CC BY-NC-ND 4.0 · Thromb Haemost 2022; 122(05): 726-738
DOI: 10.1055/s-0041-1735972
Cellular Haemostasis and Platelets

Effects of Platelet Agonists and Priming on the Formation of Platelet Populations

Alicia Veninga
1   Department of Biochemistry, Cardiovascular Research Institute Maastricht, Maastricht University, The Netherlands
,
Constance C. F. M. J. Baaten
1   Department of Biochemistry, Cardiovascular Research Institute Maastricht, Maastricht University, The Netherlands
2   Institute for Molecular Cardiovascular Research, University Hospital Aachen, RWTH Aachen University, Germany
,
Ilaria De Simone
1   Department of Biochemistry, Cardiovascular Research Institute Maastricht, Maastricht University, The Netherlands
3   Institute for Cardiovascular and Metabolic Research, University of Reading, Reading, United Kingdom
,
Bibian M. E. Tullemans
1   Department of Biochemistry, Cardiovascular Research Institute Maastricht, Maastricht University, The Netherlands
,
1   Department of Biochemistry, Cardiovascular Research Institute Maastricht, Maastricht University, The Netherlands
4   Thrombosis Expertise Center, Heart and Vascular Center, Maastricht University Medical Center, Maastricht, The Netherlands
,
1   Department of Biochemistry, Cardiovascular Research Institute Maastricht, Maastricht University, The Netherlands
,
Paola E. J. van der Meijden
1   Department of Biochemistry, Cardiovascular Research Institute Maastricht, Maastricht University, The Netherlands
4   Thrombosis Expertise Center, Heart and Vascular Center, Maastricht University Medical Center, Maastricht, The Netherlands
› Author Affiliations
Funding This work is supported by the Landsteiner Foundation for Blood Transfusion Research Grant No. 1711. C.C.F.M.J.B. received funding from The Dutch Heart Foundation (2020T020) and the START-Program of the Faculty of Medicine at the RWTH Aachen University (105/20). I.D.S. is supported by a joint PhD scholarship of Maastricht and Reading Universities, and by the European Union's Horizon 2020 research and innovation program under the Marie Sklodowska-Curie grant agreement No. 766118.

Abstract

Platelets from healthy donors display heterogeneity in responsiveness to agonists. The response thresholds of platelets are controlled by multiple bioactive molecules, acting as negatively or positively priming substances. Higher circulating levels of priming substances adenosine and succinate, as well as the occurrence of hypercoagulability, have been described for patients with ischaemic heart disease. Here, we present an improved methodology of flow cytometric analyses of platelet activation and the characterisation of platelet populations following activation and priming by automated clustering analysis.

Platelets were treated with adenosine, succinate, or coagulated plasma before stimulation with CRP-XL, 2-MeSADP, or TRAP6 and labelled for activated integrin αIIbβ3 (PAC1), CD62P, TLT1, CD63, and GPIX. The Super-Enhanced Dmax subtraction algorithm and 2% marker (quadrant) setting were applied to identify populations, which were further defined by state-of-the-art clustering techniques (tSNE, FlowSOM).

Following activation, five platelet populations were identified: resting, aggregating (PAC1 + ), secreting (α- and dense-granules; CD62P + , TLT1 + , CD63 + ), aggregating plus α-granule secreting (PAC1 + , CD62P + , TLT1 + ), and fully active platelet populations. The type of agonist determined the distribution of platelet populations. Adenosine in a dose-dependent way suppressed the fraction of fully activated platelets (TRAP6 > 2-MeSADP > CRP-XL), whereas succinate and coagulated plasma increased this fraction (CRP-XL > TRAP6 > 2-MeSADP). Interestingly, a subset of platelets showed a constant response (aggregating, secreting, or aggregating plus α-granule secreting), which was hardly affected by the stimulus strength or priming substances.

Supplementary Material



Publication History

Received: 14 December 2020

Accepted: 12 July 2021

Article published online:
24 October 2021

© 2021. The Author(s). This is an open access article published by Thieme under the terms of the Creative Commons Attribution-NonDerivative-NonCommercial License, permitting copying and reproduction so long as the original work is given appropriate credit. Contents may not be used for commercial purposes, or adapted, remixed, transformed or built upon. (https://creativecommons.org/licenses/by-nc-nd/4.0/)

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