VEGF transiently disrupts gap junctional communication in endothelial cells

Abstract

Vascular endothelial growth factor, VEGF, stimulates angiogenesis by directly acting on endothelial cells. The effects of VEGF are mediated by two tyrosine kinase receptors, VEGFR-1 (Flt-1) and VEGFR-2 (Flk-1/KDR) that are highly related to receptors of the platelet derived growth factor (PDGF) receptor family. We are interested in early signalling events downstream from VEGF receptors that affect blood vessel homeostasis. Endothelial cells form multiple types of cell-cell junctions that are required for cellular organization into complex networks. These junctions also regulate communication among adjacent cells. Stimulation by various growth factors such as epidermal growth factor (EGF) or PDGF has been shown to disrupt cell-cell junctions, consequently affecting cell-to-cell communication. We investigated gap junctional communication (GJC) by monitoring the transfer of a low molecular mass fluorescent tracer molecule between adjacent cells using immunofluorescence microscopy. VEGF maximally blocked GJC 15 minutes after growth factor administration. The cells resumed communication via gap junctions within 1–2 hours after treatment. This early effect of VEGF on communication correlated with changes in the phosphorylation state of one of the proteins involved in gap junction formation, connexin 43 (Cx43). The signalling mechanisms involved in this phenomenon depend on activation of VEGFR-2, impinge on a tyrosine kinase of the Src family and activate the Erk family of MAP kinases. The function of VEGF-mediated disruption of GJC might be to restrict an increase in endothelium permeability to the environment affected by local injury to blood vessels.

Modulation of gap junction channels and hemichannels by growth factors

“…Similar to other growth factors, VEGF induced a fast (o15 min) and reversible reduction in dye coupling evaluated with Lucifer yellow in human umbilical vein endothelial cells (HUVEC) and Ea.hy926 cells. 152 The effect was mediated by VEGFR-2, required the activation of Src and MAPK, and was associated with Cx43 phosphorylation, apparently at serine/threonine residues, without changes in total Cx43 levels. VEGF also abolished gap junctional communication within 30 min in primary cultures of rat ventricular capillary endothelial cells.…”

The Role of Connexin in Ophthalmic Neovascularization and the Interaction between Connexin and Proangiogenic Factors

“…However, after approximately one hour, this function can be reversed. Researchers hypothesize that VEGF-induced GJC blockade may act by increasing endothelial permeability in injured blood vessels [ 55 ]. VEGF inhibits the Ca 2+ burst in pregnant sheep by phosphorylating Cx43 at Tyr-265 and Ser-279/282 via the VEGFR2 Src and ERK pathways.…”

“…Some related studies have found that gap junction communication in vascular endothelial cells is disrupted during the early stages of VEGF stimulation and then resumes 1-2 hours after treatment. This disruption is attributed to VEGF's effect on the phosphorylation state of Cx43, which ultimately affects the permeability of vascular endothelial cells [ 55 ] ( Figure 1 and Table 2 ).…”

“…Connexin type Comment VEGF [52] Cx43 GJIC Activation of VEGFR2 by VEGF-E alone inhibited GJIC VEGF [48] Cx43 GJIC EGF promoted GJIC in glioblastoma multiforme cells VEGF [48] Cx43 GJ VEGF increased the expression of gap junction proteins in U-251 glioblastoma multiforme cells VEGF [39] Cx43 VEGF enhanced the expression of Cx43 in endothelial progenitor cells VEGF [51] Cx43 GJ Exposure of primary porcine pulmonary artery endothelial cells to VEGF markedly suppressed Cx43 GJs within 1 hour VEGF [52] Cx43 GJ VEGF induced Cx43 phosphorylation leading to a decline in pregnancy-adapted Ca 2+ burst function VEGF [49] Cx43 VEGF120 increased expression of Cx43 in heart VEGF [50] Cx43 GJIC VEGF improved intercellular communication at the Cx43 gap junction VEGF [54] Cx43 GJIC In coronary capillary endothelium, VEGF disrupted intercellular communication and promoted Cx43 endocytosis and phosphorylation at the Cx43 gap junction VEGF [53] Cx43 GJIC VEGF induced phosphorylation of connexin 43 to regulate gap junction intercellular communication in HUVECs VEGF [55] Cx43 GJIC VEGF reversibly inhibited GJIC function in EA.hy926 cells and HUVECs bFGF and LPS [57] Cx43 GJ and HC Inhibited Cx43 gap junctions but stimulated hemichannels in C6 glioma cells bFGF and LPS [57] Cx43 GJ and HC Inhibited Cx43 hemichannels function in HeLa cells.…”

Normalizing Tumor Vasculature to Reduce Hypoxia, Enhance Perfusion, and Optimize Therapy Uptake

“…Given that these plasma membrane microdomains are vital for the exchange of solutes and water at the interface of tissue and vasculature, tumor endothelial cells are often more porous compared with normal counterparts [43]. Abnormal VEGF signaling in tumor endothelial cells also leads to downregulation of connexin expression, causing gap junction dysfunction, increasing vascular fenestrations, and increasing vascular permeability [44][45][46]. In fact, VEGF was initially identified based on its ability to increase vascular permeability and extravasation of plasma proteins, such as fibrinogen [47].…”