P values of 0.05 were considered significant. proteins, and immune cells between the vessel lumen and the interstitial space (Dejana, 2004; Pries and Kuebler, 2006). Dysregulation of endothelial permeability is a hallmark of several inflammatory and vascular diseases and can result in uncontrolled vascular leakage leading to severe fluid loss and organ dysfunction (Mehta and Malik, 2006; Bakker et al., 2009; Lee and Slutsky, 2010). Paracellular permeability of the endothelium can be altered by Nazartinib S-enantiomer soluble factors such as thrombin, bradykinin, TNF-, histamine, and vascular endothelial (VE) growth factor (VEGF; Mehta Nazartinib S-enantiomer and Malik, 2006) through a mechanism that relies on the discrete widening and tightening of endothelial cell (EC)Ccell junctions (Giannotta et al., 2013). Two types of intercellular junctions, namely adherens junctions and tight junctions, are most crucial in regulating the barrier properties of the endothelium. The main molecular component of endothelial adherens junctions is VE-cadherin (Navarro et al., 1998; Dejana, 2004; Giannotta et al., 2013), whereas tight junctions rely on clusters of claudins, occludins, and junction adhesion molecules (Furuse et al., 1993, 1998; Martn-Padura et al., 1998). In addition to cellCcell contacts, the endothelial barrier is also influenced by molecular interactions with the basement membrane through integrins (Zaidel-Bar and Geiger, 2010; Oldenburg and de Rooij, 2014). Finally, a third component, the cytoskeleton, has gained attention as a critical regulator of barrier function. As a dynamic intracellular network of actin fibers, microtubules, and intermediate filaments (Ingber, 2002), the cytoskeleton links junctional complexes and focal adhesions, coordinating tension forces that affect both cell shape and intercellular contacts (Fanning et al., 1998; Giannotta et al., 2013). Adhesive molecules of tight junctions directly interact with zonula occludin proteins (ZO-1, ZO-2, and ZO-3), which anchor the actin cytoskeleton to these junctional complexes (Itoh et al., 1999a,b). Similarly, the cytoplasmic tail of VE-cadherin is connected to the actin bundles via – and -catenin proteins (Dejana, 2004). This association to the actin cytoskeleton is essential for junction assembly, strength, and maintenance (Nelson et al., 2004; Huveneers et al., 2012; Hong et al., 2013). In this manner, the cytoskeleton has the capacity to quickly alter both cellCcell and cellCmatrix interactions. Cytoskeletal organization and dynamics are regulated by Rho GTPases such as RhoA, Rac1, and Cdc42. In turn, these GTPases Gadd45a have major effects on endothelial barrier regulation and permeability (Wojciak-Stothard and Ridley, 2002; Dejana, 2004; Mehta and Malik, 2006; Goddard and Iruela-Arispe, 2013). Traditionally, activation of Rac1 and Cdc42 has been linked to barrier maintenance and stabilization. In contrast, RhoA has been associated with actin stress fiber formation, leading to junctional destabilization and loss of barrier integrity (Amado-Azevedo et al., 2014). Furthermore, other GTPases such as RhoB and Ras-related protein-1 small GTPase (Rap1) have expanded the framework of regulatory proteins that Nazartinib S-enantiomer contribute to barrier function (Cullere et al., 2005; Fukuhara et al., 2005a; Amado-Azevedo et al., 2014). The activation state of small GTPases is controlled by a large number of regulatory proteins that translate various extracellular stimuli into adequate levels of GTPase activity. These include guanosine nucleotide exchange factors (GEFs) that catalyze the activation step of Rho proteins, the GTPase-activating proteins that promote inactivation, and the GDP dissociation inhibitors that regulate the stability and subcellular localization of GTPases depending on the cell stimulation state (Zheng, 2001; Cherfils and Zeghouf, 2013). Thus, 150 GTPase regulatory molecules have been described, including the Vav family of GEFs (Vav1, Vav2, and Vav3; Bustelo, 2014). Despite this, our current understanding of their specific effects on vascular barrier function remains fragmentary (Amado-Azevedo et al., 2014). Importantly, regulation of vascular permeability differs across vascular beds, and the molecular bases for the diversity of organ-specific.
