Platelet alpha(llb)beta(3) Integrin and Myosin II ATPase Guide Fibrin Network Structure and Enhance Clot Stiffness During Clot Formation
Brackett, Nicole, Biomedical Engineering - School of Engineering and Applied Science, University of Virginia
Lawrence, Michael, Department of Biomedical Engineering, University of Virginia
At the site of vascular injury, coagulation is critical for the restoration of hemostasis. Despite the life-preserving function of coagulation, clotting abnormalities are associated with numerous vascular disorders including myocardial infarction, stroke, and hemophilia. Platelets, the primary cellular component of blood clots, drive clot progression through interactions with the clot fibrin network by platelet αIIbβ3 integrin-mediated adhesion to fibrin and cytoskeleton-mediated tensioning of the fibrin network.
Previous research has identified clot stiffness as a valuable metric for quantifying hypercoagulability or hypocoagulability affiliated with vascular disease. Similarly, previous studies have used features of abnormal clot structure to identify patients at risk of thrombotic events and to evaluate the efficacy of anti-thrombotic drugs. Given the prominent function of the platelet αIIbβ3 integrin and the platelet cytoskeleton in coagulation, and given the value of clot structure and stiffness as metrics of vascular disease, this study aimed to quantify the contribution of the platelet αIIbβ3 integrin and platelet cytoskeletal myosin II ATPase to fibrin network structure and clot stiffness during clot formation. We hypothesized that the platelet αIIbβ3 integrin and platelet myosin II ATPase enhance clot stiffness and regulate fibrin network structure. Additionally, we hypothesized that a network model of stiffness would find that platelet-mediated changes to fibrin network structure could be used to predict changes in clot stiffness.
To test our hypothesis, the contribution of the platelet αIIbβ3 integrin and platelet myosin II ATPase to whole blood and plasma clot stiffness was evaluated using sonorheometry and electromagnetic force techniques. The results found that inhibition of the platelet αIIbβ3 integrin or platelet myosin II ATPase significantly attenuated clot stiffness, indicating that platelet αIIbβ3 integrin-mediated binding to the fibrin network and myosin II ATPase-mediated tensioning of the fibrin network are critical components of clot stiffening during coagulation.
Next, to evaluate if changes in clot stiffness could be linked to platelet-mediated changes in clot structure, we used confocal microscopy and image analysis to quantify the contribution of the platelet αIIbβ3 integrin and platelet myosin II ATPase to fibrin network structure. The results found that inhibition of the platelet αIIbβ3 integrin or platelet myosin II ATPase produced clots with longer fibrin fibers, fewer fiber junctions, increased clot porosity, and reduced clot heterogeneity, suggesting that platelet αIIbβ3 integrin-mediated binding to the clot fibrin network and myosin II ATPase-mediated tensioning of the network are critical modulators of fibrin network structure during coagulation. Finally, the application of a simple network model found that platelet αIIbβ3 integrin- and platelet myosin II ATPase-mediated changes in fibrin network structure are useful predictors of relative changes in clot stiffness.
Quantifiable features of fibrin network structure and clot mechanical stability are a valuable means to guide the detection of vascular disease. In this study, we demonstrated that platelet myosin II ATPase is a critical modulator of clot stiffening during coagulation. In addition, we quantified the contribution of the platelet αIIbβ3 integrin and platelet myosin II ATPase to specific structural features of the clot fibrin network, and suggest that the platelet-mediated changes in clot structure could predict changes in clot stiffness. Finally, the study highlighted the utility of existing technologies for measuring clot structure and strength (confocal microscopy and sonorheometry), and presented a novel technique that offers the potential for simultaneous assessment of clot structure and stiffness (electromagnetic force).
PHD (Doctor of Philosophy)
Blood coagulation, Platelet, Fibrin network