Defining an Upstream VEGF (Vascular Endothelial Growth Factor) Priming Signature for Downstream Factor-Induced Endothelial Cell-Pericyte Tube Network Coassembly.
OBJECTIVE
In this work, we have sought to define growth factor requirements and the signaling basis for different stages of human vascular morphogenesis and maturation. Approach and Results: Using a serum-free model of endothelial cell (EC) tube morphogenesis in 3-dimensional collagen matrices that depends on a 5 growth factor combination, SCF (stem cell factor), IL (interleukin)-3, SDF (stromal-derived factor)-1α, FGF (fibroblast growth factor)-2, and insulin (factors), we demonstrate that VEGF (vascular endothelial growth factor) pretreatment of ECs for 8 hours (ie, VEGF priming) leads to marked increases in the EC response to the factors which includes; EC tip cells, EC tubulogenesis, pericyte recruitment and proliferation, and basement membrane deposition. VEGF priming requires VEGFR2, and the effect of VEGFR2 is selective to the priming response and does not affect factor-dependent tubulogenesis in the absence of priming. Key molecule and signaling requirements for VEGF priming include RhoA, Rock1 (Rho-kinase), PKCα (protein kinase C α), and PKD2 (protein kinase D2). siRNA suppression or pharmacological blockade of these molecules and signaling pathways interfere with the ability of VEGF to act as an upstream primer of downstream factor-dependent EC tube formation as well as pericyte recruitment. VEGF priming was also associated with the formation of actin stress fibers, activation of focal adhesion components, upregulation of the EC factor receptors, c-Kit, IL-3Rα, and CXCR4 (C-X-C chemokine receptor type 4), and upregulation of EC-derived PDGF (platelet-derived growth factor)-BB, PDGF-DD, and HB-EGF (heparin-binding epidermal growth factor) which collectively affect pericyte recruitment and proliferation.


CONCLUSIONS
Overall, this study defines a signaling signature for a separable upstream VEGF priming step, which can activate ECs to respond to downstream factors that are necessary to form branching tube networks with associated mural cells.
['Bowers|Stephanie L K|SLK|', 'Kemp|Scott S|SS|', 'Aguera|Kalia N|KN|', 'Koller|Gretchen M|GM|', 'Forgy|Joshua C|JC|', 'Davis|George E|GE|']
D043925:Angiogenesis Inducing Agents D002450:Cell Communication D002478:Cells, Cultured D018920:Coculture Techniques D005786:Gene Expression Regulation D061307:Human Umbilical Vein Endothelial Cells D006801:Humans D018919:Neovascularization, Physiologic D020286:Pericytes D010766:Phosphorylation D015398:Signal Transduction D042461:Vascular Endothelial Growth Factor A D040301:Vascular Endothelial Growth Factor Receptor-2
endothelial cells stem cell factor insulin fibroblast growth factor interleukin-3