Multiple integrin-ligand interactions synergize in shear-resistant platelet adhesion at sites of arterial injury in vivo

Abstract

Damage to the integrity of the vessel wall results in exposure of the subendothelial extracellular matrix (ECM), which triggers integrin-dependent adhesion and aggregation of platelets. The role of platelet ␤1 integrins in these processes remains mostly undefined. Here, we demonstrate by intravital fluorescence microscopy that platelet adhesion and thrombus growth on the exposed ECM of the injured carotid artery is not significantly altered in ␣2-null mice and even in mice with a Cre/loxP-mediated loss of all ␤1 integrins on their platelets. In contrast, inhibition of ␣IIb␤3 integrin on platelets in wild-type mice blocked aggregate formation and reduced platelet adhesion by 60.0%. Strikingly, ␣IIb␤3 inhibition had a comparable effect in ␣2-null mice, demonstrating that other receptors mediate shear-resistant adhesion in the absence of functional ␣2␤1 and ␣IIb␤3. These were identified to be ␣5␤1 and/or ␣6␤1 as ␣IIb␤3 inhibition abrogated platelet adhesion in ␤1-null mice. We conclude that shear-resistant platelet adhesion on the injured vessel wall in vivo is a highly integrated process involving multiple integrin-ligand interactions, none of which by itself is essential. IntroductionAt sites of vascular injury, the subendothelial extracellular matrix (ECM) is exposed to the flowing blood, which triggers adhesion and aggregation of platelets. 1 This process is crucial to limit posttraumatic blood loss but may also lead to occlusion of diseased vessels and infarction of vital organs. Integrins are the major class of receptors that mediate firm adhesion and aggregation of platelets. Integrins are heterodimeric transmembrane receptors composed of an ␣ and a ␤ subunit. 2,3 Platelets express integrins of the ␤1 and the ␤3 family that are present on the membrane in a low-affinity state. Once the platelets become activated the integrins shift to a high-affinity state and efficiently bind their ligands. 2,[4][5][6] The subendothelial ECM contains many different macromolecular constituents that are suitable substrates for integrin-mediated platelet adhesion including collagens, laminins, fibronectin, and von Willebrand factor (VWF). Collagen has a central role as it not only supports platelet adhesion but also strongly activates the cells. 7,8 This activation is mediated by the low-affinity collagen receptor glycoprotein VI (GPVI). 9 GPVI belongs to the immunoglobulin (Ig) superfamily 10 and is noncovalently associated with the signal-transducing FcR␥ chain. 11,12 Under conditions of elevated shear, platelet adhesion on collagen strictly depends on the interaction of GPIb-V-IX with collagen-bound VWF 13 and that of GPVI with collagen. 5 During this process, ligation of GPVI 5 (and GPIb 14 ) leads to platelet activation and the shift of ␤1 and ␤3 integrins to a high-affinity state via "inside-out" signals, enabling the platelet to establish firm adhesion contacts that resist the shear forces in the bloodstream (shear-resistant adhesion) and subsequent thrombus growth. The importance of these initial processes in arterial ...

Red blood cells mediate the onset of thrombosis in the ferric chloride murine model

“…This finding may explain the lack of a major phenotype for collagen receptor glycoprotein VI and integrin a2b1 deficiency observed in some studies. 7,17,18 However, for reasons that remain unclear, other investigators also have reported a severe thrombotic defect for glycoprotein VI deficiency in this model. 13,19 Although they express multiple adhesion molecules, RBCs are not believed to be able to adhere to any other cell types under normal physiological conditions.…”

Platelet receptors and signaling in the dynamics of thrombus formation

“…30 Mice deficient in α2 or β1 have normal bleeding times, and display minor defects on platelet adhesion and aggregation to collagen. 31,32 Studies of experimental in vivo arterial thrombosis in these mice have led to conflicting results showing both unaltered formation of thrombi 32 and mildly delayed, reduced and unstable thrombi. 33,34 Like other integrins, conformational activation of α2β1 increases its affinity for collagen, and seems to require inside-out signaling events that might be driven by engagement of GP VI with collagen 23 and/or by activation of αIIbβ3.…”

Integrins in Wound Healing

“…3; Table 2). [46][47][48][49][50][51][52][53][54][55][56][57][58][59][60][61][62][63][64][65] Integrins can be divided into four subfamilies based on their ligand specificity and/or phylogenetic comparison of the a subunits (Fig. 3).…”