The direct thrombin inhibitors (argatroban, bivalirudin and lepirudin) and the indirect Xa-inhibitor (danaparoid) increase fibrin network porosity and thus facilitate fibrinolysis

2010 ◽  
Vol 103 (05) ◽  
pp. 1076-1084 ◽  
Author(s):  
Margareta Blombäck ◽  
Niklas Bark ◽  
Hans Johnsson ◽  
N. Hakan Wallen ◽  
Shu He
Keyword(s):  
Dose Response ◽  
Response Curve ◽  
Dose Response Curve ◽  
Thrombin Inhibitors ◽  
Therapeutic Window ◽  
Bleeding Complications ◽  
Direct Thrombin Inhibitors ◽  
Vitro Assay ◽  
Fibrin Network

SummaryThe present study aimed to assess whether the fibrin network structure is modified by the direct thrombin-inhibitors lepirudin, argatroban or bivalirudin and by the indirect Xa-inhibitor danaparoid. Using an in vitro assay that imitates the physiological process of coagulation from thrombin generation to fibrin formation, we examined a normal plasma pool spiked with one of the inhibitors. At concentrations considered to be the plasma levels observed during therapy, almost no influence was detected for lepirudin despite clear-cut effects on “clotting time”. However, argatroban, bivalirudin and danaparoid increased the fibrin gel permeability (Ks) to a similar extent. At concentrations higher than the “therapeutic” levels, the dose-response curve in the Ks assay became very steep for lepirudin while those were shallow for the others. In parallel with the drug-induced increases of Ks, larger network pores in 3D-microscopic images and significant shortenings in “clot lysis time” induced by addition of rtPA were observed. Recombinant factor VIII (rFVIII) added to danaparoid-treated samples profoundly counteracted the increase of Ks but had only a slight or no effect on the other drugs. Thus, in vitro, argatroban, bivalirudin and danaparoid have comparable anticoagulating effects, rendering the fibrin network more permeable and less resistant to fibrinolysis. For lepirudin, the steep dose-response curve supports previous clinical findings, i.e. this thrombin inhibitor has a narrow therapeutic window. Furthermore, our data suggest that the haemostatic agent, rFVIII, might be effective in treatment of bleeding complications induced by danaparoid.

An In Vitro Study To Compare Argatroban with Bivalirudin, Lepirudin and Danaparoid Concerning the Effects on Fibrin Gel Permeability, APTT and INR.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 1886-1886
Author(s):  
Shu He ◽  
Margareta Blomback ◽  
Hans Johnsson ◽  
Hakan Wallen
Keyword(s):  
Dose Response ◽  
Response Curve ◽  
Thrombin Generation ◽  
In Vitro Study ◽  
Dose Response Curve ◽  
Vitro Study ◽  
Therapeutic Window ◽  
Fibrin Gel ◽  
Fibrin Network

