Synergistic outside-in regulation of platelet activation by GPIIb/IIIa ligand-induced conformation and oligomerization.
Full platelet activation with serotonin secretion and thromboxane A(2) (TxA(2)) formation induced by a low dose of thrombin receptor agonist peptide (TRAP) or high dose ADP requires platelet aggregation. This requirement can be replaced by pretreatment of platelets with a combination of reagents including: GPIIb/IIIa inhibitors yielding ligand-induced binding sites (LIBS), either arginine-glycine-aspartate-serine (RGDS) peptide or Ro 43-5054, cytochalasin to disrupt actin filaments and crosslinking by a GPIIb/IIIa mAb (pl-62). Crosslinking is required since Fab fragments of pl-62 do not support activation. Engagement of the Fc receptor by the mAb Fc domain is not required for pl-62 augmentation, since it is not blocked by the anti-Fc receptor mAb, IV-3. Another GPIIb/IIIa inhibitor, Ro 44-9883, not yielding LIBS epitopes, serves as a negative control and shows a requirement for LIBS in addition to crosslinking. Focal adhesion kinase tyrosine phosphorylation induced by TRAP is blocked by these GPIIb/IIIa antagonists, but restored by pl-62 crosslinking independent of LIBS induction. Tyrosine phosphorylation of a peptide comigrating with p38 MAP kinase is also inhibited by these antagonists and restored by pl-62 crosslinking. However, p38 MAP kinase activation by low dose TRAP is not affected by these aggregation inhibitors. Tyrosine phosphorylation of a 34-kDa phosphoprotein in the absence of aggregation or TxA(2) formation was uniquely augmented by Ro 43-5054 but not Ro 44-9883 under the above activation conditions.
Wang
X F
XF
Department of Anesthesiology, University of Tennessee, Graduate School of Medicine, Knoxville, TN, USA.
Liu
F
F
Morris
S A
SA
Carroll
R C
RC
D000085:Acetates / Q000494:pharmacology; D000244:Adenosine Diphosphate / Q000494:pharmacology; D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism / Q000494:pharmacology; D001665:Binding Sites / Q000187:drug effects; D006224:Cricetinae; D003432:Cross-Linking Reagents / Q000378:metabolism / Q000494:pharmacology; D019281:Dimerization; D004357:Drug Synergism; D008024:Ligands; D009842:Oligopeptides / Q000494:pharmacology; D015539:Platelet Activation / Q000187:drug effects* / Q000502:physiology; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000502:physiology*; D011487:Protein Conformation / Q000187:drug effects; D012701:Serotonin / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D013928:Thromboxane A2 / Q000096:biosynthesis; D014443:Tyrosine / Q000031:analogs & derivatives* / Q000494:pharmacology
D000085:Acetates; D000911:Antibodies, Monoclonal; D003432:Cross-Linking Reagents; D008024:Ligands; D009842:Oligopeptides; D010975:Platelet Aggregation Inhibitors; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; C078174:Ro 43-5054; D012701:Serotonin; D014443:Tyrosine; D013928:Thromboxane A2; D000244:Adenosine Diphosphate; C084062:lamifiban