scholarly journals Thrombin-induced platelet membrane glycoprotein IIb and IIIa complex formation. An electron microscope study.

1981 ◽  
Vol 154 (4) ◽  
pp. 1058-1068 ◽  
Author(s):  
M J Polley ◽  
L L Leung ◽  
F Y Clark ◽  
R L Nachman
Keyword(s):  
Complex Formation ◽  
Keyhole Limpet Hemocyanin ◽  
Human Platelets ◽  
Platelet Membrane ◽  
Macromolecular Complex ◽  
Membrane Glycoprotein ◽  
Platelet Surface ◽  
Platelet Membrane Glycoprotein ◽  
Normal Platelet ◽  
Glycoprotein Iiia

The topographic relationships of platelet membrane glycoprotein IIb and glycoprotein IIIa have been studied in stimulated and unstimulated human platelets using immunoelectron microscopy. An indirect approach with ferritin-conjugated goat anti-rabbit gamma-globulin was used to localize the rabbit antibody to glycoprotein IIIa. The second ultrastructural label was keyhole limpet hemocyanin conjugated directly to antibody to glycoprotein IIb. Using the double labels, it was demonstrated that glycoprotein IIb and glycoprotein IIIa were distributed randomly in the unstimulated platelet membrane. After platelet stimulation with thrombin, large clusters of glycoprotein IIb-glycoprotein IIIa complexes were formed. No complex formation between glycoprotein Ib and glycoprotein IIb was observed in control experiments. These observations suggest that thrombin stimulation initiates the specific glycoprotein IIb-glycoprotein IIIa macromolecular complex formation on the platelet surface, which may act as the active fibrinogen-binding site required for normal platelet aggregation.

DN-9693: A PHOSPHODIESTERASE INHIBITOR WITH A PLATELET MEMBRANE EFFECT

1987 ◽  
Author(s):  
J Ball ◽  
M Greaves ◽  
C Jackson ◽  
J Peel ◽  
F E Preston
Keyword(s):  
Platelet Rich Plasma ◽  
Phosphodiesterase Inhibitor ◽  
Water Soluble ◽  
Platelet Membrane ◽  
Membrane Glycoprotein ◽  
Glycoprotein Ib ◽  
Platelet Surface ◽  
Platelet Membrane Glycoprotein ◽  
Plasma Factor ◽  
Glycoprotein Iiia

We have examined the effect of DN-9693 (piperidinyl - imidazo - quinazoline: Daiichi Seiyaku, Japan) a water soluble phosphodiesterase inhibitor on platelet aggregation, secretion and thromboxane B2 (TXB2) production. In platelet rich plasma and at concentrations of 2uM and 5uM the drug significantly inhibited aggregation induced by adenosine diphosphate, collagen and sodium arachidonate. TXB2 production and release of adenosine tri-phosphate and 14C 5-hydroxytryptamine were also significantly inhibited by the drug. Cyclic adenosine mono-phosphate accumulation was enhanced. Inhibition of ristocetin induced platelet agglutination was an unexpected finding and further experiments were undertaken to explore this. These suggested no specific effect against a plasma factor (von Willebrand factor) and reduced expression of the platelet membrane glycoprotein Ib was implicated. To investigate this further we examined the effect of DN-9693 on the binding of a monoclonal antibody (McAb) to platelet membrane glycoprotein lb (AN51). This was assessed by a FACS IV flow cytofluorimeter utilising a goat anti-mouse fluorescein isothiocyanate (FITC) labelled secondary antibody. Similar experiments were also performed with McAbs to the membrane glycoprotein complex IIb/IIIa (M148) and also to glycoprotein IIIa (C17). In platelet rich plasma, at concentrations which have been shown to inhibit aggregation, DN-9693 significantly reduced the mean fluorescence intensity of the cells coated with McAb AN51 in a dose related manner. This strongly suggested a drug effect against the glycoprotein Ib receptor site. Also, the drug appeared to enhance the binding of McAb C17 to glycoprotein IIIa. This study indicates that in addition to potent phosphodiesterase inhibitor activity, DN-9693 causes a platelet surface membrane change which is associated with reduced expression of membrane glycoprotein Ib.

An additional HpaII polymorphism in exon 2 of the human platelet membrane glycoprotein IIIa gene

1994 ◽  
Vol 93 (3) ◽  
pp. 353-354 ◽  
Author(s):  
Bruno Perichon ◽  
Sylvie Clemenceau ◽  
Alain Romand ◽  
Jacques Elion ◽  
Cecile Kaplan ◽  
...  
Keyword(s):  
Human Platelet ◽  
Platelet Membrane ◽  
Membrane Glycoprotein ◽  
Exon 2 ◽  
Platelet Membrane Glycoprotein ◽  
Glycoprotein Iiia

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 Platelet membrane topography: colocalization of thrombospondin and fibrinogen with the glycoprotein IIb-IIIa complex

Blood ◽  
1985 ◽  
Vol 66 (4) ◽  
pp. 926-934 ◽  
Author(s):  
AS Asch ◽  
LL Leung ◽  
MJ Polley ◽  
RL Nachman
Keyword(s):  
Thin Section ◽  
Platelet Activation ◽  
Immunoelectron Microscopy ◽  
Human Platelets ◽  
The Body ◽  
Platelet Membrane ◽  
Macromolecular Complex ◽  
Platelet Surface ◽  
Membrane Topography

