Galactosyltransferase and membrane glycoprotein abnormality in human platelets from Tn-syndrome donors

Nature ◽  
1979 ◽  
Vol 282 (5739) ◽  
pp. 621-623 ◽  
Author(s):  
J. P. Cartron ◽  
A. T. Nurden
Keyword(s):  
Human Platelets ◽  
Membrane Glycoprotein

Differential Ability of Agonists to Express Distinct Pools of Fibrinogen (Gpllb/llla) Receptors which Can Mediate the Aggregation of Human Platelets

1989 ◽  
Vol 62 (03) ◽  
pp. 955-961 ◽  
Author(s):  
Ian S Watts ◽  
Rebecca J Keery ◽  
Philip Lumley
Keyword(s):  
Platelet Aggregation ◽  
Human Platelet ◽  
Surface Membrane ◽  
Blood Platelet ◽  
Human Platelets ◽  
Membrane Glycoprotein ◽  
Platelet Surface ◽  
Human Platelet Aggregation ◽  
Differential Ability ◽  
Blood Platelet Aggregation

SummaryWe have investigated the effect of two procedures that modify human platelet surface membrane glycoprotein (Gp) IIb and IIIa complexes upon whole blood platelet aggregation to a range of agonists. (A) Irreversible disruption of complexes by temporary (30 min) Ca2+-deprivation with EGTA at 37° C. (B) Binding of a monoclonal antibody M148 to the complex. EGTA exposure abolished aggregation to ADP, adrenaline and PAF. In contrast, full aggregation curves to collagen and U-46619 could still be established. EGTA exposure reduced M148 binding to platelets by 80%. Excess M148 abolished aggregation to ADP, PAF, collagen and U-46619. However, upon removal of unbound antibody from platelets full aggregation curves to collagen and U-46619 but not to ADP and PAF could be re-established. Thus human platelet aggregation to ADP, PAF and adrenaline appears absolutely dependent upon surface membrane GpIIb/IIIa complexes. In contrast, collagen and U-46619 cause expression of an additional distinct pool of Gp complexes inaccessible to EGTA and M148 in unstimulated platelets which is intimately involved in aggregation to these agonists.

The Use of Monoclonal Antibodies to Human Platelet Membrane Glycoprotein IIb-IIIa to Quantitate Platelet Deposition on Artificial Surfaces

1987 ◽  
Vol 58 (02) ◽  
pp. 724-731 ◽  
Author(s):  
Juliette N Mulvihill ◽  
Han G Huisman ◽  
Jean-Pierre Cazenave ◽  
Jan A van Mourik ◽  
Willem G van Aken
Keyword(s):  
Whole Blood ◽  
Human Albumin ◽  
Human Platelets ◽  
Membrane Glycoprotein ◽  
Bovine Collagen ◽  
Glass Capillaries ◽  
Platelet Deposition ◽  
A New Technique ◽  
Albumin Adsorption

SummaryA new technique is described to quantitate platelet deposition in vitro on artifical surfaces, based on a surface phase radioimmunoassay using the monoclonal antibody 6C9, directed specifically against the membrane glycoprotein complex IIb-IIIa of human platelets. Results correlate in linear fashion with those obtained using 111Indium labeled platelets. The method offers particular advantages for the measurement of platelet deposition in whole blood, since platelet separation, washing and labeling procedures are eliminated, together with the ensuing possible selection of platelet populations. In vitro perfusion is performed in glass capillaries of precisely defined diameter (0.80 or 0.56 mm i.d.). Blood flow is laminar and accurately controlled over wall shear rates ranging from venous to capillary (50-4,000 s-1). Using glass capillaries precoated with purified human albumin or fibrinogen or bovine collagen, platelet deposition from suspensions of washed human platelets in Tyrode's-albumin buffer in the presence of a 40% hematocrit is (platelets/mm2): 11,000 (albumin), 78,000 (fibrinogen) and 306,000 (collagen) after 5 min perfusion at 2,000 s-1. In heparin, citrate or hirudin anticoagulated whole blood, surfaces are passivated, probably by albumin adsorption from plasma (platelets/mm2): 400 (albumin), 3,600 (fibrinogen) and 48,000 (collagen) after 5 min perfusion in the presence of 13 mM citrate.

