THE PLATELET “REBOUND-PHENOMENON” DURING PROLONGED, CONTINUOUS PGI2 INFUSION OCCURS AT THE RECEPTOR LEVEL

1987 ◽  
Author(s):  
G Steurer ◽  
H Sinzinger ◽  
P Fitscha
Keyword(s):  
Binding Sites ◽  
Venous Blood ◽  
Intermittent Infusion ◽  
Receptor Level ◽  
Binding Data ◽  
Post Infusion ◽  
Number Of Binding Sites ◽  
Rebound Phenomenon ◽  
Infusion Regimen

During earlier attempts in optimizing the therapeutic regimen with PGI2 we were able to discover an “ intra- and post-infusion platelet rebound” being characterized by an activated platelet function and a diminished responsiveness of platelets to the action of PGI2 in-vitro.In order to verify this phenomenon at the receptor level we infused continuously 6 patients suffering from peripheral vascular disease (PVD) with PGI2 at a rate of 5 ng/kg/min for 5 days. Anticoagulated venous blood has been drawn at different intervals. Saturation binding experiments on platelet membrane fraction have been performed using [3H]iloprost, a stable PGI2 analoque. Analysis of the binding data according to Scatchard demonstrated a decrease of receptor affinity with an increased number of binding sites.It is concluded, that intrainfusion rebound occurs at the receptor level, whereas the postinfusion rebound does not. This is a further piece of evidence that an intermittent infusion regimen is preferable.

Abnormal Platelet Serotonin Uptake and Binding Sites in Myeloproliferative Disorders

1984 ◽  
Vol 51 (03) ◽  
pp. 349-353 ◽  
Author(s):  
C Caranobe ◽  
P Sié ◽  
F Fernandez ◽  
J Pris ◽  
S Moatti ◽  
...  
Keyword(s):  
Binding Sites ◽  
High Capacity ◽  
Specific Inhibitor ◽  
Myeloproliferative Disorders ◽  
Normal Subjects ◽  
Binding Data ◽  
Full Explanation ◽  
Number Of Binding Sites ◽  
5 Ht Uptake ◽  
Kinetics Of

SummaryA simultaneous investigation of the kinetics of serotonin (5 HT) uptake and of binding sites was carried out in the platelets of normal subjects and of 10 patients affected with various types of myeloproliferative disorders (MD). The 5 HT uptake was analysed according to the Lineweaver-Burk and the Eadie-Hofstee methods. With the two methods, the patient’s platelets exhibited a dramatic reduction of the Vi max and of the Km; in some patients the Eadie-Hofstee analysis revealed that a passive diffusion phenomenon is superimposed on the active 5 HT uptake at least for the higher concentration used. The binding data were analysed with the Scatchard method. Two classes of binding sites (high affinity - low capacity, low affinity - high capacity) were found in normal subjects and patients. Pharmacological studies with imipramine, a specific inhibitor of 5 HT uptake, suggested that both the sites are involved in 5 HT uptake. The number of both binding sites was significantly decreased in patient’s platelets while the affinity constants of these binding sites were not significantly reduced in comparison with those of the control subjects. No correlations were found between Vi max, Km and the number of binding sites. These results suggest that a reduction in the number of platelet membrane acceptors for 5 HT commonly occurs in myeloproliferative disorders but does not provide a full explanation of the uptake defect.

The Binding Of 125I-Von Willebrand Factor To Human Platelets In The Presence Of Ristocetin

1981 ◽  
Author(s):  
P Silber ◽  
T H Finlay
Keyword(s):  
Von Willebrand Factor ◽  
Binding Sites ◽  
Binding Constant ◽  
Specific Binding ◽  
Human Platelets ◽  
Scatchard Analysis ◽  
Binding Data ◽  
Number Of Binding Sites ◽  
Von Willebrand ◽  
Willebrand Factor

The effect of ristocetin on the binding of 125I-porcine von Willebrand factor to human platelets was studied. Previously, we had shown that 125I-porcine von Willebrand factor binds to human platelets in the absence of ristocetin. The present work demonstrates that binding is stimulated by ristocetin and this stimulation is maximal at a ristocetin concentration of 2 mg/ml. At a ristocetin concentration of 0.5 mg/ml, Scatchard analysis indicates a binding constant of 5.18 × 10-9M and the presence of 105,000 binding sites. This compares with our previous finding, in the absence of ristocetin, of a binding constant of 2.92 × 10-7M and 4760 binding sites. These binding data assume the porcine von Willebrand factor to be a tetramer with a molecular weight of 9 × 105. This study indicates that ristocetin causes tighter binding and increases the number of binding sites on human platelets for porcine von Willebrand factor. Unlabelled porcine von Willebrand factor competitively inhibits the specific binding of the labelled protein and gives a binding constant of 0.17 × 10-9M. Similar results were obtained using human von Willebrand factor.

