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A. De Erkenez, Y.S. Ng; An ex vivo Model for Studying Angiogenic Vessel Maturation and Stabilization . Invest. Ophthalmol. Vis. Sci. 2005;46(13):3264.
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Purpose:Pericyte recruitment and the subsequent association of pericytes with the EC in developing vessels is critical for maintaining vascular integrity and stability. The platelet–derived growth factor B (PDGF–B)/PDGF receptor system plays an important role in this process. The molecular mechanism of PDGF–B/receptor mediated pericyte recruitment and EC–association remains unclear, due to the lack of an appropriate in–vitro model. The aim of this study is to establish an ex–vivo organ culture model to study pericyte recruitment and pericyte–EC interactions during vessel formation, with a particular focus on the roles of the VEGF/VEGF receptor and the PDGF–B/receptor systems in this process. Methods:Thoracic aortic rings from 6 week old male Sprague–Dawley rats were cultured on a type I and type IV collagen matrix in serum–free media with or without growth factors/reagents that interrupt the VEGF/receptor and PDGF–B/receptor systems. To examine pericyte recruitment and EC–pericyte interaction, the aortic rings were fixed after 7 days of culture and stained for EC (PECAM–1, VE–Cadherin), pericyte (NG2), and smooth muscle cells (smooth muscle actin) markers. Results:Immunohistochemical results showed that angiogenic vessel sprouts contain EC and tightly associated pericytes, similar to the organization in vivo. Reagents that disrupt the PDGFB/receptor system (excess PDGF–BB, PDGFRß/Fc, anti–PDGF–B aptamer) disrupted pericyte recruitment and the EC–pericyte interaction leaving shortened angiogenic vessels with loosely associated or no pericyte coverage. Pegaptanib sodium, the anti–VEGF aptamer, reduced the number and length of angiogenic vessel sprouts, suggesting that vessel sprouting is VEGF–dependent in this model. The combination of the anti–VEGF aptamer and reagents that disrupt the PDGF–B/receptor system are more effective at inhibiting vessel sprouting than either individually. Conclusions: These results suggest that the ex–vivo aortic culture model recapitulates the complex in–vivo process of pericyte recruitment and EC–pericyte interaction during angiogenesis. Disruption of the PDGF–B/receptor system results in vessels that lack the appropriate EC–pericyte interactions. Disruption of the VEGF/VEGF receptor system results in a reduction in vessel sprouting. The combination of anti–VEGF and anti–PDGF–B treatments is the most effective at inhibiting vessel sprouting, suggesting that the PDGF–B/receptor system is involved in vascular maturation and stabilization. This model is useful in studying the interaction between VEGF and PDGF–B in angiogenesis.
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