scholarly journals Monoclonal antibodies to human platelet glycoprotein IIb beta that initiate distinct platelet responses

Blood ◽  
1985 ◽  
Vol 65 (5) ◽  
pp. 1112-1119
Author(s):  
LK Jennings ◽  
DR Phillips ◽  
WS Walker
Keyword(s):  
Monoclonal Antibodies ◽  
Platelet Aggregation ◽  
Platelet Activation ◽  
Human Platelet ◽  
Platelet Glycoprotein ◽  
Membrane Glycoprotein ◽  
Spleen Cells ◽  
Glanzmann’S Thrombasthenia ◽  
Glanzmann's Thrombasthenia ◽  
Mouse Myeloma

Hybridomas secreting monoclonal antibodies (MoAbs) to human platelet membrane glycoprotein IIb (GPIIb) were prepared by fusing cells of a mouse myeloma line to spleen cells from a BALB/c mouse immunized with purified GPIIb. Six of the hybridomas secreted MoAbs that recognized epitopes on the 23,000-dalton, disulfide-linked subunit of GPIIb, GPIIb beta. All six of these MoAbs agglutinated platelets in the absence of calcium. The agglutination titers of three of the MoAbs, however, were enhanced between 2 and 6 log2 dilutions when titrated in the presence of mmol/L of calcium. The enhancement in titer was the result of MoAb- induced platelet activation followed by platelet aggregation, a reaction that could also be initiated by the monovalent Fab fragments prepared from one of the MoAbs. The MoAbs did not significantly agglutinate platelets from patients with Glanzmann's thrombasthenia, confirming biochemical evidence that there is a paucity of GPIIb beta in the membranes of these cells. Our results show that MoAbs to epitopes on GPIIb beta initiate distinct platelet responses; therefore, they should be useful for studying the ways in which regions of surface glycoproteins are involved in platelet-platelet interactions. In addition, these reagents may prove of value in diagnosing and typing patients with Glanzmann's thrombasthenia.

scholarly journals Monoclonal antibodies to human platelet glycoprotein IIb beta that initiate distinct platelet responses

Blood ◽  
1985 ◽  
Vol 65 (5) ◽  
pp. 1112-1119 ◽  
Author(s):  
LK Jennings ◽  
DR Phillips ◽  
WS Walker
Keyword(s):  
Monoclonal Antibodies ◽  
Platelet Aggregation ◽  
Platelet Activation ◽  
Human Platelet ◽  
Platelet Glycoprotein ◽  
Membrane Glycoprotein ◽  
Spleen Cells ◽  
Glanzmann’S Thrombasthenia ◽  
Glanzmann's Thrombasthenia ◽  
Mouse Myeloma

Abstract Hybridomas secreting monoclonal antibodies (MoAbs) to human platelet membrane glycoprotein IIb (GPIIb) were prepared by fusing cells of a mouse myeloma line to spleen cells from a BALB/c mouse immunized with purified GPIIb. Six of the hybridomas secreted MoAbs that recognized epitopes on the 23,000-dalton, disulfide-linked subunit of GPIIb, GPIIb beta. All six of these MoAbs agglutinated platelets in the absence of calcium. The agglutination titers of three of the MoAbs, however, were enhanced between 2 and 6 log2 dilutions when titrated in the presence of mmol/L of calcium. The enhancement in titer was the result of MoAb- induced platelet activation followed by platelet aggregation, a reaction that could also be initiated by the monovalent Fab fragments prepared from one of the MoAbs. The MoAbs did not significantly agglutinate platelets from patients with Glanzmann's thrombasthenia, confirming biochemical evidence that there is a paucity of GPIIb beta in the membranes of these cells. Our results show that MoAbs to epitopes on GPIIb beta initiate distinct platelet responses; therefore, they should be useful for studying the ways in which regions of surface glycoproteins are involved in platelet-platelet interactions. In addition, these reagents may prove of value in diagnosing and typing patients with Glanzmann's thrombasthenia.

scholarly journals Human platelet aggregation by murine monoclonal antiplatelet antibodies is subtype-dependent

Blood ◽  
1993 ◽  
Vol 81 (7) ◽  
pp. 1792-1800 ◽  
Author(s):  
S De Reys ◽  
C Blom ◽  
B Lepoudre ◽  
PJ Declerck ◽  
M De Ley ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Platelet Aggregation ◽  
Platelet Activation ◽  
Human Platelet ◽  
Human Platelets ◽  
Antigen Recognition ◽  
Metabolic Inhibitors ◽  
Antigen Binding ◽  
Human Platelet Aggregation ◽  
Murine Monoclonal Antibodies

