower quantities of FXaI16L may be effective in vivo. We also showed that rivaroxaban dramatically prolongs the half-life of FXa in plasma, possibly by competing with ATIII and TFPI for FXa binding. This work provides a starting point for the development of a long half-life reversal strategy for the emerging FXa inhibitors. Disclosures: Patel-Hett: Pfizer: Employment. Jasuja:Pfizer: Employment. Fruebis:Pfizer: Employment. Pittman:Pfizer: Employment. Camire:Pfizer: Consultancy, Patents & Royalties, Research Funding; Alnylam: Consultancy.

Performances of an Artificial Reagent for the One-stage Factor IX Assay

1975 ◽  
Vol 33 (03) ◽  
pp. 547-552 ◽  
Author(s):  
L Meunier ◽  
J. P Allain ◽  
D Frommel
Keyword(s):  
Human Plasma ◽  
Factor Ix ◽  
Severe Haemophilia ◽  
Normal Human Plasma ◽  
Haemophilia B ◽  
One Stage ◽  
Normal Human ◽  
The One

SummaryA mixture of adsorbed normal human plasma and chicken plasma was prepared as reagent for factor IX measurement using a one-stage method. The substrate was found to be specific for factor IX. Its performances tested on samples displaying factor IX activity ranging from <l%–2,500% compared favorably with those obtained when using the plasma of severe haemophilia B patients as substrate.

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