Abstract Numerous thrombin/Xa-inhibitors have been used in prophylaxis/treatment of thromboembolic diseases. We were interested to compare argatroban (direct thrombin inhibitor, DTI) with bivalirudin (DTI), lepirudin (DTI) and danaparoid (indirect Xa inhibitor) concerning the effects on fibrin gel permeability (Ks), APTT and INR. We were also interested to assess whether the drug-induced impairment of haemostasis reacts differently to the addition of two coagulants-rVIII or rVIIa. Ks was measured in a normal plasma pool (NPP) mixed with one of the inhibitors in increasing concentrations. Coagulation was initiated by adding recombinant tissue factor (rTF, 5pM), purified phospholipids (PPL, 4mM) and CaCl2. Since the activity of rVIIa is dependent on the presence of activated platelets, rTF and PPL were replaced by washed-frozen platelets when the influence of rVIIa was evaluated. APTT and INR were tested in the same inhibitor-spiked NPP using reagents from Stago, France. We found that all inhibitors investigated made the fibrin network more porous in a concentration-dependent way. However, danaparoid was less potent to change Ks than argatroban or bivalirudin. When a comparison was performed between samples containing argatroban or bivaluridin, the former drug brought increases in Ks which appeared gentler than those by the latter. Regarding the effect by lepirudin, a dose-response curve in Ks assay was very steep. APTT and INR were prolonged by all the inhibitors examined, but only argatroban and bivaluridin led to prolongations which were significantly correlated to Ks. rFVIII overcame the anticoagulant effect on Ks to some degree. Similar influence by rVIIa was seen in samples containing argatroban or bivalirudin but not in those containing lepirudin. The effect of danaparoid on Ks was almost neutralised by addition of the two recombinant coagulants. Conclusions: In the in vitro study, argatroban, bivalirudin, lepirudin and danaparoid can inhibit thrombin generation and thus depress coagulation, shown as prolonged time-to-start of detectable fibrin formation and increased porosity of fibrin network. The latter effect may favour fibrinolysis partly by facilitating the transportation of fibrinolytic components. The steep dose-response curve for Ks shown in the samples containing lepirudin may implicate a narrow therapeutic window. Further work are needed to clarify 1) whether the dissimilarities between argatroban and other inhibitors concerning the effects on Ks, APTT or INR implicate different therapeutic effects or safety in patients; 2) whether the dose-dependent influence of argatroban/bivalirudin on APTT and INR can be shown in vivo, thereby determining if these two simple methods are sensitive enough for therapeutic monitoring; 3) whether rVIII or rVIIa can be used to treat patients with haemorrhagic complications to the therapies. Additionally, the fibrin gel assay which employs a tiny dose of TF (together with PPL) as the thrombin generation trigger to test the end stage of coagulation, i.e. fibrin network formation, may reflect the effects of multiple protease activation and inhibition in a physiologically relevant way. We consider that in contrast to the conventional methods such as APTT and INR, the fibrin gel assay can give additional and important information on drug effects, and thus contribute with important information in the development of new anticoagulant compounds.

Comparison of Activated Partial Thromboplastin Time Ratios with Ecarin Clotting Time Ratios for Monitoring Direct Thrombin Inhibitor Therapy: In-Vitro Study and Case Study of Lepirudin Therapy.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 4149-4149
Author(s):  
Harry L. Messmore ◽  
Nancy J. Fabbrini ◽  
Ketty Badrinath ◽  
Richard Harriman ◽  
Omar Iqbal ◽  
...  
Keyword(s):  
Dose Response ◽  
Partial Thromboplastin Time ◽  
Response Curve ◽  
Activated Partial Thromboplastin Time ◽  
Dose Response Curve ◽  
Thrombin Inhibitors ◽  
Thrombin Inhibitor ◽  
Blood Levels ◽  
Clotting Time

Abstract The direct thrombin inhibitors lepirudin and argatroban are widely used to treat heparin induced thrombocytopenia (HIT). It has been suggested that the Ecarin™ (Echis carinatus venom) clotting time may be superior to the activated partial thromboplastin time (APTT) for monioring purposes. We have prepared standard curves for lepirudin (Refludan™) and Argatroban covering therapeutic drug levels and corresponding APTT ratios (clotting time/control clotting time). Ecarin™ clotting time ratios were performed to demonstrate the practical application of these curves in the clinical care of patients. We report the case of an 80 year old man with HIT/HITT syndrome that occurred during therapy for deep vein thrombosis (DVT) with a low molecular weight heparin (LMWH). His initial coagulation studies were abnormal due to warfarin and LMWH therapy. The patient had a moderate impairment of renal function. Lepirudin therapy in a bolus dose of 14 mg (patient weight: 103.0 kg) resulted in supratherapeutic blood levels of drug and hematuria (Platelet count: 〉200 x 103). Dosage adjustment to maintain an APTT ratio of 1.5 for five days caused no hematuria, but thromboembolic complications occurred at that ratio. The in-vitro dose response curve for Lepirudin was compared with the Ecarin™ clotting time (ECT) ratio at those same concentration ranges in the same plasma. For comparison, Argatroban dose response curves in-vitro were made as well. ECT ratios were very similar to the APTT ratios in the patient’s samples. Representative ratios after the initial bolus, during the infusion period of five days and at the termination of that period are shown in the following table: Comparison of APTT and ECT Ratios APTT Ratio ECT Ratio 1.43 1.07 3.98 3.08 2.91 1.95 2.74 1.95 2.79 1.90 2.58 1.78 2.44 2.42 3.83 4.78 Conclusion: The ECT ratios reflect a steeper dose response curve than that observed with the APTT ratios. This may permit more accurate measurement of blood levels using ECT ratios.