Abstract Twenty murine monoclonal antibodies (MoAbs) generated against different human platelet antigens induced clumping of human platelets in plasma and buffer. Whereas one MoAb could agglutinate platelets, clumping for 19 MoAbs was blocked by metabolic inhibitors, indicating that these induce platelet activation. Fifteen MoAbs were of IgG1, two of IgG2a, and two of IgG2b subtype. F(ab')2 fragments of these did not evoke an aggregatory response, but specifically inhibited aggregations by and binding of their respective intact MoAbs to platelets. Single-platelet counting technology indicated that the MoAbs bind through their antigen- binding and Fc domains mainly to the surface of the same platelet, rather than cause interplatelet-binding. Despite these similarities, the mechanism of action was nevertheless subtype-dependent. Aggregation induced by all IgG1 antibodies could consistently be prevented by blocking the Fc gamma II-receptor, whereas aggregations induced by all IgG2 antibodies still occurred with blocked Fc-receptor, provided functional complement was present. We therefore conclude that platelet activation by MoAb-binding is initiated by antigen recognition followed by an Fc domain-dependent step, which involves the Fc gamma II-receptor for IgG1-type MoAbs and complement-binding for IgG2-type MoAbs. Thus, antibodies of different subtypes can aggregate platelets via different pathways.

Effect of Three Japanese Kampo Medicines on Platelet Activation by Monoclonal Anti-platelet Membrane Glycoprotein Antibodies

1994 ◽  
Vol 22 (01) ◽  
pp. 71-76 ◽  
Author(s):  
Toshihiro Kawakatsu ◽  
Shosaku Nomura ◽  
Hirofumi Kido ◽  
Kazuyuki Yamaguchi ◽  
Tsutomu Fukuroi ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Flow Cytometry ◽  
Platelet Aggregation ◽  
Platelet Activation ◽  
Antibody Binding ◽  
Platelet Glycoprotein ◽  
Membrane Glycoprotein ◽  
Fc Gamma Receptor ◽  
Gamma Receptor ◽  
Induced Aggregation

We studied the effect of three Japanese kampo medicines on platelet activation by an anti-CD9 monoclonal antibody (NNKY1-19) and an anti-human Fc gamma receptor II monoclonal antibody (NNKY3-2). Sho-saiko-to (TJ-9) and Sairei-to (TJ-114) partially suppressed platelet aggregation induced by NNKYl-19, while Juzen-taiho-to (TJ-48) suppressed aggregation induced by NNKY3-2. TJ-9 and TJ-114 also suppressed collagen-induced aggregation, but TJ-48 did not. Flow cytometry showed that the three medicines did not affect antibody binding to the platelets. Thus, all three kampo medicines suppressed platelet activation by anti-platelet glycoprotein antibodies without inhibiting antibody binding.

scholarly journals Human platelet aggregation by murine monoclonal antiplatelet antibodies is subtype-dependent

Blood ◽  
1993 ◽  
Vol 81 (7) ◽  
pp. 1792-1800 ◽  
Author(s):  
S De Reys ◽  
C Blom ◽  
B Lepoudre ◽  
PJ Declerck ◽  
M De Ley ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Platelet Aggregation ◽  
Platelet Activation ◽  
Human Platelet ◽  
Human Platelets ◽  
Antigen Recognition ◽  
Metabolic Inhibitors ◽  
Antigen Binding ◽  
Human Platelet Aggregation ◽  
Murine Monoclonal Antibodies

Twenty murine monoclonal antibodies (MoAbs) generated against different human platelet antigens induced clumping of human platelets in plasma and buffer. Whereas one MoAb could agglutinate platelets, clumping for 19 MoAbs was blocked by metabolic inhibitors, indicating that these induce platelet activation. Fifteen MoAbs were of IgG1, two of IgG2a, and two of IgG2b subtype. F(ab')2 fragments of these did not evoke an aggregatory response, but specifically inhibited aggregations by and binding of their respective intact MoAbs to platelets. Single-platelet counting technology indicated that the MoAbs bind through their antigen- binding and Fc domains mainly to the surface of the same platelet, rather than cause interplatelet-binding. Despite these similarities, the mechanism of action was nevertheless subtype-dependent. Aggregation induced by all IgG1 antibodies could consistently be prevented by blocking the Fc gamma II-receptor, whereas aggregations induced by all IgG2 antibodies still occurred with blocked Fc-receptor, provided functional complement was present. We therefore conclude that platelet activation by MoAb-binding is initiated by antigen recognition followed by an Fc domain-dependent step, which involves the Fc gamma II-receptor for IgG1-type MoAbs and complement-binding for IgG2-type MoAbs. Thus, antibodies of different subtypes can aggregate platelets via different pathways.

