High-throughput elucidation of thrombus formation reveals sources of platelet function variability

Ankie van Geffen, Sanne Brouns, Joana Batista, Harriet McKinney, Carly Kempster, Magdi Nagy, Suthesh Sivapalaratnam, Constance Baaten, Nikki Bourry, Mattia Frontini, Kerstin Jurk, Manuela Krause, Daniele Pilliteri, Frauke Swieringa, Remco Verdoold, Rachel Cavill, Marijke Kuijpers, Willem H. Ouwehand, Kate Downes, Johan Heemskerk

Research output: Contribution to journalArticleAcademicpeer-review

Abstract

In combination with microspotting, whole-blood microfluidics can provide high-throughput information on multiple platelet functions in thrombus formation. Based on assessment of the inter-and intra-subject variability in parameters of microspot-based thrombus formation, we aimed to determine the platelet factors contributing to this variation. Blood samples from 94 genotyped healthy subjects were analyzed for conventional platelet phenotyping: i.e. hematologic parameters, platelet glycoprotein (GP) expression levels and activation markers (24 parameters). Furthermore, platelets were activated by ADP, CRP-XL or TRAP. Parallel samples were investigated for whole-blood thrombus formation (6 microspots, providing 48 parameters of adhesion, aggregation and activation). Microspots triggered platelet activation through GP Ib-V-IX, GPVI, CLEC-2 and integrins. For most thrombus parameters, inter-subject variation was 2-4 times higher than the intra-subject variation. Principal component analyses indicated coherence between the majority of parameters for the GPVI-dependent microspots, partly linked to hematologic parameters, and glycoprotein expression levels. Prediction models identified parameters per microspot that were linked to variation in agonist-induced alpha(IIb)beta(3) activation and secretion. Common sequence variation of GP6 and FCER1G, associated with GPVI-induced alpha(IIb)beta(3) activation and secretion, affected parameters of GPVI-and CLEC-2-dependent thrombus formation. Subsequent analysis of blood samples from patients with Glanzmann thrombasthenia or storage pool disease revealed thrombus signatures of aggregation-dependent parameters that were subject-dependent, but not linked to GPVI activity. Taken together, this high-throughput elucidation of thrombus formation revealed patterns of inter-subject differences in platelet function, which were partly related to GPVI-induced activation and common genetic variance linked to GPVI, but also included a distinct platelet aggregation component.

Original languageEnglish
Pages (from-to)1256-1267
Number of pages12
JournalHaematologica-the Hematology Journal
Volume104
Issue number6
Early online date13 Dec 2018
DOIs
Publication statusPublished - 31 May 2019

Keywords

  • DISORDERS
  • RESPONSES
  • DIFFERENCE
  • ADHESION

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