Control of thrombus embolization and fibronectin internalization by integrin αIIbβ3 engagement of the fibrinogen γ chain

Abstract

Fibrin(ogen) deficiency (Fg -/-) was shown previously to be compatible with rapid thrombus growth within injured arterioles, but platelet fibronectin content was increased and newly formed thrombi were unstable. To further define the role of fibrin(ogen) in thrombus formation and stabilization, platelet biology was examined in mice expressing a form of fibrinogen that clots normally but lacks the ␥ chain C-terminal binding site for ␣IIb␤3 (Fg␥⌬5). Thrombus growth within the arterioles of Fg␥⌬5 mice appeared faster than in wild-type mice despite a far greater emboli formation. Unlike Fg -/-mice, the emboli were relatively small and released from the top of thrombi, rather than by fracture at the vessel wall. The fibronectin content in Fg␥⌬5 platelets was also dramatically increased through a ␤3 integrindependent mechanism. The following has been concluded: (1) Fibrin formation contributes to, but is not sufficient for, the stabilization of arterial thrombi. Platelet receptor engagement of the C-terminal of the Fg ␥ chain contributes to the stable incorporation of platelets into thrombi. IntroductionArrest of bleeding at the site of injury is mediated by the adhesion and aggregation of platelets and the formation of the polymerized fibrin matrix. The same general process contributes to the generation of inopportune thrombi within atherosclerotic arteries. Thrombosis in coronary or cerebral arteries is a major cause of morbidity and mortality worldwide. It has been demonstrated that platelet membrane glycoprotein (GP) Ib complex and its ligand, von Willebrand factor (VWF), are involved in initiating platelet adhesion, particularly at high shear. 1 Subsequent stable platelet adhesion and aggregation are mediated by several integrin receptors and their ligands, such as ␣2␤1/collagen and ␣IIb␤3/fibrinogen (Fg) or VWF. 1,2 The GPIb complex is also involved in platelet aggregation. [3][4][5] In contrast to static or low shear conditions, where fibrinogen is necessary for platelet aggregation, 6,7 at high shear efficient platelet aggregation can occur independently of Fg 8-10 and even independently of both VWF and Fg. 10 Under these conditions, fibronectin (Fn) 11 and perhaps other molecules are able to support platelet aggregation. However, fibrinogen and the local conversion of fibrinogen to fibrin by thrombin are generally thought to be indispensable in stabilizing the thrombus. 9,10 Fibrinogen is a 340-kDa glycoprotein dimer consisting of 2 sets of 3 polypeptide chains, termed the A␣, B␤, and ␥ chains. Fibrinogen both provides the fundamental building block for assembly of provisional fibrin matrices and supports cell adhesion through specific integrin and nonintegrin receptor binding motifs. The C-terminal of the ␥ chain contains a critical binding site for the platelet integrin receptor, ␣IIb␤3 (GPIIbIIIa), which has been demonstrated to be required for platelet aggregation. 12,13 Fg from Fg␥⌬5 mice, carrying the genetically modified form of the ␥ chain gene, which eliminates the last 5 residues (QAGDV) of the ␥ ...

Plasma fibronectin depletion enhances platelet aggregation and thrombus formation in mice lacking fibrinogen and von Willebrand factor

“…4,21 Injury was induced by topical application of 30 L of 250 mM FeCl 3 , and the characteristics of thrombus formation were compared based on (1) number of fluorescent platelets deposited on the vessel wall during the first 3 to 5 minutes after injury, (2) time required for the formation of the first 20-m thrombus, and (3) time to complete vessel occlusion. 4,7,21,25 Cremaster arterial thrombosis model. To quantitatively study platelet accumulation within growing thrombi after the vascular injury in vivo, we used a laser to induce an arteriole thrombus in the cremaster muscle as previously described.…”

Therapeutic approaches in arterial thrombosis

“…2) [23,24]. While these results may have a simple explanation, resulting from the inhibition of fibrin formation by the anticoagulant, where fibrin contributes to thrombus stability under arterial shear rate [25], the emerging data indicate that the anti-thrombotic synergisms may originate from the complementation of the signaling pathways in platelets. Activation of the receptor function of GP IIb-IIIa is optimal when engagement of G 12/13 or G q signaling pathways is combined with G i stimulation [26,27].…”

The Glycoprotein VI-Phospholipase Cγ2 Signaling Pathway Controls Thrombus Formation Induced by Collagen and Tissue Factor In Vitro and In Vivo

“…Furthermore, in vivo experiments where thrombus formation was induced with free radical-forming FeCl 3 allowed us to study the importance of this process in a thrombosis model known to rely on thrombin generation and coagulation. 16 The data showed that also in vivo the signaling cascade from GPVI to PLC␥2 led to coagulant activity and enhanced thrombus formation. …”