Absence of the 145,000 Molecular Weight, Soluble Platelet Membrane Glycoprotein - Lack of Platelet Agglutination

1979 ◽  
Vol 42 (05) ◽  
pp. 1626-1629 ◽  
Author(s):  
Nils Olav Solum ◽  
Inger Hagen ◽  
Miroslav Peterka ◽  
Torbjørn Gjemdal
Keyword(s):  
Molecular Weight ◽  
Factor Viii ◽  
Human Platelets ◽  
Platelet Membrane ◽  
Membrane Glycoprotein ◽  
Related Protein ◽  
Giant Platelets ◽  
Platelet Membrane Glycoprotein ◽  
Human Factor Viii ◽  
Bovine Factor

SummaryOne step in the function of platelets in hemostasis is their adhesion to subendothelial tissue. The human factor VIII related protein (von Willebrand factor) is considered to be involved in the adhesion phenomenon (Baumgartner et al. 1977). One manifestation of the protein-cell interaction can be observed as a platelet agglutination after addition to the human platelets of a combination of the human protein and the glycopeptide ristocetin, or after addition of the bovine protein alone. The bovine factor VIII related protein as such directly binds to the platelet membrane (Kirby and Sha May Tang 1977) and thus represents a simpler system than ristocetin plus the human cofactor which may have to interact with each other before excerting their effect on the platelet membrane. The present paper concerns the se.One of the characteristics of the agglutination of human platelets brought about by the bovine factor VIII related protein (as well as by ristocetin plus the human cofactor) is that it is independent of the energy metabolism and the internal organization of the platelet. One would therefore expect that modified platelets and platelet “ghosts” would agglutinate as long as certain structures on the outer cell surface are chemically and sterically intact. Because of the hydrophilic character of the carbohydrate side chains, the membrane glycoproteins are considered of special importance for cell contact phenomena. Thus it has already been known for some years that giant platelets of the Bernard-Soulier type which do not agglutinate with the bovine protein (Bithell et al. 1972), contain a reduced amount of sialic acid related to protein content and surface area (Grottum and Solum 1969), and show a reduced glycoprotein stain in the GP I region on SDS polyacrylamide gel electrophoresis (Nurden and Caen 1975).This paper presents five observations which support a working hypothesis stating that the presence on the platelet membrane of the 145,000 molecular weight, soluble platelet membrane glycoprotein called GPS or glycocalicin is a prerequisite to the agglutination of human platelets by bovine factor VIII related protein.

scholarly journals Characterization of human platelet glycoprotein antigens giving rise to individual immunoprecipitates in crossed-immunoelectrophoresis

Blood ◽  
1981 ◽  
Vol 58 (6) ◽  
pp. 1190-1197 ◽  
Author(s):  
TJ Kunicki ◽  
AT Nurden ◽  
D Pidard ◽  
NR Russell ◽  
JP Caen
Keyword(s):  
Sodium Dodecyl ◽  
Electrophoretic Analysis ◽  
Human Platelets ◽  
Soluble Proteins ◽  
Platelet Glycoprotein ◽  
Membrane Glycoprotein ◽  
Membrane Glycoproteins ◽  
Reference Pattern ◽  
Crossed Immunoelectrophoresis ◽  
Normal Individuals

Abstract Washed human platelets were labeled with 125I by the lactoperoxidase- catalyzed method and solubilized in 1% Triton X-100. The soluble proteins were analyzed by crossed-immunoelectrophoresis in 1% agarose, employing a rabbit antibody raised against whole human platelets. Analysis of autoradiograms developed from dried agarose gels led to the establishment of a normal reference pattern that was consistent for platelets obtained from more than 50 normal individuals. Six platelet membrane glycoprotein antigens contained in four distinguishable precipitates were identified. Each identification was based on direct sodium dodecyl sulfate-polyacrylamide gel electrophoretic analysis of 125I-antigens contained in individually excised precipitates. These platelet antigens include major membrane glycoproteins previously designated la, lb, lla, llb, llla, and lllb. Glycoproteins llb and llla were shown to be contained in a single immunoprecipitate, while glycoproteins la and lla were routinely detected in a single different immunoprecipitate. Analysis of soluble proteins from platelets of five patients with Glanzmann's thrombasthenia demonstrated either a complete absence or a marked reduction of only one radiolabeled precipitate, that containing membrane glycoproteins llb and llla. Platelet samples from two patients with Bernard-Soulier syndrome were devoid of a different precipitate, that containing membrane glycoprotein lb.

scholarly journals Isolation and structural characterization of the polypeptide subunits of membrane glycoprotein IIb-IIIa from human platelets

Blood ◽  
1982 ◽  
Vol 59 (1) ◽  
pp. 80-85 ◽  
Author(s):  
RP McEver ◽  
JU Baenziger ◽  
PW Majerus
Keyword(s):  
High Performance ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Structural Features ◽  
Human Platelets ◽  
Membrane Glycoprotein ◽  
Radiolabeled Peptides ◽  
The Relationship ◽  
Peptide Maps

Abstract We have previously demonstrated the isolation of platelet membrane glycoprotein IIb-IIIa by affinity chromatography with a specific monoclonal antibody. We have now separated the polypeptide subunits IIb and IIIa of the isolated glycoprotein by preparative sodium dodecyl sulfate polyacrylamide gel electrophoresis and have compared their structural features. Both IIb and IIIa contain approximately 15% carbohydrate, but IIIa contains a larger percentage of mannose residues, suggesting the presence of high mannose as well as complex N- linked oligosaccharide chains. The amino acid compositions are sufficiently similar to imply areas of sequence homology between the two subunits. To examine further the relationship between the subunits, we digested a mixture of 125I-IIb and 131I-IIIa with trypsin and then separated the radiolabeled peptides by high performance liquid chromatography. The resultant peptide maps of IIb and IIIa are completely different. This indicates that neither subunit is derived from the other and suggests that polypeptides IIb and IIIa are products of separate genes.