Abnormal Platelet Serotonin Uptake And Binding Sites In Myeloproliferative Disorders

1981 ◽  
Author(s):  
C Caranobe ◽  
P Sié ◽  
C Nouvel ◽  
G Laurent ◽  
J Pris ◽  
...  
Keyword(s):  
Binding Sites ◽  
High Capacity ◽  
Myeloproliferative Disorders ◽  
Binding Assays ◽  
Dense Granules ◽  
Plot Analysis ◽  
Binding Data ◽  
Plasmatic Membrane ◽  
Number Of Binding Sites ◽  
5 Ht Uptake

We previously shown that the platelets of patients suffering from myeloproliferative disorders (MD) present not only a reduced capacity to store serotonin (5 HT) in their dense granules but also a dramatic reduction in the initial velocity (Vi) of 5 HT uptake ; this suggests that the abnormalities are not restricted to the dense granules but involve the transport mechanism accross the platelet membrane.The present study concerns the 5 HT binding sites of MD platelets which present such a reduction of the Vi Max (Li- neweaver-Burke plot) of the 5 HT uptake. Binding assays were performed according to the method of Schik et al. (Biochem. Pharmacol. 1979, 28, 2667). Schatchard plot analysis of the binding data revealed two binding sites both in normals and patients : site A with a high affinity and a low capacity and site B with a low affinity and a high capacity.Thus the abnormal 5 HT transport accross the plasmatic membrane is the consequence of a quantitative reduction of the 5 HT binding sites and not of a qualitative defect of these sites. Nevertheless, in spite of the reduction of the number of binding sites, 5 HT-induced platelet aggregation was found normal in these patients.

Effects of hypophysectomy and GH administration on bovine and human GH binding to rat liver membranes

1985 ◽  
Vol 249 (1) ◽  
pp. E56-E62
Author(s):  
J. L. Messina ◽  
S. Eden ◽  
J. L. Kostyo
Keyword(s):  
Binding Sites ◽  
Plasma Membranes ◽  
Specific Binding ◽  
Human Growth ◽  
Male Rats ◽  
Normal Male ◽  
Number Of Binding Sites ◽  
Liver Membranes ◽  
Isolated Liver

Experiments were conducted to investigate the specific binding of highly purified bovine and human growth hormones (bGH and hGH) to purified liver plasma membranes of male rats at various times after hypophysectomy and after the acute intravenous administration of bGH. Liver membranes prepared from hypophysectomized male rats showed a two- to threefold increase in the specific binding of either [125I]iodo-bGH or [125I]iodo-hGH, when compared with membranes prepared from the livers of age-matched normal male rats. The increase in GH binding was apparent within 3 days after hypophysectomy and persisted for a number of weeks after the operation. The increase in GH binding produced by hypophysectomy appeared to be due to an increase in the number of binding sites present on the membranes. The intravenous injection of 200 micrograms of bGH into hypophysectomized male rats 5-60 min before they were killed markedly reduced the ability of liver membranes prepared from these animals to bind [125I]iodo-bGH specifically. This decrease in GH binding seen after the injection of bGH may have been due to the development of a slowly dissociating hormone-binding site complex, which thereby reduced the number of available binding sites. This conclusion is supported by the finding that bGH, which is bound in vitro to isolated liver membranes, dissociates slowly and incompletely in the presence of an excess of unlabeled hormone. Moreover, the degree to which the bound hormone can dissociate appears to depend on the length of time that association is allowed to occur.(ABSTRACT TRUNCATED AT 250 WORDS)

Spectrometric, Thermodynamic, pH Metric and Viscometric Studies on the Binding of TEALS as Surfactant with Albumin as Biopolymer

2020 ◽  
Vol 10 (1) ◽  
pp. 47-64
Author(s):  
Shveta Acharya ◽  
Arun Kumar Sharma
Keyword(s):  
Binding Sites ◽  
Equilibrium Dialysis ◽  
Protein Molecule ◽  
Structure Characterization ◽  
Binding Interaction ◽  
Practical Applications ◽  
Egg Protein ◽  
Binding Data ◽  
Number Of Binding Sites ◽  
Lower Temperature

