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Thromb Haemost 2020; 120(05): 776-792
DOI: 10.1055/s-0040-1709524
Cellular Haemostasis and Platelets
Georg Thieme Verlag KG Stuttgart · New York

Platelet Activation via Shear Stress Exposure Induces a Differing Pattern of Biomarkers of Activation versus Biochemical Agonists

Yana Roka-Moiia
1   Department of Medicine, Sarver Heart Center, University of Arizona, Tucson, Arizona, United States
,
Ryan Walk
1   Department of Medicine, Sarver Heart Center, University of Arizona, Tucson, Arizona, United States
,
Daniel E. Palomares
1   Department of Medicine, Sarver Heart Center, University of Arizona, Tucson, Arizona, United States
,
Kaitlyn R. Ammann
1   Department of Medicine, Sarver Heart Center, University of Arizona, Tucson, Arizona, United States
,
Annalisa Dimasi
2   Department of Electronics, Information and Bioengineering, Politecnico di Milano, Milan, Italy
,
Joseph E. Italiano
3   Brigham and Woman's Hospital, Harvard Medical School, Boston, Massachusetts, United States
,
Jawaad Sheriff
4   Department of Biomedical Engineering, Stony Brook University, Stony Brook, New York, United States
,
Danny Bluestein
4   Department of Biomedical Engineering, Stony Brook University, Stony Brook, New York, United States
,
Marvin J. Slepian
1   Department of Medicine, Sarver Heart Center, University of Arizona, Tucson, Arizona, United States
4   Department of Biomedical Engineering, Stony Brook University, Stony Brook, New York, United States
› Author Affiliations Funding This work is supported by the National Institute of Health grants U01 EB012487 (to D.B., M.J.S., J.S.) and U01 HL131052 (to D.B., M.J.S., J.S.), and by the Arizona Center for Accelerated Biomedical Innovation of the University of Arizona (to Y.R.M., D.P., K.A., M.J.S.).
Further Information

Publication History

29 August 2019

02 March 2020

Publication Date:
05 May 2020 (online)

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Abstract

Background Implantable cardiovascular therapeutic devices, while hemodynamically effective, remain limited by thrombosis. A driver of device-associated thrombosis is shear-mediated platelet activation (SMPA). Underlying mechanisms of SMPA, as well as useful biomarkers able to detect and discriminate mechanical versus biochemical platelet activation, are poorly defined. We hypothesized that SMPA induces a differing pattern of biomarkers compared with biochemical agonists.

Methods Gel-filtered human platelets were subjected to mechanical activation via either uniform constant or dynamic shear; or to biochemical activation by adenosine diphosphate (ADP), thrombin receptor-activating peptide 6 (TRAP-6), thrombin, collagen, epinephrine, or arachidonic acid. Markers of platelet activation (P-selectin, integrin αIIbβ3 activation) and apoptosis (mitochondrial membrane potential, caspase 3 activation, and phosphatidylserine externalization [PSE]) were examined using flow cytometry. Platelet procoagulant activity was detected by chromogenic assay measuring thrombin generation. Contribution of platelet calcium flux in SMPA was tested employing calcium chelators, ethylenediaminetetraacetic acid (EDTA), and BAPTA-AM.

Results Platelet exposure to continuous shear stress, but not biochemical agonists, resulted in a dramatic increase of PSE and procoagulant activity, while no integrin αIIbβ3 activation occurred, and P-selectin levels remained barely elevated. SMPA was associated with dissipation of mitochondrial membrane potential, but no caspase 3 activation was observed. Shear-mediated PSE was significantly decreased by chelation of extracellular calcium with EDTA, while intracellular calcium depletion with BAPTA-AM had no significant effect. In contrast, biochemical agonists ADP, TRAP-6, arachidonic acid, and thrombin were potent inducers of αIIbβ3 activation and/or P-selectin exposure. This differing pattern of biomarkers seen for SMPA for continuous uniform shear was replicated in platelets exposed to dynamic shear stress via circulation through a ventricular assist device-propelled circulatory loop.

Conclusion Elevated shear stress, but not biochemical agonists, induces a differing pattern of platelet biomarkers—with enhanced PSE and thrombin generation on the platelet surface. This differential biomarker phenotype of SMPA offers the potential for early detection and discrimination from that mediated by biochemical agonists.

Keywords

shear-mediated platelet activation - shear stress - cardiovascular therapeutic device - phosphatidylserine externalization - biomarker

Authors' Contributions

Y.R.-M. designed and performed experiments, analyzed and interpreted data, and wrote the manuscript; R.W., D.E.P., and A.D. performed experiments; K.A. and J.E.I. participated in discussions and manuscript preparation; J.S. and D.B. provided acetylated prothrombin, developed the PAS assay protocol, participated in discussions, and manuscript preparation; M.J.S. designed research, interpreted data, and wrote the manuscript.


Supplementary Material

 

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