scholarly journals Stimulus-response coupling in human platelets activated by monoclonal antibodies to the CD9 antigen, a 24 kDa surface-membrane glycoprotein

1990 ◽  
Vol 266 (2) ◽  
pp. 527-535 ◽  
Author(s):  
R C Carroll ◽  
R E Worthington ◽  
C Boucheix
Keyword(s):  
Monoclonal Antibody ◽  
Protein Kinase C ◽  
Platelet Aggregation ◽  
Protein Kinase ◽  
Phospholipase C ◽  
Surface Membrane ◽  
Human Platelets ◽  
Membrane Glycoprotein ◽  
Kinase C ◽  
Autocrine Stimulation

The CD9 molecule is a 24 kDa surface-membrane glycoprotein present on platelets and a variety of haematopoetic and non-haematopoetic tissues. In the present study we utilized specific inhibitors of thromboxane A2 (TxA2) formation (aspirin), protein kinase C [H-7 [1-(5-isoquinolinesulphonyl)-2-methylpiperazine]] and autocrine stimulation by secreted ADP (apyrase) to modify platelet activation by a monoclonal antibody ALB-6 to the CD9 antigen. This activation is only partially inhibited by aspirin alone but, in combination with either H-7 or apyrase, more than 50% inhibition of platelet aggregation and secretion was observed. This combination of inhibitors was also required to inhibit effectively the phosphorylation of myosin light chain and the 47 kDa substrate of protein kinase C. Intracellular Ca2+ flux monitored by the fluorescent dye fura-2 showed that this was almost completely mediated by the aspirin-sensitive TxA2 pathway. We suggest that the aspirin-insensitive pathway is primarily mediated by phospholipase C formation of diacylglycerol to activate protein kinase C. The inhibition by apyrase suggests a strong dependency on autocrine stimulation by secreted ADP to fully activate both phospholipase C and express fibrinogen-binding sites mediating platelet aggregation. This alternate pathway of phospholipase C activation by ALB-6 may be mediated by cytoplasmic alkalinization [monitored by SNARF-1 (5′(6′)-carboxy-10-bismethylamino-3-hydroxy-spiro-[7H- benzo[c]xanthine-1′,7(3H)-isobenzofuran]-3′-one) fluorescence of the dye]. Both activation pathways are dependent on intact antibodies, since F(ab′)2 fragments of SYB-1, a monoclonal antibody against the CD9 antigen with activation characteristics identical with those of ALB-6, do not elicit activation. Besides thrombin, collagen is another physiological agonist shown to induce aspirin-insensitive activation. Similarities to ALB-6 in collagen sensitivity to apyrase in combination with aspirin inhibitors were noted with respect to aggregation and secretion, as well as a complete block of Ca2+ flux by aspirin. However, it is unlikely that collagen activation is mediated by the CD9 antigen, since SYB-1 F(ab′)2 fragments had no effect on collagen activation and aspirin also completely blocked the alkalinization response to collagen, in contrast with ALB-6.

scholarly journals Isolation and structural characterization of the polypeptide subunits of membrane glycoprotein IIb-IIIa from human platelets

Blood ◽  
1982 ◽  
Vol 59 (1) ◽  
pp. 80-85 ◽  
Author(s):  
RP McEver ◽  
JU Baenziger ◽  
PW Majerus
Keyword(s):  
High Performance ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Structural Features ◽  
Human Platelets ◽  
Membrane Glycoprotein ◽  
Radiolabeled Peptides ◽  
The Relationship ◽  
Peptide Maps

We have previously demonstrated the isolation of platelet membrane glycoprotein IIb-IIIa by affinity chromatography with a specific monoclonal antibody. We have now separated the polypeptide subunits IIb and IIIa of the isolated glycoprotein by preparative sodium dodecyl sulfate polyacrylamide gel electrophoresis and have compared their structural features. Both IIb and IIIa contain approximately 15% carbohydrate, but IIIa contains a larger percentage of mannose residues, suggesting the presence of high mannose as well as complex N- linked oligosaccharide chains. The amino acid compositions are sufficiently similar to imply areas of sequence homology between the two subunits. To examine further the relationship between the subunits, we digested a mixture of 125I-IIb and 131I-IIIa with trypsin and then separated the radiolabeled peptides by high performance liquid chromatography. The resultant peptide maps of IIb and IIIa are completely different. This indicates that neither subunit is derived from the other and suggests that polypeptides IIb and IIIa are products of separate genes.

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