Abstract-Endothelial cell migration is essential to angiogenesis. This motile process is directionally regulated by chemotactic, haptotactic, and mechanotactic stimuli and further involves degradation of the extracellular matrix to enable progression of the migrating cells. It requires the activation of several signaling pathways that converge on cytoskeletal remodeling. Then, it follows a series of events in which the endothelial cells extend, contract, and throw their rear toward the front and progress forward. The aim of this review is to give an integrative view of the signaling mechanisms that govern endothelial cell migration in the context of angiogenesis. (Circ Res. 2007;100:782-794.)Key Words: endothelial cells Ⅲ chemotaxis Ⅲ haptotaxis Ⅲ mechanotaxis Ⅲ migration Ⅲ angiogenesis Ⅲ VEGF Ⅲ integrins Ⅲ actin Ⅲ remodeling E ndothelial cells derive from the successive differentiation of mesodermal cells into hemangioblasts, which leads to the formation of the first vascular structures that are called primitive blood islands. The hemangioblasts from the center of the islands give rise to the hematopoietic stem cells, whereas the peripheral hemangioblasts differentiate into angioblasts, the precursors of mature endothelial cells. Under the influence of vascular endothelial growth factor (VEGF), the angioblasts and newly formed endothelial cells migrate on a matrix constituted mainly of collagen and hyaluronan, allowing the fusion of the blood islands, their remodeling into tubular structures, and the formation of the first primitive vascular plexus. These tubules remodel through vasculogenesis into larger vessels, leading to vascularization of the embryo. 1 In contrast to vasculogenesis, angiogenesis refers to the formation of new blood vessels from preexisting ones ( Figure 1). It is required in many physiological and pathological conditions, including embryonic development, wound healing, tissue regeneration, and tumor growth. The process is regulated by a tight balance between pro-and antiangiogenic agents and involves a cascade of events of which migration of capillary endothelial cells is an essential component. 2 The basic concepts of the cellular and molecular machinery underlying endothelial cell migration have been obtained mostly from observations of cell culture systems (see the online data supplement, available at http://circres.ahajournals.org). However, the 3D environment within a whole organism is far more complex, and the cells must move between and among other cells while they interpret attractive and repulsive cues to choose their path. They must integrate and coordinate their adhesion with their surroundings and sense when to start and stop moving. Genetic dissection using in particular zebrafish 3,4 and mouse embryo 5 now offers the possibility to understand endothelial cell migration in a physiological context. The aim of this review is to make an integrative update on the signaling and physiological aspects of endothelial cell migration in angiogenesis.Original received October 18, 200...

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VEGFR-2 is the major receptor that regulates the different functions of VEGF in adults. We have previously reported that following VEGF treatment of endothelial cells, VEGFR-2 is phosphorylated on Tyr 1214 upstream of the Cdc42-SAPK2/p38-MAPKAP K2 pathway. However, little is known of the earliest molecular events that compose the SAPK2/p38 pathway following VEGFR-2 activation. In this study, we address this question using HA-tagged constructs of either wild-type VEGFR-2 or Y1214F VEGFR-2 mutant in immunoprecipitation assays. We show that the Src family kinase member Fyn, but not c-Src itself, is recruited to VEGFR-2 and is activated in a pϳTyr 1214 -dependent manner. We also report that the SH2 domain-containing adapter molecule Nck, but not Grb2, is recruited to VEGFR-2 in a pϳTyr 1214 -dependent manner and that it associates with Fyn. Moreover, PAK-2 is phosphorylated in a Fyn-dependent manner. Using chemical and genetic inhibitors, we show that Fyn activity is required for SAPK2/p38 but not for FAK activation in response to VEGF. In contrast, c-Src permits activation of FAK, but not that of SAPK2/p38. In addition, Fyn is required for stress fiber formation and endothelial cell migration. We propose a model in which Fyn forms a molecular complex with Nck and PAK-2 and suggest that this complex assembles in a pϳTyr 1214 -dependent manner within VEGFR-2 following VEGF treatment. In turn, this triggers the activation of the SAPK2/p38 MAP kinase module, and promotes stress fiber formation and endothelial cell migration.

Medial vascular calcification is a common complication of chronic kidney disease (CKD). Although elevated inorganic phosphate stimulates vascular smooth muscle cell (VSMC) osteogenic transdifferentiation and calcification, the mechanisms involved in their calcification during CKD are not fully defined. Because hypoxic gene activation is linked to CKD and stimulates bone cell osteogenic differentiation, we used in vivo and in vitro rodent models to define the role of hypoxic signaling during elevated inorganic phosphate-induced VSMC calcification. Cell mineralization studies showed that elevated inorganic phosphate rapidly induced VSMC calcification. Hypoxia strongly enhanced elevated inorganic phosphate-induced VSMC calcification and osteogenic transdifferentiation, as seen by osteogenic marker expression. Hypoxia-inducible factor-1 (HIF-1), the key hypoxic transcription factor, was essential for enhanced VSMC calcification. Targeting HIF-1 expression in murine VSMC blocked calcification in hypoxia with elevated inorganic phosphate while HIF-1 activators, including clinically used FG-4592/Roxadustat, recreated a procalcifying environment. Elevated inorganic phosphate rapidly activated HIF-1, even in normal oxygenation; an effect mediated by HIF-1α subunit stabilization. Thus, hypoxia synergizes with elevated inorganic phosphate to enhance VSMC osteogenic transdifferentiation. Our work identifies HIF-1 as an early CKD-related pathological event, prospective marker, and potential target against vascular calcification in CKD-relevant conditions.