Project 4. Platelet accumulation is both a hallmark of hemostasis and a contributor to heart attacks and strokes. Previous studies have focused on identifying individual signaling molecules that support platelet activation. Here we attempt a paradigm shift, first, by approaching platelet activation and the hemostatic response as the product of the global platelet signaling network rather than any one pathway, and, second, by linking testable ideas about the function of that network to real time observations of platelet activation in vivo and in vitro. We and others have shown that hemostatic thrombi have a characteristic structure in which a core of fully-activated platelets is overlaid with an unstable shell of less-activated platelets. We have recently extended this model by showing that the core and shell are regulated by different elements of the platelet signaling network and demonstrating that increased packing density in the core affects thrombin distribution, contact-dependent signaling and the influx of plasma-borne molecules. The proposed studies will build upon this background, merging mechanism-driven and observational approaches to determine how thrombus structure evolves define its relationship the platelet signaling network.
In Aim #1 we will use intravital confocal and multi-photon microscopy to compare platelet activation in the micro- and macro-vasculature. Dyslipidemia will be used to study the impact of acquired disorders of platelet reactivity.
In Aim #2 we will use human platelets, transgenic mouse lines and pharmacologic agents to examine the relationships between the platelet signaling network and thrombus structure.
In Aim #3 we will focus on contact-dependent signaling events between platelets, testing the novel hypothesis that these events are segregated spatially and temporally into pathways that either promote or restrain the thrombus core. Through these aims, we hope to define the mechanisms that drive thrombus formation, account for differences in the clinical impact and bleeding risk associated with different antiplatelet agents, and show how pathological conditions can subvert normal responses by their impact on the platelet signaling network

Public Health Relevance

Platelet activation is part of the normal response to vascular injury, producing a plug that limits blood loss. Thrombosis occurs when platelets are activated inappropriately, blocking blood flow and damaging tissues such as the heart and brain. The goal of this project is to better understand the molecular basis of platelet activation and translate that understanding into improved methods to prevent thrombosis..

National Institute of Health (NIH)
National Heart, Lung, and Blood Institute (NHLBI)
Research Program Projects (P01)
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Heart, Lung, and Blood Initial Review Group (HLBP)
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University of Pennsylvania
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Tomaiuolo, Maurizio; Stalker, Timothy J; Welsh, John D et al. (2014) A systems approach to hemostasis: 2. Computational analysis of molecular transport in the thrombus microenvironment. Blood 124:1816-23
Welsh, John D; Stalker, Timothy J; Voronov, Roman et al. (2014) A systems approach to hemostasis: 1. The interdependence of thrombus architecture and agonist movements in the gaps between platelets. Blood 124:1808-15
Lian, Lurong; Suzuki, Aae; Hayes, Vincent et al. (2014) Loss of ATE1-mediated arginylation leads to impaired platelet myosin phosphorylation, clot retraction, and in vivo thrombosis formation. Haematologica 99:554-60
Stalker, Timothy J; Welsh, John D; Tomaiuolo, Maurizio et al. (2014) A systems approach to hemostasis: 3. Thrombus consolidation regulates intrathrombus solute transport and local thrombin activity. Blood 124:1824-31
Stalker, Timothy J; Welsh, John D; Brass, Lawrence F (2014) Shaping the platelet response to vascular injury. Curr Opin Hematol 21:410-7
Kowalska, M Anna; Zhao, Guohua; Zhai, Li et al. (2014) Modulation of protein C activation by histones, platelet factor 4, and heparinoids: new insights into activated protein C formation. Arterioscler Thromb Vasc Biol 34:120-6
Min, Sang H; Suzuki, Aae; Stalker, Timothy J et al. (2014) Loss of PIKfyve in platelets causes a lysosomal disease leading to inflammation and thrombosis in mice. Nat Commun 5:4691
Brass, Lawrence F; Tomaiuolo, Maurizio; Stalker, Timothy J (2013) Harnessing the platelet signaling network to produce an optimal hemostatic response. Hematol Oncol Clin North Am 27:381-409
Lu, Qiongyu; Dong, Ningzheng; Wang, Qi et al. (2013) Increased levels of plasma soluble Sema4D in patients with heart failure. PLoS One 8:e64265
Stalker, Timothy J; Traxler, Elizabeth A; Wu, Jie et al. (2013) Hierarchical organization in the hemostatic response and its relationship to the platelet-signaling network. Blood 121:1875-85

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