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Identification of platelet function defects by multi-parameter assessment of thrombus formation.

de Witt SM, Swieringa F, Cavill R, Lamers MM, van Kruchten R, Mastenbroek T, Baaten C, Coort S, Pugh N, Schulz A, Scharrer I, Jurk K, Zieger B, Clemetson KJ, Farndale RW, Heemskerk JW, Cosemans JM - Nat Commun (2014)

Bottom Line: Three types of thrombus formation can be identified with a predicted hierarchy of the following receptors: glycoprotein (GP)VI, C-type lectin-like receptor-2 (CLEC-2)>GPIb>α6β1, αIIbβ3>α2β1>CD36, α5β1, αvβ3.Application with patient blood reveals distinct abnormalities in thrombus formation in patients with severe combined immune deficiency, Glanzmann's thrombasthenia, Hermansky-Pudlak syndrome, May-Hegglin anomaly or grey platelet syndrome.We suggest this test may be useful for the diagnosis of patients with suspected bleeding disorders or a pro-thrombotic tendency.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), Maastricht University, Universiteitssingel 50, 6229 ER Maastricht, The Netherlands.

ABSTRACT
Assays measuring platelet aggregation (thrombus formation) at arterial shear rate mostly use collagen as only platelet-adhesive surface. Here we report a multi-surface and multi-parameter flow assay to characterize thrombus formation in whole blood from healthy subjects and patients with platelet function deficiencies. A systematic comparison is made of 52 adhesive surfaces with components activating the main platelet-adhesive receptors, and of eight output parameters reflecting distinct stages of thrombus formation. Three types of thrombus formation can be identified with a predicted hierarchy of the following receptors: glycoprotein (GP)VI, C-type lectin-like receptor-2 (CLEC-2)>GPIb>α6β1, αIIbβ3>α2β1>CD36, α5β1, αvβ3. Application with patient blood reveals distinct abnormalities in thrombus formation in patients with severe combined immune deficiency, Glanzmann's thrombasthenia, Hermansky-Pudlak syndrome, May-Hegglin anomaly or grey platelet syndrome. We suggest this test may be useful for the diagnosis of patients with suspected bleeding disorders or a pro-thrombotic tendency.

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Related in: MedlinePlus

Protein surfaces used for flow studies and assignment map of platelet receptors interacting with indicated proteins.Assignment of interactions of platelet receptors to immobilized protein or peptide ligands was as represented in Supplementary Table 1. Surfaces were numbered 1–52, based on unsupervised hierarchical cluster analysis of thrombus parameters (see Fig. 4). Colour code: white, not involved; black, involved at low/high wall shear rate.
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f1: Protein surfaces used for flow studies and assignment map of platelet receptors interacting with indicated proteins.Assignment of interactions of platelet receptors to immobilized protein or peptide ligands was as represented in Supplementary Table 1. Surfaces were numbered 1–52, based on unsupervised hierarchical cluster analysis of thrombus parameters (see Fig. 4). Colour code: white, not involved; black, involved at low/high wall shear rate.

Mentions: To compare the roles of established platelet-adhesive receptors in thrombus formation, we selected ligands of these receptors that are present in either the vessel wall or platelet aggregates. In addition, we used a number of chemically synthesized peptides binding to the same receptors (Supplementary Table 1). In preliminary experiments, we used different coating concentrations to assure that the selected proteins and peptides: (i) remained bound to degreased glass coverslips after repeated rinsing (Coomassie staining), and (ii) showed optimal platelet adhesion under static conditions (S.M.d.W., personal communication). Optimized surfaces for platelet adhesion consisted of the following proteins: collagens I and III, decorin, fibrinogen, fibronectin, laminin 511/521, osteopontin, thrombospondin-1, vitronectin and/or vWF (Fig. 1). Also included were the snake venom rhodocytin, four collagen-mimetic triple-helical peptides, that is, GFOGER-(GPP)n, GFOGER-(GPO)n, (GPO)n and a vWF-binding peptide (vWF-BP). Immobilized bovine serum albumin (BSA) served as negative control surface. Optimally effective coating concentrations of the collagen-mimetic peptides were established before2437.


