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J. Cao, Y. Liu, N. Gale, J. S. Rudge, G. D. Yancopoulos, S. J. Wiegand; Genetic Deletion or Pharmacological Inhibition of Angiopoietin-2 Reduces Injury-Induced Corneal Neovascularization, Lymphangiogenesis and Inflammation. Invest. Ophthalmol. Vis. Sci. 2007;48(13):3198.
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To evaluate the role of endogenous Angiopoietin-2 (Ang2) in the development of corneal neovascularization and inflammation.
Adult mice (7-14 weeks of age) homozygous for Ang2 deletion / LacZ substitution (Ang2-/-), their wild type (WT) littermates, and adult male C57Bl/6 mice were used in these studies. Corneal neovascularization (NV) was induced by intrastromal placement of 3 nylon sutures in the right eye (day 0). In C57Bl/6 mice, a neutralizing Ang2 antibody (25mg/kg) or a control protein (human Fc, 10mg/kg) was injected subcutaneously on days 0, 3 and 6 following corneal injury. On day 9 after corneal injury, the vasculature was labeled by intravenous injection of fluorescein conjugated lectin (lycopersicon esculentum), and corneal NV was evaluated in flat-mounts. Lymphatic vessels were co-localized in the same specimens by LYVE-1 immunostaining. Macrophage infiltration was evaluated in cross-sections stained with rat anti-mouse F4/80 monoclonal antibody. The Scion Image program was used for analysis of the area and length of corneal neovessels. Corneal thickness also was measured in cross-sections, as an index of edema.
The vasculature at the corneal limbus exhibited abnormalities in adult Ang2-/- mice, most notably a reduction in the caliber of the major circumferential veins and arteries. Following suture injury, corneal NV was significantly reduced in the Ang2-/- mice compared to their WT littermates (p <0.001) and also altered qualitatively, with the neovessels being much finer in the Ang2-/- mice. Genetic deletion of Ang2 also dramatically reduced the infiltration of macrophages into the damaged cornea. Systemic treatment with Ang2 antibody significantly inhibited corneal NV (p < 0.001) in C57Bl.6 mice. Ang2 antibody treatment also markedly decreased the growth of lymphatic vessels into the injured cornea and reduced corneal edema.
These findings demonstrate that endogenous Ang2 plays an important role in the development of neovascularization, lymphangiogenesis and inflammation following corneal injury.
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