title: A novel ex vivo murine retina angiogenesis (EMRA) assay.
abstract: Pathological retinal angiogenesis results from the imbalance of pro-angiogenic and anti-angiogenic factors. In particular, vascular endothelial growth factor (VEGF) plays a pivotal role in retinal neovascularization and various therapeutic VEGF blockers have evolved over time. Nevertheless, new retinal angiogenesis models are crucial for investigating anti-angiogenic therapies and bringing them to patients. Here, we developed a novel ex vivo murine retina angiogenesis (EMRA) assay in which endothelial sprouts originate from mature and quiescent retinal vessels. In this model, retina fragments from adult mice are embedded in a three-dimensional fibrin gel in the presence of human recombinant VEGF. Starting from the 3rd-4th day of incubation, endothelial cell sprouts invading the fibrin gel can be observed under an inverted microscope and measured at different time points thereafter. The effect of VEGF is dose-dependent, maximal stimulation being observed at day 7 for retina fragments stimulated with 25-75 ng/ml of the growth factor. To assess whether the EMRA assay is suitable for testing the activity of anti-angiogenic compounds, retina fragments were incubated with VEGF in the presence of the neutralizing anti-VEGF antibodies bevacizumab and ranibizumab. The results demonstrate that both antibodies inhibit VEGF activity in a dose-dependent manner. In conclusion, the EMRA assay represents a new ex vivo model of retinal neovascularization suitable for the rapid screening of novel anti-angiogenic therapeutics.
authors: Department of Molecular and Translational Medicine, University of Brescia, Viale Europa 11, 25123 Brescia, Italy; Department of Ophthalmology, University of Brescia, Brescia, Italy.
authors: Sara
authors: S
authors: Rezzola
authors: Mirella
authors: M
authors: Belleri
authors: Domenico
authors: D
authors: Ribatti
authors: Ciro
authors: C
authors: Costagliola
authors: Marco
authors: M
authors: Presta
authors: Francesco
authors: F
authors: Semeraro
mesh_terms: D020533:Angiogenesis Inhibitors / Q000494:pharmacology; D000818:Animals; D061067:Antibodies, Monoclonal, Humanized / Q000494:pharmacology; D057134:Antibodies, Neutralizing / Q000494:pharmacology; D000068258:Bevacizumab; D001681:Biological Assay / Q000379:methods*; D004195:Disease Models, Animal; D004305:Dose-Response Relationship, Drug; D004730:Endothelium, Vascular / Q000187:drug effects*; D005337:Fibrin; D051379:Mice; D008810:Mice, Inbred C57BL; D009924:Organ Culture Techniques; D000069579:Ranibizumab; D011994:Recombinant Proteins; D012160:Retina / Q000187:drug effects; D015861:Retinal Neovascularization / Q000188:drug therapy / Q000209:etiology*; D012171:Retinal Vessels / Q000187:drug effects / Q000473:pathology*; D042461:Vascular Endothelial Growth Factor A / Q000037:antagonists & inhibitors / Q000633:toxicity
chemical_list: D020533:Angiogenesis Inhibitors; D061067:Antibodies, Monoclonal, Humanized; D057134:Antibodies, Neutralizing; D011994:Recombinant Proteins; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A; D000068258:Bevacizumab; D005337:Fibrin; D000069579:Ranibizumab