Quantitation of microparticles released from coated‐platelets

Abstract

Summary. Dual agonist stimulation of platelets with thrombin and convulxin results in generation of coated-platelets, a sub-population of cells known formerly as COAT-platelets (collagen and thrombin). Coated-platelets retain several procoagulant proteins on their surface and express phosphatidylserine (PS). In this report, we utilize a new methodology to demonstrate that coated-platelets also release microparticles. Platelets were prelabeled with 2.5 lM Bodipy-maleimide and then stimulated with convulxin plus thrombin. Microparticles, 0.3-0.5 lm in diameter, were observed by fluorescence confocal microscopy. Confocal microscopy was also used to demonstrate that microparticles were positive for glycoprotein IIb/IIIa, glycoprotein Ib, CD9, and PS, but negative for fibrinogen and thrombospondin. Furthermore, microparticles released from Bodipy-labeled platelets were observed by flow cytometry, and activation with convulxin plus thrombin produced 15 ± 5 microparticles per coated-platelet. In contrast, platelets stimulated with thrombin or convulxin alone produced few microparticles. Phenylarsine oxide and diamide, both of which potentiate the mitochondrial permeability transition pore and coated-platelet production, significantly increased the number of microparticles released per coated-platelet.

Expression of complement components and inhibitors on platelet microparticles

“…It will be interesting to explore whether PMP generated by platelet stimulation with combinations of other agonists express differing capacities for classical complement pathway activation. Expression of procoagulant activity and Factor V on activated platelets, for example, is especially pronounced in subsets of platelets referred to as "coated" platelets, activated by thrombin and collagen or thrombin and A23187 [29,30].…”

Developmental Endothelial Locus-1 (Del-1) Mediates Clearance of Platelet Microparticles by the Endothelium

“…After 6 hours, blood was drawn from the retroorbital plexus, and microparticles in mouse plasma were quantified as described previously. 28 …”

Coated Platelet Levels Correlate With Bleed Volume in Patients With Spontaneous Intracerebral Hemorrhage

“…Analyses included calculation of descriptive statistics, independent t tests, bivariate and partial correlations for comparison of continuous variables, and 2 or Fisher exact tests for comparison of categorical variables with significance set at PՅ0.05.…”

“…1 The biochemical characteristics of coated platelets include a robust prothrombinase activity, retention of several procoagulant proteins on the cell surface, and release of microparticles. 1,2 The critical nature of coated platelets in hemostasis is revealed by a genetic disorder in dogs, in which deficiency of coated platelets results in spontaneous mucosal bleeding and postsurgical hemorrhage. 3 Previous studies in ischemic stroke indicated that coated platelet levels are elevated in patients with nonlacunar stroke as compared with control subjects, a finding in keeping with the thrombotic nature of these events.…”