Monoclonal antibody AG-1 initiates platelet activation by a pathway dependent on glycoprotein IIb-IIIa and extracellular calcium.
The biochemical responses of intact human platelets to the monoclonal antibody (mAb) AG-1 were investigated. AG-1 is a murine IgG mAb that recognizes a series of platelet membrane glycoproteins (Gp) from M(r) 21,000 to 29,000, one of which is the M(r) 24,000 (p24) receptor for anti-CD9 mAbs. AG-1 causes platelet aggregation and secretion. Platelets binding AG-1 demonstrate a dose- and time-dependent breakdown of phosphatidylinositol 4,5-bisphosphate (PIP2), production of diacylglycerol, and generation of phosphatidic acid (PA). These events are associated with the activation of protein kinase C (PKC), an increase in intracellular calcium, and fibrinogen binding. Platelet PA generation and PKC activation in response to AG-1 are inhibited by mAbs to platelet GpIIb-IIIa or by extracellular EGTA, but not by a mAb to platelet GpIb or by inhibiting platelet Na+/H+ exchange with 5-(N-ethyl-N-isopropyl)amiloride. Platelet cytoplasmic free calcium ([Ca2+]i) is elevated in response to AG-1, and this elevation is inhibited by mAbs to GpIIb-IIIa, an RGDS peptide or by chelating extracellular calcium. These results suggest that AG-1 binding to a unique platelet-surface glycoprotein initiates platelet responses through the activation of PIP2-specific phospholipase C, and that this occurs through a signal pathway that is dependent on GpIIb-IIIa and extracellular calcium.
Kroll
M H
MH
VA Medical Center, Houston, TX.
Mendelsohn
M E
ME
Miller
J L
JL
Ballen
K K
KK
Hrbolich
J K
JK
Schafer
A I
AI
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal; D002118:Calcium / Q000378:metabolism*; D002413:Cations, Divalent; D004789:Enzyme Activation; D005340:Fibrinogen / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000494:pharmacology; D010712:Phosphatidic Acids / Q000096:biosynthesis; D019269:Phosphatidylinositol 4,5-Diphosphate; D010716:Phosphatidylinositols / Q000096:biosynthesis; D010766:Phosphorylation; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology; D010980:Platelet Membrane Glycoproteins / Q000276:immunology / Q000378:metabolism*; D011493:Protein Kinase C / Q000378:metabolism; D015398:Signal Transduction
D000911:Antibodies, Monoclonal; D002413:Cations, Divalent; D009842:Oligopeptides; D010712:Phosphatidic Acids; D019269:Phosphatidylinositol 4,5-Diphosphate; D010716:Phosphatidylinositols; D010975:Platelet Aggregation Inhibitors; D010980:Platelet Membrane Glycoproteins; D005340:Fibrinogen; C046617:arginyl-glycyl-aspartyl-serine; D011493:Protein Kinase C; D002118:Calcium