Mechanisms of histamine-induced contraction of canine airway smooth muscle

1983 ◽  
Vol 55 (1) ◽  
pp. 22-26 ◽  
Author(s):  
S. Shore ◽  
C. G. Irvin ◽  
T. Shenkier ◽  
J. G. Martin
Keyword(s):  
Smooth Muscle ◽  
Dose Response ◽  
Response Curve ◽  
Line Tension ◽  
Isometric Tension ◽  
Dose Response Curve ◽  
Base Line ◽  
Histamine Concentration ◽  
Release Of Acetylcholine

We studied the effects of atropine (10(-10) to 10(-6) M), tetrodotoxin (TTX) (10(-6) g/ml), and neostigmine (10(-7) M) on the histamine dose-response curve of canine tracheal smooth muscle (TSM) in vitro. Pretreatment with atropine or TTX reduced base-line tension in some TSM samples, whereas neostigmine invariably caused contraction of TSM. All concentrations of atropine reduced the maximum isometric tension produced by histamine (Tmax). With 10(-6), 10(-8), and 10(-10) M atropine, Tmax was 57, 74, and 88%, respectively, of its value in paired control samples. Atropine, 10(-9) to 10(-6) M, increased the concentration of histamine which produced 20% of Tmax, whereas 10(-6) M also increased the concentration required to produce 50% of Tmax. TTX reduced tension produced by low concentrations of histamine but had no effect at higher concentrations. Neostigmine shifted the histamine dose-response curve and caused greater tension for any given histamine concentration; Tmax increased by 30% (P less than 0.05). Our data are consistent with spontaneous release of acetylcholine from cholinergic nerves in the airway tissue and suggest that histamine either accelerates this release or interacts supra-additively with the acetylcholine at the smooth muscle.

scholarly journals Cd2 Sets Quantitative Thresholds in T Cell Activation

1999 ◽  
Vol 190 (10) ◽  
pp. 1383-1392 ◽  
Author(s):  
Martin F. Bachmann ◽  
Marijke Barner ◽  
Manfred Kopf
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Dose Response ◽  
Response Curve ◽  
Cell Activation ◽  
Dose Response Curve ◽  
Lymphocyte Function

It has been proposed that CD2, which is highly expressed on T cells, serves to enhance T cell–antigen presenting cell (APC) adhesion and costimulate T cell activation. Here we analyzed the role of CD2 using CD2-deficient mice crossed with transgenic mice expressing a T cell receptor specific for lymphocytic choriomeningitis virus (LCMV)-derived peptide p33. We found that absence of CD2 on T cells shifted the p33-specific dose–response curve in vitro by a factor of 3–10. In comparison, stimulation of T cells in the absence of lymphocyte function–associated antigen (LFA)-1–intercellular adhesion molecule (ICAM)-1 interaction shifted the dose–response curve by a factor of 10, whereas absence of both CD2–CD48 and LFA-1–ICAM-1 interactions shifted the response by a factor of ∼100. This indicates that CD2 and LFA-1 facilitate T cell activation additively. T cell activation at low antigen density was blocked at its very first steps, as T cell APC conjugate formation, TCR triggering, and Ca2+ fluxes were affected by the absence of CD2. In vivo, LCMV-specific, CD2-deficient T cells proliferated normally upon infection with live virus but responded in a reduced fashion upon cross-priming. Thus, CD2 sets quantitative thresholds and fine-tunes T cell activation both in vitro and in vivo.

scholarly journals Interim In vitro Dose-Response Curve for the Dicentric Biodosimeter Assay from a Philippine Radiotherapy Facility using a Linear Accelerator

