Abstract
Angiogenesis is crucial for tissue development and homeostasis; however, excessive angiogenesis can lead to diseases, including arthritis and cancer metastasis. Some antiangiogenic drugs are available, but side effects remain problematic. Thus, alternative angiogenesis inhibition strategies are needed. Fibulin-7 (Fbln7) is a newly discovered member of the fibulin protein family, a group of cell-secreted glycoproteins, that functions as a cell adhesion molecule and interacts with other extracellular matrix (ECM) proteins as well as cell receptors. We previously showed that a recombinant C-terminal Fbln7 fragment (Fbln7-C) inhibits tube formation by human umbilical vein endothelial cells (HUVECs) in vitro. In the present study, we examined the in vivo antiangiogenic activity of recombinant full-length Fbln7 (Fbln7-FL) and Fbln7-C proteins using a rat corneal angiogenesis model. We found that both Fbln7-FL and Fbln7-C inhibited neovascularization. Fbln7-C bound to vascular endothelial growth factor receptor 2 (VEGFR2), inhibiting VEGFR2 and ERK phosphorylation and resulting in reduced HUVEC motility. HUVEC attachment to Fbln7-C occurred through an interaction with integrin α5β1 and regulated changes in cellular morphology. These results suggest that Fbln7-C action may target neovascularization by altering cell/ECM associations. Therefore, Fbln7-C could have potential as a therapeutic agent for diseases associated with angiogenesis.
Highlights
Integrins are membrane-associated molecules that regulate endothelial cell adhesion to extracellular matrix (ECM) at focal adhesion sites during angiogenesis[6,7]
We examined the antiangiogenic activity of Fbln7-C in vivo using a rat corneal angiogenesis model
This model is characterized by the induction of neovascularization by the pro-angiogenic, pro-inflammatory lipid 7KCh13. 7KCh was previously reported to be a very potent inducer of vascular endothelial growth factor (VEGF) production in vivo and in vitro14. 7KCh induced VEGF expression and secretion in endothelial cells and monocytes recruited to implants, suggesting that VEGF is a crucial regulator of neovascularization in the rat eye corneal model[13]
Summary
Integrins are membrane-associated molecules that regulate endothelial cell adhesion to ECM at focal adhesion sites during angiogenesis[6,7]. They play an important role in the synergy among growth factor receptors during angiogenesis. HUVEC morphology was altered after Fbln7-C treatment and was associated with RhoA inhibition[12] It is not clear how HUVEC morphology is regulated under angiogenic conditions, nor whether Fbln7-C has antiangiogenic properties in vivo. We found that Fbln7-C inhibited neovascularization in vivo, and that the inhibition was mediated by Fbln7-C binding to VEGFR2, which subsequently decreased endothelial cell motility by inhibiting the downstream VEGF signaling pathway with induction of focal adhesion site maturation. Our results suggest that Fbln7-C may have potential as a therapeutic agent for diseases associated with angiogenesis
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