Extracellular matrix deposition by fibroblasts is necessary to promote capillary-like tube formation in vitro.
The contribution of the cellular and fibrillar microenvironment to angiogenesis still remains unclear. Our purpose was to evaluate the effect of the extracellular matrix deposited by fibroblasts on the capacity of human endothelial cells to form capillaries in vitro. We have drastically decreased the amount of extracellular matrix surrounding fibroblasts in our model of endothelialized-reconstructed connective tissue (ERCT) by culturing it without ascorbate. Under these conditions, the number of capillary-like tubes (CLT) formed by endothelial cells was reduced by up to 10-fold after 31 days of culture compared to controls. This decrease was due neither to a variation of MMP-2 and MMP-9 secretion, nor to a reduction in the number of fibroblasts and/or endothelial cells, or a diminution of fibroblast growth factor 2 (FGF2) synthesis. The secretion of vascular endothelial growth factor (VEGF) by fibroblasts accounted for 25-70% of the capillary-like tube formation when tissues were cultured in the presence or absence of ascorbate, as demonstrated by VEGF-blocking studies. The culture of endothelial cells on a similar extracellular matrix but in the absence of living fibroblasts did not promote the formation of CLT, even when tissues were fed with fibroblast-conditioned medium. Thus, the deposition of a rich extracellular matrix by living fibroblasts appeared necessary, but not sufficient to promote capillary-like formation. Fibroblasts seem to induce endothelial cells to spontaneously form CLT by secreting and organizing an abundant extracellular matrix, which creates a microenvironment around cells that could in turn trap growth factors produced by fibroblasts and promote three-dimensional cell organization.
Berthod
François
F
Laboratoire d'Organogénèse Expérimentale (LOEX), Centre hospitalier affilié universitaire de Québec, Hôpital du Saint-Sacrement, Québec, Canada. Francois.berthod@chg.ulaval.ca
Germain
Lucie
L
Tremblay
Nathalie
N
Auger
François A
FA
D001205:Ascorbic Acid / Q000494:pharmacology; D002196:Capillaries / Q000166:cytology / Q000648:ultrastructure; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D018920:Coculture Techniques; D017077:Culture Media, Conditioned / Q000494:pharmacology; D042783:Endothelial Cells / Q000187:drug effects / Q000502:physiology* / Q000648:ultrastructure; D005109:Extracellular Matrix / Q000378:metabolism*; D016222:Fibroblast Growth Factor 2 / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D007797:Laminin / Q000032:analysis; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism; D046529:Microscopy, Electron, Transmission; D018919:Neovascularization, Physiologic / Q000187:drug effects / Q000502:physiology; D019408:Platelet Endothelial Cell Adhesion Molecule-1 / Q000032:analysis; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism; D014841:von Willebrand Factor / Q000032:analysis
D017077:Culture Media, Conditioned; D007797:Laminin; D019408:Platelet Endothelial Cell Adhesion Molecule-1; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A; D014841:von Willebrand Factor; D016222:Fibroblast Growth Factor 2; D020778:Matrix Metalloproteinase 2; D020780:Matrix Metalloproteinase 9; D001205:Ascorbic Acid