ADP‐stimulated fibrinogen binding is necessary for some of the inositol phospholipid changes found in ADP‐stimulated platelets

Abstract

ADP-stimulation of washed human platelets suspended in Tyrode/albumin solution containing Ca' + (2 mM) and fibrinogen (0.4 mg/ml) causes extensive, reversible aggregation without appreciable secretion of granule contents. Under these conditions ADP (10 pM) stimulation decreased the amounts of phosphatidylinositol 4,5-bisphosphate (PtdInsP,) and phosphatidylinositol 4-phosphate (PtdInsP) at 10 s. Omitting fibrinogen from the suspending medium or blocking fibrinogen binding to the platelets using Arg-Gly-Asp-Ser (RGDS, 0.23 mM) inhibited these decreases in PtdInsP, and PtdinsP. in contrast, ADP-induced decreases in PtdInsP, and increases in PtdInsP at 60 s compared to 10 s were not affected by RGDS or the absence of fibrinogen. in platelets prelabelled with [3H]glycerol and ['2P]phosphate, changes in labelling of the inositol phospholipids paralleled the changes in amount. The ADP-induced changes in phosphatidic acid (PtdOH) at 10 s were unaffected by RGDS; this finding supported previous reports that phospholipase C was not the cause of the early decreases in PtdInsP, and PtdinsP. These results indicate that the early decreases in PtdInsP, and PtdInsP at 10 s are dependent on fibrinogen binding to the platelets and occur after fibrinogen binding which is activated by ADP stimulation. It is proposed that the fibrinogen-dependent changes in PtdInsP, and PtdInsP inay have a feedback role augmenting platelet aggregation or other responses of platelets that might occur after fibrinogen binding, possibly due to effects on actin polymerisation.The receptor for fibrinogen on platelets, the glycoprotein IIbAIIa complex (integrin al&, [l]), has been shown to undergo conformational change in activated platelets to expose the fibrinogen binding site [2-41. In addition, evidence is now accumulating that the binding of fibrinogen to the glycoprotein IIb/IIIa complex (GPIIbDIIa) may cause changes in the complex which affect internal processes in the platelet. An early report to this effect by Isenberg et al. [ 5 ] showed that clustering of GPIIb/IIIa complexes in ADP-stimulated platelets required binding of fibrinogen and did not involve cross-linking by fibrinogen. In a subsequent study, Isenberg et al.[6] demonstrated stimulation of actinomysin-dependent clustering of GPIIb/IIIa in the surface-connected canalicular system of otherwise unstimulated platelets caused by addition of antibodies to GPIIb (Tab) and GPIIIa (AP3). There have been other reports in which monoclonal antibodies to GPIIbAIIa have been shown to cause platelet aggregation [7-91. Since in these cases aggregation was shown to be fibrinogen-dependent, aggregation is not due to platelet cross-linking by the antibodies and demonstration that aggregation was inhibitable by agents (such as prostaglandin E,) which block internal signal transduction pathways indicated that the confornational change in fibrinogen receptors was not only directly induced by antibody binding to GPIIb

HDL3-Mediated Inhibition of Thrombin-Induced Platelet Aggregation and Fibrinogen Binding Occurs via Decreased Production of Phosphoinositide-Derived Second Messengers 1,2-Diacylglycerol and Inositol 1,4,5-tris-Phosphate

“…This apparent discrepancy is not surprising, taking into account that PI-PLC does not appear to play a major role in the ADP-induced platelet activation. [42][43][44][45] Oxidative modification of lipoproteins, which may occur in vivo or during isolation, is thought to change their physicochemical and biological properties. If the inhibitory activity of HDL 3 is a consequence of oxidation, one would expect in vitro-oxidized HDL 3 to act on platelet activation more potently than native HDL 3 .…”

Molecular Basis for ADP-induced Platelet Activation

“…Acting extracellularly, ADP causes a number of intracellular events including rapid calcium influx (6,7), mobilization of intracellular calcium stores (8), and inhibition of adenylyl cyclase (9). An increase in intracellular Ca 2ϩ may be due to an increase in inositol trisphosphate (10 -13) but this finding remains controversial (14,15). In addition, arachidonic acid, liberated from platelet membranes due to activation of phospholipase A 2 , is converted to thromboxane A 2 , itself a powerful platelet agonist.…”