scholarly journals Effects of Lectins on Blood Platelets from Various Species

1977 ◽  
Author(s):  
O. Tangen ◽  
B. Karlstam ◽  
S. Bygdeman
Keyword(s):  
Shape Change ◽  
Blood Platelets ◽  
Human Platelets ◽  
Serotonin Release ◽  
Variable Degree ◽  
Con A ◽  
Platelet Release ◽  
Induced Aggregation ◽  
Aggregation Of Platelets ◽  
Platelet Shape

Earlier it has been shown that different lectins induce a variable degree of aggregation of platelets. The present study confirmed previous data and demonstrated that wheat germ agglutinin (WGA) was very active, 1eucoagglutinin had about a tenth of the activity of WGA on a concentration basis, and Con A had a weak aggregating effect on human gel filtered platelets (GFP). Soy bean lectin did not aggregate human GFP.The fact that adenosine inhibited WGA- and leucoagglutinin-induced aggregation that WGA and Con A caused serotonin release, and that the aggregation- curves indicated platelet shape change are indications that the lectins influenced glycosyl moieties involving one or more molecules relevant to release and aggregation reaction.GFP were markedly more responsive to the lectins than platelets in plasma, probably due to interfering glycosyl groups amongst the plasma constituents.Platelets from man, rabbit, rat, cow and pig reacted differently towards the lectins, human platelets being the most reactive and bovine and porcine platelets being almost unreactive. These results pose intriguing questions regarding the glycosyl content of platelet membranes in different species and their relation to platelet release and aggregation.

Platelet Aggregation Caused by Dithiothreitol

1984 ◽  
Vol 51 (01) ◽  
pp. 119-124 ◽  
Author(s):  
M B Zucker ◽  
N C Masiello
Keyword(s):  
Shape Change ◽  
Blood Platelets ◽  
Dense Granule ◽  
Metabolic Inhibitors ◽  
Electrophoretic Patterns ◽  
Discernible Effect ◽  
Variable Extent ◽  
Triton X ◽  
Induced Aggregation ◽  
Platelet Shape

SummaryMacIntyre et al. showed that over 1 mM dithiothreitol (DTT) aggregates blood platelets in the presence of fibrinogen; aggregation is not inhibited by prostaglandin E1. We confirmed their data and found that 70 mM 2-mercaptoethanol was also active. DTT- induced aggregation was not associated with platelet shape change or secretion of dense granule contents, was not inhibited by tetracaine or metabolic inhibitors, was prevented at pH 6.5, and prevented, reversed, or arrested by EDTA, depending on when the EDTA was added. DTT did not cause aggregation of thrombasthenic, EDTA-treated, or cold (0° C) platelets, which also failed to aggregate with ADP. Platelets stimulated with DTT bound 125I-labeled fibrinogen. Thus DTT appears to “expose” the fibrinogen receptors. SDS gel electrophoresis of platelet fractions prepared by use of Triton X-114 showed that aggregating concentrations of DTT reduced proteins of apparent Mr 69,000 and 52,000 (probably platelet albumin) and, to a variable extent, glycoproteins Ib, IIb and III. Exposure of unlabeled or 125I- labeled platelets to ADP had no discernible effect on the electrophoretic patterns.

IRREVERSIBLE BLOCKADE OF THE THROMBOXANE A2/PROSTAGLANDIN H2 RECEPTOR OF HUMAN PLATELETS BY AZIDO-BSP

1987 ◽  
Author(s):  
H Zehender ◽  
E C Witte ◽  
K Stegemeier ◽  
A Patscheke
Keyword(s):  
Shape Change ◽  
Thromboxane A2 ◽  
High Specificity ◽  
Short Wave ◽  
Human Platelets ◽  
Serotonin Release ◽  
Irreversible Inhibition ◽  
High Concentrations ◽  
Very High ◽  
Platelet Shape

