Shear stress is a fundamental determinant of vascular homeostasis, regulating vascular remodelling, cardiac development and atherogenesis, but the mechanisms of transduction are poorly understood. Previous work showed that the conversion of integrins to a high-affinity state mediates a subset of shear responses, including cell alignment and gene expression. Here we investigate the pathway upstream of integrin activation. PECAM-1 (which directly transmits mechanical force), vascular endothelial cell cadherin (which functions as an adaptor) and VEGFR2 (which activates phosphatidylinositol-3-OH kinase) comprise a mechanosensory complex. Together, these receptors are sufficient to confer responsiveness to flow in heterologous cells. In support of the relevance of this pathway in vivo, PECAM-1-knockout mice do not activate NF-kappaB and downstream inflammatory genes in regions of disturbed flow. Therefore, this mechanosensing pathway is required for the earliest-known events in atherogenesis.

Vascular endothelial cadherin, VE-cadherin, mediates adhesion between endothelial cells and may affect vascular morphogenesis via intracellular signaling, but the nature of these signals remains unknown. Here, targeted inactivation (VEC-/-) or truncation of the beta-catenin-binding cytosolic domain (VECdeltaC/deltaC) of the VE-cadherin gene was found not to affect assembly of endothelial cells in vascular plexi, but to impair their subsequent remodeling and maturation, causing lethality at 9.5 days of gestation. Deficiency or truncation of VE-cadherin induced endothelial apoptosis and abolished transmission of the endothelial survival signal by VEGF-A to Akt kinase and Bcl2 via reduced complex formation with VEGF receptor-2, beta-catenin, and phosphoinositide 3 (PI3)-kinase. Thus, VE-cadherin/ beta-catenin signaling controls endothelial survival.

Tight junctions are the most apical components of endothelial and epithelial intercellular cleft. In the endothelium these structures play an important role in the control of paracellular permeability to circulating cells and solutes. The only known integral membrane protein localized at sites of membrane–membrane interaction of tight junctions is occludin, which is linked inside the cells to a complex network of cytoskeletal and signaling proteins. We report here the identification of a novel protein (junctional adhesion molecule [JAM]) that is selectively concentrated at intercellular junctions of endothelial and epithelial cells of different origins. Confocal and immunoelectron microscopy shows that JAM codistributes with tight junction components at the apical region of the intercellular cleft. A cDNA clone encoding JAM defines a novel immunoglobulin gene superfamily member that consists of two V-type Ig domains. An mAb directed to JAM (BV11) was found to inhibit spontaneous and chemokine-induced monocyte transmigration through an endothelial cell monolayer in vitro. Systemic treatment of mice with BV11 mAb blocked monocyte infiltration upon chemokine administration in subcutaneous air pouches. Thus, JAM is a new component of endothelial and epithelial junctions that play a role in regulating monocyte transmigration.

Intercellular junctions mediate adhesion and communication between adjoining endothelial and epithelial cells. In the endothelium, junctional complexes comprise tight junctions, adherens junctions, and gap junctions. The expression and organization of these complexes depend on the type of vessels and the permeability requirements of perfused organs. Gap junctions are communication structures, which allow the passage of small molecular weight solutes between neighboring cells. Tight junctions serve the major functional purpose of providing a "barrier" and a "fence" within the membrane, by regulating paracellular permeability and maintaining cell polarity. Adherens junctions play an important role in contact inhibition of endothelial cell growth, paracellular permeability to circulating leukocytes and solutes. In addition, they are required for a correct organization of new vessels in angiogenesis. Extensive research in the past decade has identified several molecular components of the tight and adherens junctions, including integral membrane and intracellular proteins. These proteins interact both among themselves and with other molecules. Here, we review the individual molecules of junctions and their complex network of interactions. We also emphasize how the molecular architectures and interactions may represent a mechanistic basis for the function and regulation of junctions, focusing on junction assembly and permeability regulation. Finally, we analyze in vivo studies and highlight information that specifically relates to the role of junctions in vascular endothelial cells.