2021 ◽  
Vol 55 (1) ◽  
Author(s):  
Antonio Carlo D. De Guzman ◽  
Carmencita D. Padilla ◽  
Henri Cartier S. Co ◽  
Elrick T. Inocencio ◽  
Edsel Allan G. Salonga
Keyword(s):  
Radiation Exposure ◽  
Dose Response ◽  
Linear Accelerator ◽  
Response Curve ◽  
Dose Response Curve ◽  
Quadratic Model ◽  
Linear Quadratic ◽  
Response Curves ◽  
Energy Agency

Background. Accidental radiation exposure can occur anytime. Biodosimeters help in quantifying the absorbed dose of individuals who are not equipped with personal dosimeters during radiation exposure. The dicentric assay can quantify radiation damage by correlating radiation dose exposure with the frequency of dicentric chromosomes in the peripheral lymphocytes extracted from exposed individuals. Objective. The study aims to present the interim results of the reference dose-response curve for a Philippine radiotherapy facility constructed using a 6MV linear accelerator (ClinacX, Varian). Methods. Samples of peripheral blood from healthy volunteers were irradiated in a customized water phantom of doses 0.10 to 5.0 Gray using a linear accelerator. The irradiated samples were cultured and analyzed following the International Atomic Energy Agency Cytogenetic Dosimetry Protocol (2011) with modifications. Linear-quadratic model curve fitting and further statistical analysis were done using CABAS (Chromosome Aberration Calculation Software Version 2.0) and Dose Estimate (Version 5.2). Interim results of the samples were used to generate these curves. Results. The dose-response curve generated from the preliminary results were comparable to published dose response curves from international cytogenetic laboratories. Conclusion. The generated dose-response calibration curve will be useful for medical triage of the public and radiologic staff accidentally exposed to radiation during medical procedures or in the event of nuclear accidents.

A mathematical approach for the transactivation of hERα

2008 ◽  
Vol 366 (1874) ◽  
pp. 2253-2263 ◽  
Author(s):  
Enrique Castano ◽  
Ruben D Flores-Saaib
Keyword(s):  
Gene Transcription ◽  
Dose Response ◽  
Response Curve ◽  
Bimolecular Reaction ◽  
Dose Response Curve ◽  
Kinetic Properties ◽  
Response Curves ◽  
Transcription System ◽  
Regulated Gene Expression

Several reports over the last few years have documented the dose–response curve for steroid hormone induction of gene transcription as a modulated property of a given receptor–agonist complex that varies with the changing concentration of a variety of factors including: homologous receptor, co-activators, co-repressors and selected co-factors. In each report, the dose–response curves are sigmoidal and show an excellent fit with the curve generated by Michaelis–Menten kinetics. In addition, even the overall function of human oestrogen receptors (hERs) can show a similar graph for the determination of sex versus oestrogen compounds in reptiles. Thus, the kinetic properties of the simple bimolecular reaction of A+B→C appear, surprisingly, to be sufficient to describe the dose–response curve of the multi-step process of steroid-regulated gene induction that involves several molecules. Any advance in explaining why the dose–response curve for steroid-regulated gene expression is sigmoidal would assist in understanding what parameters are key factors of the dose–response curve and can benefit in the design of new oestrogenic substances. We have constructed and analysed a multi-step model of hER-induced gene transcription that explains the multiple forms of a simple dose–response curve in an in vitro transcription system.

scholarly journals THE IN VITRO ACTION OF HYDROCORTISONE ON LEUCOCYTE MIGRATION

1958 ◽  
Vol 107 (2) ◽  
pp. 211-218 ◽  
Author(s):  
Melvin M. Ketchel ◽  
Cutting B. Favour ◽  
Somers H. Sturgis ◽  
Keyword(s):  
Dose Response ◽  
Response Curve ◽  
Dose Response Curve ◽  
Logarithmic Plot ◽  
Leucocyte Migration

Hydrocortisone inhibits the ameboid migration of human leucocytes when added in vitro. The dose-response curve for the reaction between this steroid and leucocytes can be best expressed by a logarithmic plot of the steroid concentrations. Tetrahydrocortisone and desoxycorticosterone had no effect on in vitro leucotyte migration.

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