Azido-BSP (4-[2-(4-azido-benzenesulphonylamino)-ethyl]phen-oxyacetic acid) is a photolabile derivative of the competitive thromboxane A2 /prostaglandin H2 (TXA2/PGH2) receptor antagonist sulotroban (=BM 13.177). If protected from short wave light, azido-BSP reversibly inhibited the platelet shape change induced by the PGH2 analogue U 46619 but notthe shape change induced by ADP or PAF. Schild analysis revealed an apparent KD=0.2 μM with washed platelets. The irreversible inhibition requiredirradiation of the platelet suspensionwith UVlight (254 nm) for 5 minutes in the presenceof azido-BSP. After this treatment,the platelets were washed twice and used forplatelet function tests. Treatment with 0.5 μM of azido-BSP suppressed the U 46619(10 μM)-induced (3H)serotonin release and 1 μM of azido-BSP was necessary to block the U 46619(2 μM)-inducedaggregation.The platelet shape change induced by U 46619 (0.01μM) was only partially inhibited, even at very high concentrations (50μM) of the antagonist.This suggests that a small portion of the TXA2/PGH2 receptors could not be blocked bythe photoaffinity treatment with azido-BSP. After treatment with 1 μM azido-BSP, the shape change stimulated by ADP or PAF was not reduced. This indicates a high specificity of thephotoaffinity ligand for the TXA2/PGH2 receptor. It is concluded that UV irradiation of azido-BSP generates anitrene intermediate that covalently links to the majority of the TXA2/PGH2 receptors. Azido-BSP provides a specific tool for tagging and subsequent purification of the TXA2/PGH2 receptor of platelets.(Supported by the Deutsche Forschungsgemeinschaft, Grant Pa263).

Concanavalin A-induced Aggregation of Human Blood Platelets

1975 ◽  
Vol 33 (02) ◽  
pp. 354-360 ◽  
Author(s):  
Heinrich Patscheke ◽  
Reinhard Brossmer
Keyword(s):  
Concanavalin A ◽  
Binding Capacity ◽  
Specific Binding ◽  
Blood Platelets ◽  
Residual Effect ◽  
Independent Effect ◽  
Con A ◽  
Experimental Conditions ◽  
Main Effect ◽  
Induced Aggregation

SummaryConcanavalin A (CON A) causes platelets to aggregate. A Ca++-independent effect of CON A could be separated from a main effect which depends on Ca++. The main effect probably is a consequence of the CON A-induced platelet release reaction and therefore is platelet-specific. The weak residual effect observed in the presence of Na2EDTA may be due to a similar mechanism as has been demonstrated for CON A-induced aggregations of several other normal and malignant transformed animal cells.Na2EDTA did not inhibit the carbohydrate-specific binding capacity of CON A. Therefore, Na2EDTA appears not to demineralize the CON A molecules under these experimental conditions.α-methyl-D-glucoside inhibits the Ca++-independent as well as the Ca++-dependent effect of CON A.Pretreatment by neuraminidase stimulated the platelet aggregation induced by CON A. It is possible that removal of terminal sialic acid residues makes additional receptors accessible for the binding of CON A.

Physiological Effects of Nonimmune Platelet Associated Immunoglobulin G

1981 ◽  
Vol 45 (01) ◽  
pp. 027-033 ◽  
Author(s):  
K Sugiura ◽  
M Steiner ◽  
M Baldini
Keyword(s):  
Shape Change ◽  
Fluorescein Isothiocyanate ◽  
Recovery Phase ◽  
Human Platelets ◽  
Serotonin Release ◽  
Platelet Volume ◽  
Igg Antibody ◽  
Heterologous Antiserum ◽  
Igg Binding ◽  
Series Of Experiments

SummaryThe function of nonimmune IgG associated with platelets is unknown. In a series of experiments we have investigated this problem, relating amount of platelet-associated IgG (PAIgG) to platelet volume, serotonin release, adherence of platelets to monocytes and platelet senescence. Most of these studies were performed with human platelets. Platelets freed of preexisting PAIgG by incubation at 22° C were incubated with IgG in a series of concentrations ranging from 0.4 — 27.0 X10-6 M. The IgG preparations used were demonstrably free of aggregated forms of the protein. The amount of PAIgG bound to platelets was determined by the use of fluorescein isothiocyanate-conjugated anti-IgG antibody (F-anti-IgG antibody) which was quantified in a fluorospectrophotometer. Newly bound IgG was assayed similarly by the use of F-IgG. A dose-dependent increase in platelet volume was associated with the binding of nonimmune IgG by platelets. The process which leveled off at an IgG concentration of 1.2 —1.5 X10-5 M was almost fully reversible and was not due to platelet shape change or aggregation. Release of serotonin from IgG-treated platelets was relatively small but to the extent that it occurred was positively related to the IgG concentration to which platelets were exposed. Adherence to autologous monocytes studied quantitatively by the use of formaldehyde-fixed cells was also positively related to the amount of IgG on the platelets. Normal or IgG-defident serum had a potent inhibitory (noncompetitive) action on the binding of F-IgG and F-anti-human IgG antibody to human platelets. Cohorts of platelets prepared in rabbits during the recovery phase of immunological thrombocytopenia induced by injection of heterologous antiserum, showed an age-dependent increase of PAIgG and of IgG binding. These results suggest that PAIgG plays a role in the clearance of senescent platelets.