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endothelial growth factor (VEGF), induce the tyrosine phosphorylation of VE-cadherin, which accompanies an increase in vascular permeability and leukocyte diapedesis; in addition, the internalization and cleavage of VE-cadherin can cause AJs to be dismantled. From the knowledge of how AJ organization can be modulated, it is possible to formulate several pharmacological strategies to control the barrier function of the endothelium. We discuss the possible use of inhibitors of SRC and other kinases, of agents that increase cAMP levels, and of inhibitors of lytic enzymes as pharmacological tools for decreasing endothelial permeability. Journal of Cell Science 2116 paracellular permeability is governed by the opening and closing of cell junctions, which implies that a complex rearrangement of adhesion proteins and the related cytoskeleton must occur. It is likely that the two pathways are interconnected in some way, because many permeability-increasing agents increase vesicular transport and also disrupt the integrity of endothelial cell-cell junctions (Dejana, 2004;Feng et al., 1999;Weis and Cheresh, 2005); however, whether this occurs in the same vessels and at the same time is still a matter of debate. It is possible that, in some areas of the vasculature, such as the microvasculature of the glands, the transcellular pathway is better developed, whereas in others, such as the postcapillary venules, the paracellular pathway is favored. It will be of interest, once more knowledge is available on the vesicular-transport systems in endothelial cells, to try to integrate both systems into a more comprehensive picture. This Commentary focuses on the role of cell-cell junctionsparticularly adherens junctions (AJs) -in the barrier function of the endothelium. We pay particular attention to the function, molecular organization and regulation of vascular endothelial (VE)-cadherin, the endothelium-specific transmembrane component of AJs. Finally, we show that pharmacological strategies for modulating endothelial permeability can be obtained by using our knowledge of the structure and function of AJs. Adherens junctions and vascular integrityPrevious work has described the molecular organization of the different types of endothelial cell-cell junctions, and has established a basis for understanding how these structures might crosstalk and reciprocally interact (Bazzoni and Dejana, 2004;Muller, 2003;Vestweber, 2007;Wallez and Huber, 2007). Endothelial-cell junctions present a particularly complex network of adhesion proteins that are linked to intracellular cytoskeletal and signaling partners. These proteins are organized into distinct structures called tight junctions (TJs) and AJs. In addition, several adhesion proteins [such as platelet endothelial cell adhesion molecule (PECAM1), MUC18, intercellular adhesion molecule 2 (ICAM2), CD34, endoglin and others] cluster at cell-cell contacts that are distinct from TJs and AJs. There are data to suggest that the assembly of AJs is required for the correct organization of TJs, but...

SUMMARY A key function of blood vessels, to supply oxygen, is impaired in tumors because of abnormalities in their endothelial lining. PHD proteins serve as oxygen sensors and may regulate oxygen delivery. We therefore studied the role of endothelial PHD2 in vessel shaping by implanting tumors in PHD2+/− mice. Haplodeficiency of PHD2 did not affect tumor vessel density or lumen size, but normalized the endothelial lining and vessel maturation. This resulted in improved tumor perfusion and oxygenation and inhibited tumor cell invasion, intravasation, and metastasis. Haplodeficiency of PHD2 redirected the specification of endothelial tip cells to a more quiescent cell type, lacking filopodia and arrayed in a phalanx formation. This transition relied on HIF-driven upregulation of (soluble) VEGFR-1 and VE-cadherin. Thus, decreased activity of an oxygen sensor in hypoxic conditions prompts endothelial cells to readjust their shape and phenotype to restore oxygen supply. Inhibition of PHD2 may offer alternative therapeutic opportunities for anticancer therapy.

The blood–brain barrier (BBB) is confined to the endothelium of brain capillaries and is indispensable for fluid homeostasis and neuronal function. In this study, we show that endothelial Wnt/β-catenin (β-cat) signaling regulates induction and maintenance of BBB characteristics during embryonic and postnatal development. Endothelial specific stabilization of β-cat in vivo enhances barrier maturation, whereas inactivation of β-cat causes significant down-regulation of claudin3 (Cldn3), up-regulation of plamalemma vesicle-associated protein, and BBB breakdown. Stabilization of β-cat in primary brain endothelial cells (ECs) in vitro by N-terminal truncation or Wnt3a treatment increases Cldn3 expression, BBB-type tight junction formation, and a BBB characteristic gene signature. Loss of β-cat or inhibition of its signaling abrogates this effect. Furthermore, stabilization of β-cat also increased Cldn3 and barrier properties in nonbrain-derived ECs. These findings may open new therapeutic avenues to modulate endothelial barrier function and to limit the devastating effects of BBB breakdown.