Fc-receptor Dependent Platelet Aggregation Induced by Monoclonal Antibodies against Platelet Glycoprotein Ib or von Willebrand Factor

2001 ◽  
Vol 85 (04) ◽  
pp. 679-685 ◽  
Author(s):  
Nancy Cauwenberghs ◽  
Agotha Schlammadinger ◽  
Stephan Vauterin ◽  
Susan Cooper ◽  
Gretel Descheemaeker ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Platelet Activation ◽  
Von Willebrand Factor ◽  
Human Platelet ◽  
Platelet Rich Plasma ◽  
Fc Receptor ◽  
Platelet Glycoprotein ◽  
Protein Tyrosine Phosphorylation ◽  
Von Willebrand ◽  
Willebrand Factor

SummaryIn this paper we describe two pathways leading to platelet activation by crosslinking glycoprotein (GP) Ib to the platelet Fc-receptor (FcγRII). First the monoclonal antibody (MoAb) 9C8, raised against human platelet GPIbα, dose-dependently induced platelet aggregation of citrate-anticoagulated platelet-rich plasma, an effect that can be inhibited by several activation inhibitors. The FcγRII-inhibitory MoAb IV.3 was able to prevent the aggregatory effects of MoAb 9C8, indicating that crosslinking of the antigen GPIbαto the FcγII-receptor is necessary for the activating effect. Secondly we observed a synergistic activating effect of two anti-von Willebrand factor (vWF) MoAbs 1C1E7 and B724, both known to enhance vWF binding to GPIbαin the presence of shear or ristocetin. When these antibodies are added together to PRP, platelet aggregation is induced without further need for an additional modulator. This effect can be blocked by either MoAb IV.3 or an inhibitory anti-GPIbαMoAb, indicating that again the platelet activation results from signaling through FcγRII crosslinked to vWF bound to GPIbα. In addition, both the anti-GPIbαMoAb 9C8, or the two anti-vWF MoAbs 1C1E7 and B724 induce genuine platelet activation, as evidenced by the secretion of ATP and protein tyrosine phosphorylation. These findings with both anti-GPIbαand anti-vWF MoAbs add further proof to recent reports demonstrating an interaction between the platelet receptors GPIbαand FcγRII, suggesting a role for the FcγII-receptor in GPIb-related signaling.

scholarly journals A Cys374Tyr homozygous mutation of platelet glycoprotein IIIa (beta 3) in a Chinese patient with Glanzmann's thrombasthenia

Blood ◽  
1996 ◽  
Vol 88 (5) ◽  
pp. 1666-1675 ◽  
Author(s):  
CM Grimaldi ◽  
F Chen ◽  
LE Scudder ◽  
BS Coller ◽  
DL French
Keyword(s):  
Platelet Aggregation ◽  
Receptor Expression ◽  
Hunan Province ◽  
Initial Slope ◽  
Platelet Glycoprotein ◽  
Homozygous Mutation ◽  
Binding Studies ◽  
Platelet Surface ◽  
Glanzmann’S Thrombasthenia ◽  
Glanzmann's Thrombasthenia

