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Ferrata Storti Foundation
Haematologica 2019 Volume 104(6):1256-1267
Platelet Biology & Its Disorders
High-throughput elucidation of thrombus formation reveals sources of platelet function variability
Johanna P. van Geffen,1 Sanne L.N. Brouns,1 Joana Batista,2,3 Harriet
McKinney,2,3 Carly Kempster,2,3 Magdolna Nagy,1 Suthesh Sivapalaratnam,2,4
Constance C.F.M.J. Baaten,1 Nikki Bourry,1 Mattia Frontini,2,3,5 Kerstin Jurk,6
Manuela Krause,7 Daniele Pillitteri,7 Frauke Swieringa,1 Remco Verdoold,1
Rachel Cavill,8 Marijke J. E. Kuijpers,1 Willem H. Ouwehand,2,3,5,9,10 Kate
Downes
2,3,9* 1* and Johan W.M. Heemskerk
1Department of Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), Maastricht University, the Netherlands; 2Department of Haematology, University of Cambridge, Cambridge Biomedical Campus, UK; 3National Health Service Blood and Transplant (NHSBT), Cambridge Biomedical Campus, UK; 4The Royal London Haemophilia Centre, London, UK; 5BHF Centre of Excellence, Division of Cardiovascular Medicine, Cambridge University Hospitals, Cambridge Biomedical Campus, UK; 6Center for Thrombosis and Hemostasis (CTH), University Medical Center of the Johannes Gutenberg University Mainz, Germany; 7DKD Helios Klinik Wiesbaden, Germany; 8Department of Data Science & Knowledge Engineering, Faculty of Humanities and Sciences, Maastricht University, the Netherlands; 9NIHR BioResource, University of Cambridge, Cambridge Biomedical Campus, UK and 10Department of Human Genetics, The Wellcome Sanger Institute, Hinxton, Cambridge, UK
ABSTRACT
In combination with microspotting, whole-blood microfluidics can pro- vide 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 convention- al 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 trig- gered 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 αIIbb3 activation and secre- tion. Common sequence variation of GP6 and FCER1G, associated with GPVI-induced αIIbb3 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 togeth- er, this high-throughput elucidation of thrombus formation revealed pat- terns 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.
*KD and JWMH contributed equally to this work.
Correspondence:
JOHAN W. M. HEEMSKERK
jwm.heemskerk@maastrichtuniversity.nl
KATE DOWNES
kd286@cam.ac.uk
Received: May 31, 2018.
Accepted: December 5, 2018. Pre-published: December 13, 2018.
doi:10.3324/haematol.2018.198853
Check the online version for the most updated information on this article, online supplements, and information on authorship & disclosures: www.haematologica.org/content/104/6/1256
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