In vivo lymphatic vessel formation in mouse cornea induced by COMP-Ang-1. (A) ALY7 immunostaining of lymphatic vessels of the cornea. COMP-Ang-1-induced migrated endothelial cells on the cornea show positive LYVE-1 immunoreactivity (arrows), indicating that lymphatic vessels were induced by COMP-Ang-1. (B) No obvious LYVE-1⁺ lymphatic vessel is seen on the cornea with a pellet containing PBS alone. (C) Double-staining of newly formed vessels with ALY (red) and anti-CD31 (green) antibodies. Both of the lymphatic (merged) and blood vessel (green) endothelial cells migrated into the cornea induced by the pellet containing COMP-Ang-1. (D) Quantitative estimation of corneal lymphangiogenesis by immunostaining with ALY7. The lymphangiogenic grading score was determined by a previously described method.¹⁰  Recombinant soluble Tie2-Fc protein suppressed the COMP-Ang-1-induced lymphatic vessel formation on mouse cornea, whereas soluble CD4-Fc protein could not inhibit corneal lymphangiogenesis. The graphs represent the mean ± SEM (n = 6); scale bars indicate 50 μm.