Abstract Number: PB1962
Meeting: ISTH 2020 Congress
Background: The vascular endothelium has an established antiplatelet and anticoagulant role, acting on flowing blood. To understand the mechanisms and kinetics of these platelet- and coagulation-inhibitory processes in vitro endothelial models need to be developed.
Aims: Establish an endothelialised microfluidic model to determine the spatio-temporal mechanisms of vascular control of collagen- and thrombin-induced platelet activation under flow.
Methods: Human umbilical vein endothelial cells (HUVEC) were cultured on collagen and TF in microfluidic channels, and subjected to whole blood perfusion. Platelet activation after incubation with HUVEC under stasis was monitored from Ca2+ fluxes using Calcium-6 dye.
Results: Culturing of HUVEC on collagen to (sub)confluency under stasis resulted in cell patches containing a glycocalyx and expressing von Willebrand factor (VWF), as shown by multicolor confocal microscopy. When Calcium-6 labelled platelets were incubated with HUVEC, platelet Ca2+ fluxes were suppressed in response to thrombin or collagen receptor agonist CRP-XL, in a time dependent manner (t1/2 = 10 minutes). Similarly, an immediate abrogation of platelet Ca+2 fluxes by stable prostacyclin pointed to the involvement (at least in part) of cAMP/PKA pathway, as confirmed by VASP phosphorylation profiles.
Subjection of the HUVEC, cultured on collagen with(out) tissue factor, to recalficied whole blood in microfluidics resulted in remarkable changes in collagen/VWF-induced platelet activation and fibrin generation, in comparison to channels not containing HUVEC. Shear-dependent platelet adhesion and activation occurred rapidly, but remained strictly confined to the collagen/TF spots in between HUVEC patches. Fibrin formation was delayed, and again only occurred in between the HUVEC patches. These effects were modified by treatment with thrombin inhibitor or heparinase (glycocalyx degradation).
Conclusions: The developed endothelial model revealed that HUVEC patches rapidly, but only locally, suppressed collagen- and thrombin-induced platelet activation under flow, thus pointing to rapid offset and onset mechanisms. Local control is provided by cAMP elevation and coagulation suppression.
To cite this abstract in AMA style:Provenzale I, Brouns SLN, Fernández DI, van der Meijden PEJ, Heemskerk JWM. Vessel-On-A-Chip Model Revealing Rapid Offset and Onset Mechanisms of Collagen- and Thrombin-Induced Platelet Activation [abstract]. Res Pract Thromb Haemost. 2020; 4 (Suppl 1). https://abstracts.isth.org/abstract/vessel-on-a-chip-model-revealing-rapid-offset-and-onset-mechanisms-of-collagen-and-thrombin-induced-platelet-activation/. Accessed December 5, 2021.
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