Identification of platelet function defects by multi-parameter assessment of thrombus formation.

de Witt SM, Swieringa F, Cavill R, Lamers MM, van Kruchten R, Mastenbroek T, Baaten C, Coort S, Pugh N, Schulz A, Scharrer I, Jurk K, Zieger B, Clemetson KJ, Farndale RW, Heemskerk JW, Cosemans JM - Nat Commun (2014)

Protein surfaces used for flow studies and assignment map of platelet receptors interacting with indicated proteins.Assignment of interactions of platelet receptors to immobilized protein or peptide ligands was as represented in Supplementary Table 1. Surfaces were numbered 1–52, based on unsupervised hierarchical cluster analysis of thrombus parameters (see Fig. 4). Colour code: white, not involved; black, involved at low/high wall shear rate.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4109023&req=5

f1: Protein surfaces used for flow studies and assignment map of platelet receptors interacting with indicated proteins.Assignment of interactions of platelet receptors to immobilized protein or peptide ligands was as represented in Supplementary Table 1. Surfaces were numbered 1–52, based on unsupervised hierarchical cluster analysis of thrombus parameters (see Fig. 4). Colour code: white, not involved; black, involved at low/high wall shear rate.
Mentions: To compare the roles of established platelet-adhesive receptors in thrombus formation, we selected ligands of these receptors that are present in either the vessel wall or platelet aggregates. In addition, we used a number of chemically synthesized peptides binding to the same receptors (Supplementary Table 1). In preliminary experiments, we used different coating concentrations to assure that the selected proteins and peptides: (i) remained bound to degreased glass coverslips after repeated rinsing (Coomassie staining), and (ii) showed optimal platelet adhesion under static conditions (S.M.d.W., personal communication). Optimized surfaces for platelet adhesion consisted of the following proteins: collagens I and III, decorin, fibrinogen, fibronectin, laminin 511/521, osteopontin, thrombospondin-1, vitronectin and/or vWF (Fig. 1). Also included were the snake venom rhodocytin, four collagen-mimetic triple-helical peptides, that is, GFOGER-(GPP)n, GFOGER-(GPO)n, (GPO)n and a vWF-binding peptide (vWF-BP). Immobilized bovine serum albumin (BSA) served as negative control surface. Optimally effective coating concentrations of the collagen-mimetic peptides were established before2437.

Bottom Line: Three types of thrombus formation can be identified with a predicted hierarchy of the following receptors: glycoprotein (GP)VI, C-type lectin-like receptor-2 (CLEC-2)>GPIb>α6β1, αIIbβ3>α2β1>CD36, α5β1, αvβ3.Application with patient blood reveals distinct abnormalities in thrombus formation in patients with severe combined immune deficiency, Glanzmann's thrombasthenia, Hermansky-Pudlak syndrome, May-Hegglin anomaly or grey platelet syndrome.We suggest this test may be useful for the diagnosis of patients with suspected bleeding disorders or a pro-thrombotic tendency.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), Maastricht University, Universiteitssingel 50, 6229 ER Maastricht, The Netherlands.

ABSTRACT
Assays measuring platelet aggregation (thrombus formation) at arterial shear rate mostly use collagen as only platelet-adhesive surface. Here we report a multi-surface and multi-parameter flow assay to characterize thrombus formation in whole blood from healthy subjects and patients with platelet function deficiencies. A systematic comparison is made of 52 adhesive surfaces with components activating the main platelet-adhesive receptors, and of eight output parameters reflecting distinct stages of thrombus formation. Three types of thrombus formation can be identified with a predicted hierarchy of the following receptors: glycoprotein (GP)VI, C-type lectin-like receptor-2 (CLEC-2)>GPIb>α6β1, αIIbβ3>α2β1>CD36, α5β1, αvβ3. Application with patient blood reveals distinct abnormalities in thrombus formation in patients with severe combined immune deficiency, Glanzmann's thrombasthenia, Hermansky-Pudlak syndrome, May-Hegglin anomaly or grey platelet syndrome. We suggest this test may be useful for the diagnosis of patients with suspected bleeding disorders or a pro-thrombotic tendency.

Show MeSH
Related in: MedlinePlus