PB1271 - Multiparameter analysis of the time-dependent roles of Platelet and Coagulation Pathways in Collagen- and Tissue Factor-Induced Thrombus Formation

Background: In hemostasis and thrombosis, the complex process of thrombus formation involves different receptors and molecular pathways of platelet and coagulation activation. These pathways are considered to operate continuously together at the same time, but this has not been fully investigated.

Aims: To elucidate the time-dependency of key pathways of thrombus and clot formation, initiated by collagen and tissue factor surfaces, where coagulation is triggered via the extrinsic route.

Methods: We adapted a microfluidics whole-blood assay with the Maastricht flow chamber to acutely block molecular pathways involved in thrombus formation by pharmacological intervention at desired time points.

Results: The microfluidic system adapted ad hoc for this study revealed crucial roles of glycoprotein VI (GPVI)-induced platelet signaling via Syk kinase as well as factor VIIa-induced thrombin generation, both of which were confined to the first minutes of thrombus formation. In contrast, platelet activation via the protease-activating receptors 1/4 (PAR1/4) and integrin αIIbβ3 activity appeared to be prolongedly active and extended to later stages of thrombus and clot formation. Finally, the data also pointed to a pivotal role of GPVI and the ITAM signaling in the generation of a procoagulant surface on platelets and therefore the enhancement of thrombin generation and consequently fibrin deposition at level of the thrombus surface.

Conclusion(s): This study presents a new efficient adaptation of the Maastricht microfluidic system, allowing disclosure of the time-dependent roles of key pathways of platelet activation and coagulation in the thrombus development. This work thereby revealed a more persistent contribution of thrombin receptor-induced platelet activation than of collagen receptor-induced platelet activation to the thrombotic process.