1. Introduction

Endothelial dysfunction, often as a result of chronic activation, is a primary determinant of the severity of disease states. Apart from other cell-specific defects, activation of endothelial cells (ECs) increases platelet binding to von Willebrand factor (vWF), a multimeric blood protein primarily synthesized, stored and secreted by ECs [1–6]. Under physiological conditions, vWF is secreted from ECs via two pathways that enable hemostasis; a continuous or basal secretory pathway that maintains a baseline blood vWF level, [7] and the other, a regulated secretory

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and eproduction in any medium, provided the original work is properly cited. © 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

pathway induced by agonists such as thrombin [8, 9]. Basal vWF secretion involves elements of regulated secretion, with vWF targeted to Weibel-Palade bodies (WPBs) after Golgi processing, but continuously secreted from the storage pool [7]. Release of WPB contents into the extracellular space upon thrombin stimulation of ECs, thought to be pathophysiologically relevant in inflammatory cardiovascular disease, occurs via several processes: (1) signaling via [Ca2+]/Gαq and Gα12/α-soluble NSF attachment protein (α-SNAP) [10] that (2) promotes engagement and activation of vesicle (v) and target (t) membrane-associated v-SNARE and t-SNARE proteins (vesicle- and target-soluble NSF attachment protein receptors) [11, 12] and (3) energydependent fusion of WPB with plasma membrane that promotes the formation of a secretion pore through which the exocytosis of WPB contents occurs [13–15].

WPBs originate from clathrin and clathrin adaptor-protein1-coated vesicles, [35, 40] which, when loaded with vWF cargo, bud from the trans-Golgi network, continue to mature and vWF contained in them becomes densely pack and highly multimeric vWF [35]. Once the vesicles had formed, they lose the clathrin coat, and the interactions between WPBs and the actin cytoskeleton are thought to either tether WPBs to the perinuclear cytosolic compartment or to relocate WPBs in the proximity of the plasma membrane in preparation for fusion and exocytosis [41]. Upon secretion of its contents, WPBs dissociate from the membrane and are recycled

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Interestingly, vWF secretion is influenced by plasma sodium level [42]. In mice subjected to mild water restriction, elevated endothelial vWF protein levels correlated with increased number of micro thrombi in capillaries [42]. Hypernatremia causes up-regulation of vWF gene expression via nuclear factor of activated T-cells 5 (NFAT5) binding to the vWF promoter [42]. Another regulator of vWF was found to be autophagy [43]. Endothelial cell-specific ATG7 (critical factor in autophagy) deficient mice have prolonged bleeding times as compared to

At least two different secretion mechanisms of WPB cargo have been reported so far: selective, time-dependent 'kiss-and-run' release [13] and differential release of the distinct components of these storage granules [8]. Apart from vWF, WPBs also contain other inflammatory and vasoactive mediators: P-selectin, Rab-27, coagulation factor XIIIa, tissue plasminogen activator, interleukin-8 (IL-8), eotaxin, endothelin-1, endothelin-converting enzyme, calcitonin-generelated peptide, α1,3-fucosyltransferase VI and angiopoietin 2 [44]. vWF and the other components—stockpiled in and liberated as needed from WPBs—provide the ECs with the ability to react acutely to restore homeostasis; vWF plays a role in hemostasis and thrombosis, and also, in the regulation of inflammation [45], vascular permeability [27] and angiogenesis [46].

vWF is synthesized exclusively by ECs, megakaryocytes and platelets and stored in specialized secretory granules. EC WPBs and platelet α-granules share several characteristics, as both organelles contain vWF, P-selectin and CD63 [47, 48]. But, in platelets, vWF detection by immune-gold labeling is restricted to a subset of spherical α-granules [48]. In this vane, spherical secretory α-granules are quite distinct from WPBs. First, the vWF content of platelets is relatively low: 0.05–0.1 U per 3 <sup>10</sup><sup>8</sup> platelets [49]. For comparison, plasma vWF concentration is 1–1.8 U/mL (10 μg/mL blood), of which only 15% of plasma vWF comes from platelets, while the majority 85% of the total vWF comes from ECs [4, 20, 34, 50]. Second, ECs combine basal and regulated vWF release, while platelets do not have a mechanism of basal secretion and, thus, substantial platelet activation would be needed for platelet vWF secretion to significantly contribute to the elevated blood vWF observed in different disease states. Finally, excessive secretion of EC-derived vWF in inflammation is attributed to the large capacity of

Thus, EC secretory granules supply the vasculature with circulating vWF to control hemostasis and thrombosis, while platelets play only a secondary role in this process [20, 49]. A balanced level of EC-derived vWF is critical to overcome the thrombotic complications in pathological settings, therefore it is important to understand the underlying endothelial constitutive and evoked exocytosis mechanisms which provide the bloodstream vWF in health

ECs to make and store vWF in EC-specific storage pool, that is, in WPBs.

control, and decreased vWF secretion of immature vWF [43].

and refilled [35].

and disease.

Once secreted from WPBs, vWF circulates in a globular conformation under resting conditions [16] and is a carrier for circulating coagulation factor VIII, thereby protecting it from degradation [17–19]. The blood vWF level in health and inflammatory disease is predominantly EC derived, while the contribution of platelets is rather minimal [20]. In response to high shear stress and inflammatory mediators [21], normally quiescent ECs secrete long vWF multimers in large quantities from the storage pool into systemic circulation. vWF multimers are then extracellularly cleaved by a metalloprotease ADAMTS-13 (A Disintegrin and Metalloproteinase with a thrombospondin type 1 motif, member 13), unfold in the circulation and self-associate into particularly linked concatemers to form ultra-large vWF strings [22]. Platelets spontaneously bind to activated vWF via glycoprotein Ibα (GpIbα) interaction with the exposed A1 domain, initiating the thrombogenic process [23–26]. In addition to its classical role in platelet binding, the vWF A1 domain also plays a role in vWF-dependent inflammatory responses [27–29]. In this chapter, we will discuss the most recent findings related to vWF secretion from ECs, the finely tuned balance of vWF level and activity in hemostasis and thrombosis, and the mechanisms by which vWF/ADAMTS-13 axis links inflammation and thrombosis. The potential of vWF as a target in antithrombotic therapies is also discussed.