A 20-year-old woman from a consanguineous family in the Hunan Province of the People's Republic of China was diagnosed as having Glanzmann's thrombasthenia based on (1) nearly a lifelong history of epistaxis, gum bleeding, petechiae, and purpura; (2) severe menorrhagia resulting in anemia and need for whole-blood transfusion; (3) normal coagulation assays; (4) prolonged bleeding time; (5) absent clot retraction; (6) decreased glass bead retention; (7) absent platelet aggregation in response to adenine diphosphate, epinephrine, and collagen; and (8) normal initial slope of platelet aggregation in response to ristocetin, but with a diminished maximal extent. The patient's platelets had a decreased level of platelet fibrinogen, but the deficiency was not as severe as in other Glanzmann's thrombasthenia patients. As judged by monoclonal antibody binding studies, surface glycoprotein (GP) IIb/IIIa (alpha IIb beta 3) expression was less than 15% of normal and alpha v beta 3 vitronectin receptor expression was 15% to 19% of normal, suggesting that the defect was in GPIIIa (beta 3). Immunoblotting of platelet lysates demonstrated decreased levels of GPIIb (approximately 30% to 35% of normal) and GPIIIa (approximately 10% of normal), and the GPIIb had undergone normal maturational processing into GPIIb heavy and light chains. Sequence analysis of the patient's GPIIIa RNA identified a G to A mutation at nucleotide 1219, predicting a Cys to Tyr substitution at residue 374. The patient's parents, who are first cousins, are asymptomatic and have only minor reductions in platelet aggregation. Direct sequencing of polymerase chain reaction-amplified cDNA and GPIIIa exon VIII indicated that the patient is homozygous and her parents are heterozygous for the mutation. Transient transfection studies in Chinese hamster ovary cells indicated that the mutation results in an 85% to 90% reduction in GPIIb/IIIa surface expression, but these cells retain the ability to mediate adhesion to immobilized fibrinogen. The relative preservation of platelet fibrinogen despite the very low level of platelet surface GPIIb/IIIa expression in this patient raises some interesting questions regarding the mechanism of fibrinogen uptake and the pathophysiology of Glanzmann's thrombasthenia.

scholarly journals A Cys374Tyr homozygous mutation of platelet glycoprotein IIIa (beta 3) in a Chinese patient with Glanzmann's thrombasthenia

Blood ◽  
1996 ◽  
Vol 88 (5) ◽  
pp. 1666-1675 ◽  
Author(s):  
CM Grimaldi ◽  
F Chen ◽  
LE Scudder ◽  
BS Coller ◽  
DL French
Keyword(s):  
Platelet Aggregation ◽  
Receptor Expression ◽  
Hunan Province ◽  
Initial Slope ◽  
Platelet Glycoprotein ◽  
Homozygous Mutation ◽  
Binding Studies ◽  
Platelet Surface ◽  
Glanzmann’S Thrombasthenia ◽  
Glanzmann's Thrombasthenia

Abstract A 20-year-old woman from a consanguineous family in the Hunan Province of the People's Republic of China was diagnosed as having Glanzmann's thrombasthenia based on (1) nearly a lifelong history of epistaxis, gum bleeding, petechiae, and purpura; (2) severe menorrhagia resulting in anemia and need for whole-blood transfusion; (3) normal coagulation assays; (4) prolonged bleeding time; (5) absent clot retraction; (6) decreased glass bead retention; (7) absent platelet aggregation in response to adenine diphosphate, epinephrine, and collagen; and (8) normal initial slope of platelet aggregation in response to ristocetin, but with a diminished maximal extent. The patient's platelets had a decreased level of platelet fibrinogen, but the deficiency was not as severe as in other Glanzmann's thrombasthenia patients. As judged by monoclonal antibody binding studies, surface glycoprotein (GP) IIb/IIIa (alpha IIb beta 3) expression was less than 15% of normal and alpha v beta 3 vitronectin receptor expression was 15% to 19% of normal, suggesting that the defect was in GPIIIa (beta 3). Immunoblotting of platelet lysates demonstrated decreased levels of GPIIb (approximately 30% to 35% of normal) and GPIIIa (approximately 10% of normal), and the GPIIb had undergone normal maturational processing into GPIIb heavy and light chains. Sequence analysis of the patient's GPIIIa RNA identified a G to A mutation at nucleotide 1219, predicting a Cys to Tyr substitution at residue 374. The patient's parents, who are first cousins, are asymptomatic and have only minor reductions in platelet aggregation. Direct sequencing of polymerase chain reaction-amplified cDNA and GPIIIa exon VIII indicated that the patient is homozygous and her parents are heterozygous for the mutation. Transient transfection studies in Chinese hamster ovary cells indicated that the mutation results in an 85% to 90% reduction in GPIIb/IIIa surface expression, but these cells retain the ability to mediate adhesion to immobilized fibrinogen. The relative preservation of platelet fibrinogen despite the very low level of platelet surface GPIIb/IIIa expression in this patient raises some interesting questions regarding the mechanism of fibrinogen uptake and the pathophysiology of Glanzmann's thrombasthenia.

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