Background:: Since the interactions of small anions with protein are very important in their transportation and distribution processes in biological systems, it is helpful to study these interactions to understand the nature of the transportation and distribution processes. Therefore, it is aimed to study the interaction of albumin with surfactant molecule by different physical methods. Objective:: Present work attempts to work on assessing the structure, characterization of the surfactants as TEALS (tri-ethanalamine lauryl sulphate) binding sites, with albumin involved in various process of living being are discussed. Method:: The binding of surfactant TEALS to egg protein has been studied at different pH values and temperatures by spectrophotometric and equilibrium dialysis methods. The binding data were found to be pH and temperature dependent. The binding data studied by the absorbance method, were found approximately identical with those obtained from the equilibrium dialysis method. Results:: The association constants and the number of binding sites were calculated from Scatchard plots and found to be at maximum at lower pH and at lower temperature. The free energy of the combining sites was lowest at higher pH and highest at low pH. Therefore, a lower temperature and a lower pH offered more sites in the protein molecule for interaction with surfactant. The ΔG (free energies of aggregation) associated with the binding interaction of the surfactants and protein were calculated. The negative values of the ΔG confirm the feasibility of interaction between the surfactant and protein. All the observations recorded in this paper indicate that the TEALS has a good affinity of binding with egg protein and the number of binding sites is dependent on various physical and chemical factors. Conclusion:: On the basis of the results of the experiments which were conducted to examine the interaction between anionic surfactant and protein by measuring the various parameters of the solutions, it is concluded that the interaction of surfactant and protein gives an idea of fundamental understanding of the structure of surfactant-protein complex and their practical applications in every field.

Unesterified Fatty Acids and the Binding of Tryptophan in Human Plasma

1974 ◽  
Vol 47 (5) ◽  
pp. 415-424 ◽  
Author(s):  
G. Curzon ◽  
June Friedel ◽  
Bharati D. Katamaneni ◽  
Monica H. Greenwood ◽  
M. H. Lader
Keyword(s):  
Human Plasma ◽  
Binding Sites ◽  
Binding Constants ◽  
Nefa Concentration ◽  
Plasma Nefa ◽  
Plasma Tryptophan ◽  
Free Tryptophan ◽  
Number Of Binding Sites ◽  
Albumin Molecule

1. The percentage of human plasma tryptophan in the free state (ultrafilterable) was positively and significantly correlated with plasma non-esterified fatty acid (NEFA) concentration. 2. Results obtained either by adding tryptophan to plasma in vitro or from plasma taken during tryptophan infusion indicate one tryptophan binding site per albumin molecule and an inverse relationship between binding constants and NEFA concentration. 3. A single normal subject was atypical as the apparent number of binding sites varied between one and two per albumin molecule (1.54±0.49 sd, four experiments). 4. Spontaneous increases of plasma NEFA during tryptophan infusion were associated with increased free tryptophan. Also addition in vitro of linoleic acid, oleic acid and palmitic acid to plasma caused free tryptophan to increase. 5. The possible significance of NEFA changes on plasma tryptophan binding in various pathological circumstances is discussed.

Effect of Glycoprotein IIb-IIIa Receptor Antagonism on Platelet Membrane Glycoproteins after Coronary Stent Placement

1998 ◽  
Vol 80 (12) ◽  
pp. 994-1001 ◽  
Author(s):  
Andreas Ruf ◽  
Franz-Josef Neumann ◽  
Gisela Pogátsa-Murray ◽  
Timm Dickfeld ◽  
Dietlind Zohlnhöfer ◽  
...  
Keyword(s):  
Stent Placement ◽  
Binding Sites ◽  
Venous Blood ◽  
Coronary Stent ◽  
Platelet Membrane ◽  
Coronary Stenting ◽  
Receptor Blockade ◽  
Membrane Glycoproteins ◽  
Heparin Group

SummaryPlatelet membrane glycoproteins play a crucial role in ischemic complications after coronary stenting. Glycoprotein IIb-IIIa blockade reduces adverse clinical events after angioplasty but is associated with rare but profound thrombocytopenia that might increase hemorrhagic complications.Changes in platelet membrane glycoproteins of patients with angina who underwent coronary stenting and were treated with the GPIIb-IIIa antagonist abciximab (n = 20) or with heparin (n = 23) were studied. GPIIb-IIIa receptor blockade and membrane glycoproteins were evaluated with immunological markers in venous blood samples taken before, 10, 24, 48, 72, and 96 h after initial treatment with either abciximab or heparin.Patients receiving abciximab therapy showed a rapid inhibition of binding of fluorochrome-conjugated mAb CD41 and c7E3 concomitant with a reduction in platelet aggregation which was restored in part in the days after termination of abciximab infusion. Induction of ligand-induced binding sites on GPIIb-IIIa was increased in patients receiving abciximab. The expression of ligand-induced binding sites correlated inversely with platelet count. No significant change in platelet membrane markers were found in the heparin group. In vitro studies showed that abciximab induces ligand-induced binding sites on isolated platelets and on nuclear cells bearing recombinant GPIIb-IIIa.Abciximab rapidly achieves GPIIb-IIIa receptor blockade after coronary stent placement that might be beneficial in high-risk settings to bridge the delayed action of ticlopidine. Significant alterations of platelet membrane glycoproteins during GPIIb-IIIa antagonism might contribute to development of acute profound thrombocytopenia.

scholarly journals Effect of oxidized glutathione and temperature on inositol 1,4,5-trisphosphate binding in permeabilized hepatocytes