A Class of Drugs that Inhibits Platelet Release and Aggregation by Apparent Interference with Anion Transport

1979 ◽  
Author(s):  
N.R. Shulman ◽  
H.B. Pollard ◽  
K. Tack-Goldman
Keyword(s):  
Pyridoxal Phosphate ◽  
Human Platelets ◽  
Anion Transport ◽  
Serotonin Release ◽  
Secretory Cells ◽  
Chloride Uptake ◽  
Potential Alternative ◽  
Osmotic Lysis ◽  
Platelet Release ◽  
Ph Range

The platelet release reaction is analogous to the process of exocytosis by which many other secretory cells release hormones or mediators from intracellular granules. Anion transport blocking (ATB) drugs Inhibit release of epinephrine from isolated chromaffin granules (CG) by blocking chloride uptake and preventing osmotic lysis. Studies on platelets analagous to those done on CG showed that increasing osmotic strength in the range 600-1000 m0sm progressively suppressed serotonin release to completion and that ATB drugs (viz, probenecid, SITS, pyridoxal phosphate and suramin) at mM concentrations completely inhibited release and aggregation of human platelets stimulated by thrombin, ADP, A23187, epinephrine or collagen. Sulfinpyrazone has the appropriate structure for anionic blocking, and may suppress platelet function as effectively by this mechanism as by cycloxy-genase inhibition. The ATB drugs acted apparently to prevent movement of OH- from the more alkaline medium into the relatively acidic granule, for platelet release was not inhibited by replacing anions with isethionate or sucrose, but was markedly dependent on OH- in the pH range 6 to 7.5 where inhibition by the ATB drugs was competitive with respect to OH-. Since the ATB compounds include some relatively nontoxic drugs in common use, and since their action on platelets differs markedly from that of aspirin, they should receive attention as potential alternative or auxiliary antithrombotic agents.

scholarly journals Transformation and motility of human platelets: details of the shape change and release reaction observed by optical and electron microscopy.

1979 ◽  
Vol 83 (1) ◽  
pp. 126-142 ◽  
Author(s):  
R D Allen ◽  
L R Zacharski ◽  
S T Widirstky ◽  
R Rosenstein ◽  
L M Zaitlin ◽  
...  
Keyword(s):  
Shape Change ◽  
Human Subjects ◽  
Blood Platelets ◽  
Time Lapse ◽  
Human Platelets ◽  
Transformation Process ◽  
Epifluorescence Microscopy ◽  
Scanning Electron Micrographs ◽  
Dense Bodies ◽  
Short Wavelength Light

Blood platelets from 10 normal human subjects have been examined with a sensitive differential interference contrast (DIC) microscope. The entire transformation process during adhesion to glass is clearly visible and has been recorded cinematographically, including the disk to sphere change of shape, the formation of sessile protuberances, the extension and retraction of pseudopodia, and the spreading, ruffling, and occasional regression of the hyalomere. The exocytosis of intact dense bodies can be observed either by DIC microscopy, or by epifluorescence microscopy in platelets stained with mepacrine. Details of fluorescent flashes indicate that the dense bodies usually release their contents extracellularly, may do so intracytoplasmically under the influence of strong, short wavelength light on some preparations of mepacrine-stained platelets. The release of one or more dense bodies leaves a crater of variable size on the upper surface of the granulomere. Such craters represent the surface component of the open canalicular system and their formation and disappearance can be directly observed. Because these techniques permit quantitation of several parameters of motility which are not readily observable by other techniques, it is suggested that high extinction DIC microscope examination may become a rapid and useful method of studying congenital and acquired platelet disorders. Many features of platelet transformation have been confirmed and extended by scanning electron micrographs. These can in turn be interpreted by reference to time-lapse films of living platelets.

Cinemicroscopic Visualization of Shape Change and Motility in Normal and Abnormal Living Human Platelets

1979 ◽  
Author(s):  
L. R. Zacharski ◽  
R. D. Allen ◽  
R. Rosenstein ◽  
S. Widirtsky
Keyword(s):  
Shape Change ◽  
Blood Platelets ◽  
Rapid Test ◽  
Human Platelets ◽  
Epifluorescence Microscopy ◽  
Optical Sectioning ◽  
Dense Bodies ◽  
Cine Film ◽  
20 Nm ◽  
Human Blood Platelets

Living human blood platelets (P) have been examined with a high extinction differential interference contrast (DIC) microscope capable of detecting structures as small as 20 nm in diameter. During the quarter hour required for complete transformation, the entire shape change sequence is clearly visible, including disk to sphere transformation, extension and retraction of pseudopodia, and spreading and ruffling of the hyalomere. The exocytosis of intact 5-hydroxy tryptamine (serotonin)-containing dense bodies has been observed both by DIC microscopy and by epifluorescence microscopy in P stained with mepacrine. The release of dense bodies is associated with the formation of one or more “craters” in the upper surface of the granulomere. With optical sectioning, it is evident that certain “craters” represent internal chambers of the open canalicular system. Using these techniques, abnormalities in P motility have been observed in hereditary P disorders. In summary, the ability to observe and record permanently on cine film the motility of living P provides a rapid test of P function which allows quantitation of normal vs. abnormal motile behavior.