Abstract The distribution of platelet thrombospondin (TSP), fibrinogen, and glycoproteins IIb-IIIa (GPIIb-IIIa) and GPIb were studied in resting and activated human platelets using frozen thin-section immunoelectron microscopy. In resting platelets, TSP and fibrinogen were found within alpha granules and not on the platelet surface. In unstimulated platelets, GPIIb-IIIa and GPIb were distributed diffusely over the platelet membrane as well as within the body of the platelets. Upon thrombin or A23187 stimulation, TSP, fibrinogen, and GPIIb-IIIa colocalized on the platelet membrane and the canalicular system as well as on pseudopodia and between adherent platelets. GPIb distribution was unchanged by platelet activation. The findings support the hypothesis that a macromolecular complex of TSP-fibrinogen and GPIIb-IIIa forms on the activated platelet membrane.

THE FUNCTIONAL DOMAIN OF PLATELET MEMBRANE GLYCOPROTEIN lb FOR VON WILLEBRAND FACTOR AND THROMBIN-BINDING

1987 ◽  
Author(s):  
M Taki ◽  
K Sato ◽  
Y Ikeda ◽  
M Yamamoto ◽  
K Watanabe
Keyword(s):  
Monoclonal Antibody ◽  
Von Willebrand Factor ◽  
Platelet Membrane ◽  
Functional Domain ◽  
Membrane Glycoprotein ◽  
Dependent Manner ◽  
Platelet Surface ◽  
Platelet Membrane Glycoprotein ◽  
Von Willebrand ◽  
Willebrand Factor

In this paper, we have examined the functional domain of platelet membrane glycoprotein lb (GPIb) by using elastase and a monoclonal antibody against GPIb which specific inhibits both von Willebrand factor (vWF) and thrombin interaction with platelets. Elastase was purified from human granulocytes by using affinity column chromatography according to the method of Okada et al.. A monoclonal antibody against platelet membrane GPIb (56-2) which inhibits both vWF and thrombin-binding to platelets was used for this study. Platelet surface glycoproteins were labelled with 3H by the method of Nurden et al.. Purified GPIb was obtained by a modification of the method of Coller et al. and labelled with 125I using chloramine-T method. Either 3H-labelled platelets or 125I-labelled GPIb was treated with elastase for various time periods. Elastase-treated l25I-GPIb was subjected to immunoaffinity chromatography using 56-2 antibody to determine the functional site of GPIb. Elastase inhibited platelet aggregation or 5-HT release by thrombin, ristocetin-induced platelet agglutination and vWF-binding to platelets in the presence of ristocetin in a dose- and time dependent manner. A fluorogram of SDS-PAGE of 3H-labelled platelets treated with elastase revealed that GPIb band was reduced gradually, and fragments with MW of 97, 70, 60, 47, 44, 37, 25 and 15 KD were released from the platelets. The 47 KD fragment was initially cleaved from the platelets, and subsequently other fragments were digested. Similar results were obtained when purified 125I-GPIb was digested by elastase. When the fragments from purified 125I-GPIb were reacted with 56-2 antibody, only three fragments with MW of 47, 44 and 25 KD were immunoisolated. The electrophoretic mobility of all these three bands was altered under reduced conditions, indicating that all these fragments contain disulfide bonds in their molecules. The 25 KD band showed a much fainter in 3H-labelling than in 125I-labelling.These results suggest that the functional domains of GPIb for both vWF and thrombin-binding may be located in a less glycosylated fragment with a MW of 25 KD on the distal portion of the GPIb molecule, which should contain at least one intramolecular disulfide bond.

scholarly journals Platelet membrane topography: colocalization of thrombospondin and fibrinogen with the glycoprotein IIb-IIIa complex

Blood ◽  
1985 ◽  
Vol 66 (4) ◽  
pp. 926-934 ◽  
Author(s):  
AS Asch ◽  
LL Leung ◽  
MJ Polley ◽  
RL Nachman
Keyword(s):  
Thin Section ◽  
Platelet Activation ◽  
Immunoelectron Microscopy ◽  
Human Platelets ◽  
The Body ◽  
Platelet Membrane ◽  
Macromolecular Complex ◽  
Platelet Surface ◽  
Membrane Topography

The distribution of platelet thrombospondin (TSP), fibrinogen, and glycoproteins IIb-IIIa (GPIIb-IIIa) and GPIb were studied in resting and activated human platelets using frozen thin-section immunoelectron microscopy. In resting platelets, TSP and fibrinogen were found within alpha granules and not on the platelet surface. In unstimulated platelets, GPIIb-IIIa and GPIb were distributed diffusely over the platelet membrane as well as within the body of the platelets. Upon thrombin or A23187 stimulation, TSP, fibrinogen, and GPIIb-IIIa colocalized on the platelet membrane and the canalicular system as well as on pseudopodia and between adherent platelets. GPIb distribution was unchanged by platelet activation. The findings support the hypothesis that a macromolecular complex of TSP-fibrinogen and GPIIb-IIIa forms on the activated platelet membrane.

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