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. 2020 Nov 13;10:19817. doi: 10.1038/s41598-020-76882-w

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© The Author(s) 2020

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

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Topical administration of ripasudil suppressed neovascularization and lymphangiogenesis in murine corneal transplants. (A) Representative slit-lamp images showing grafted corneas on day 14 post-transplantation (magnification × 25). (B) Neovascularization scores of grafted corneas 14 days after transplantation (two-way ANOVA, n = 5 per group; *p < 0.01). (C) Representative immunohistochemical images of CD31 staining of grafted corneas on day 14 post-transplantation. Upper row: whole grafted cornea. Bottom row: high magnification showing center of grafted cornea. (D) Percentages of the corneal area covered with blood vessels (CD31⁺) in the ripasudil and control groups (one-way ANOVA, n = 5 per group; *p < 0.01). (E) Representative immunohistochemical images of lymphatic vessel endothelial hyaluronan receptor-1 (LYVE-1) staining of grafted corneas on day 14 post-transplantation. Upper row: whole grafted cornea. Bottom row: high magnification showing center of grafted cornea. (F) Percentages of the corneal area covered with lymphatic vessels (LYVE-1⁺) in the ripasudil and control groups (one-way ANOVA, n = 5 per group; *p < 0.05). The area of the whole grafted cornea covered by blood or lymphatic vessels was analyzed using ImageJ version 1.53a (National Institutes of Health, Bethesda, MD, USA; available at https://rsb.info.nih.gov/ij/index.html)^57,58. Scale bar, 1000 μm. ANOVA analysis of variance.