1995 ◽  
Vol 310 (1) ◽  
pp. 185-192 ◽  
Author(s):  
D C Renard-Rooney ◽  
S K Joseph ◽  
M B Seitz ◽  
A P Thomas
Keyword(s):  
Ligand Binding ◽  
Binding Sites ◽  
Binding Activity ◽  
Oxidized Glutathione ◽  
Ip3 Receptors ◽  
Ip3 Receptor ◽  
Temperature Dependent ◽  
Inositol 1,4,5 Trisphosphate ◽  
Number Of Binding Sites

The effect of oxidized glutathione (GSSG) on inositol 1,4,5-trisphosphate (IP3) binding and the activity of IP3-gated Ca2+ channels was examined in permeabilized hepatocytes. The permeability properties of the channel were measured by using Mn2+ quenching of compartmentalized fura-2 at 37 degrees C and at 4 degrees C for comparison with IP3-binding measurements. GSSG (2 mM) increased the IP3-sensitivity of Mn2+ quenching, consistent with previous studies based on Ca(2+)-release measurements [Renard, Seitz and Thomas (1992) Biochem. J. 284, 507-512]. Measurements of [3H]IP3 binding were made at 4 degrees C after preincubation of permeabilized hepatocytes at 37 degrees C in the absence or presence of GSSG. Under these conditions GSSG stimulated IP3 binding by increasing the number of binding sites without changing the Kd. This effect was observed in the absence or presence of Ca2+, but was abolished when the preincubation with GSSG was carried out at 4 degrees C. Thimerosal also stimulated [3H]IP3 binding, but this effect was mediated both by an increase in the maximum number of binding sites and by a decrease in the Kd. The effects of thimerosal and GSSG were not additive. Further analysis of the effect of GSSG revealed that preincubation of permeabilized hepatocytes at 37 degrees C results in a progressive loss of [3H]IP3-binding sites that can be prevented and reversed by inclusion of GSSG. A parallel loss of IP3-sensitive Mn(2+)-quenchable stores was observed after incubation at 37 degrees C, and this could also be reversed by adding back GSSG. The loss of IP3 binding was not the result of IP3-receptor proteolysis, as judged by Western blotting of immunoreactive protein. The sensitivity of [3H]IP3 binding in permeabilized hepatocytes to varied ratios of GSSG and GSH suggests that the IP3 receptor responds to an oxidized redox environment such as that found in the lumen of the endoplasmic reticulum. GSSG had no direct effect on the ligand-binding activity of detergent-solubilized and partially purified IP3 receptors. We conclude that GSSG exerts an indirect effect on the IP3 receptors in permeabilized hepatocytes by preventing a temperature-dependent loss of IP3-binding sites. We suggest that the hepatic IP3 receptors may interact with a thiol-disulphide oxidoreductase that utilizes GSSG as a substrate and prevents inappropriate unfolding of the ligand-binding domain occurring after incubation of the receptor at 37 degrees C in vitro.

In vitro study of the binding between chlorpyrfos and sex hormones using headspace solid-phase microextraction combined with high-performance liquid chromatography

2015 ◽  
Vol 34 (8) ◽  
pp. 819-827 ◽  
Author(s):  
K Farhadi ◽  
R Tahmasebi ◽  
P Biparva ◽  
R Maleki
Keyword(s):  
High Performance Liquid Chromatography ◽  
Liquid Chromatography ◽  
Sex Hormones ◽  
Binding Sites ◽  
Solid Phase Microextraction ◽  
High Performance ◽  
Solid Phase ◽  
Hplc Method ◽  
Number Of Binding Sites

Endocrine-disrupting chemicals are compounds that alter the normal functioning of the endocrine system. Organophosphorus insecticides, as chlorpyrifos (CPS), receive an increasing consideration as potential endocrine disrupters. Physiological estrogens, including estrone (E1), 17β-estradiol (E2), and diethylstilbestrol (DES) fluctuate with life stage, suggesting specific roles for them in biological and disease processes. There has been great interest in whether certain organophosphorus pesticides can affect the risk of breast cancer. An understanding of the interaction processes is the key to describe the fate of CPS in biological media. The objectives of this study were to evaluate total, bound, and freely dissolved amount of CPS in the presence of three estrogenic sex hormones (ESHs). In vitro experiments were conducted utilizing a headspace solid phase microextraction (HS-SPME) combined with high-performance liquid chromatography (HPLC) method. The obtained Scatchard plot based on the proposed SPME-HPLC method was employed to determine CPS-ESHs binding constant and the number of binding sites as well as binding percentage of each hormone to CPS. The number of binding sites per studied hormone molecule was 1.10, 1, and 0.81 for E1, E2, and DES, respectively. The obtained results confirmed that CPS bound to one class of binding sites on sex hormones.

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