Thrombin-Induced Surface Changes of Human Platelets in Plasma: Their Relation to Serotonin Release

1979 ◽  
Author(s):  
K. Subbarao ◽  
V.V. Kakkar
Keyword(s):  
Membrane Proteins ◽  
Blood Platelets ◽  
Polyacrylamide Gel Electrophoresis ◽  
Protein Band ◽  
Human Platelets ◽  
Serotonin Release ◽  
Glycoprotein I ◽  
Reversible Aggregation ◽  
Coomassie Blue ◽  
Labeling Pattern

Membrane proteins of both control and thrombin-treated platelets were labeled by NaB3H4, reduction of Schiff bases formed between pyridoxal 5′-phosphate and protein amino groups. Examination of the labeled polypeptides by SDS-polyacrylamide gel electrophoresis and fluorography disclosed a different labeling pattern for thrombin-treated platelets. The distributions of Coomassie blue-stained protein from treated and untreated cells were, by contrast, almost identical. Fluorographs of control platelets showed a single intensely labeled protein band (mol wt 90,000) whereas with cells exposed to thrombin (30-60 milliunits) about 10 protein bands with mol wts ranging from 43,000 to 200,000 were typically present. Among these were: thrombin-sensitive protein (mol wt 188,000), glycoprotein I (mol wt 150,000) and actin (mol wt 43,000). When serotonin release was prevented, either by reversing platelet aggregation with low amounts of ADP (0.1-0.3 μM) or by preincubating with 3',5'-ADP (20 μM), an inhibitor of both ADP- and thrombin-induced platelet function, the labeling patterns on fluorographs were similar to the control. These results indicate that blood platelets can undergo reversible aggregation without major changes in their surface topography, whereas thrombin-induced serotonin release appears related to structural alterations in platelet membrane proteins.

scholarly journals Mechanism of Action of Halofenate, A Platelet Inhibitor

1977 ◽  
Author(s):  
G. R. Favis ◽  
R. W. Colman
Keyword(s):  
Platelet Aggregation ◽  
Platelet Rich Plasma ◽  
Shape Change ◽  
Thiobarbituric Acid ◽  
Serotonin Release ◽  
Prostaglandin Synthesis ◽  
Second Phase ◽  
Change Curve ◽  
Cellulose Column ◽  
Platelet Shape

Halofenate (Hal) has previously been shown to inhibit epinephrine (Epi) and ADP induced platelet aggregation and C14-serotonin release. We further investigated the site of action of Hal by examining platelet shape change as a membrane event and malondialdehyde (MDA) formation as a measure of prostaglandin synthesis. Platelet-rich-plasma (PRP) with and without Hal wasdiluted in an EDTA buffer and examined in a spectrophotometer modified for stirring and maintained at 37°. ADP induced increase in absorbance was recorded and the velocity of the shape change curve was plotted against ADP concentration. MDA production was measured by the thiobarbituric acid assay and utilized a DEAE-52 cellulose column to concentrate the chromogen. Hal in pharmacologic concentrations (.96mM) had no effect on Epi induced primary aggregation or on ADP induced shape change. However, at higher than pharmacologic amounts (3.36mM), Hal did inhibit ADP induced shape change. Epi-induced MDA formation (.18μM-.33μM) normally occurs concomitant with the second phase of aggregation and serotonin release but was markedly decreased by Hal (.06μM-.085μM). This inhibition was not due to a direct effect on prostaglandin synthesis since sodium arachi-donate (1mM) caused secondary aggregation in PRP treated with Hal but not PRP treated with aspirin (4mM). Hal (.96mM) does not seem to inhibit platelet aggregation through an inhibition of ADP induced shape change or of Epi induced primary aggregation. Since Hal treated platelets respond to arachidonate, Hal must work at some earlier step than arachidonate induced prostaglandin synthesis. We suggest that this may be an alteration of the platelet membrane structure which makes ADP and Epi binding sites less accessible or which impairs arachidonic acid release by phospholipase. Decreased MDA formation and inhibition of aggregation would then be secondary to this membrane change.

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