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. 2019 Sep 9;9:12903. doi: 10.1038/s41598-019-49472-8

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© The Author(s) 2019

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 license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license 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 license, visit http://creativecommons.org/licenses/by/4.0/.

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Azapeptide MPE-001 regulates inflammatory profile of subretinal MPs and reduces photoreceptor degeneration. (A–H) CD36^+/+ mice were illuminated for 5 days with blue light. Subcutaneous injections of 289 nmol/kg MPE-001 were administered after 1 day illumination and pursued daily for 7 consecutive days. (A) Upper panel: area of retinal cryosections were microdissected between ONL and RPE and visualized with green circles. Lower panel: bar graphs of IBA-1, iNOS, IL-6, IL-10 and IL-12 mRNA expression levels in microdissected retinal cryosections. Cytokine analysis were normalized to 18 s rRNA. (B) Subretinal MPs stained for F4/80 (red) and iNOS (green) on RPE flat mounts as assessed by confocal microscopy. (C) Subretinal MPs stained with IBA-1 (red) and IL-12 (green) on RPE flat mounts as assessed by confocal microscopy. (D) Subretinal MPs stained with anti-IBA-1 (green) and anti-CD206 (red) antibodies on RPE flat mounts as assessed by confocal microscopy. Scale bar: 15 μm. (E) Percentage of subretinal MPs (IBA-1⁺ or F4/80⁺) expressing INOS, IL-12 or CD206. (F) Confocal microscopy of neuroretinal flat mounts (photoreceptors side) and (G) retina cryosections from illuminated CD36^+/+ mice treated or not with MPE-001 stained with fluorescein PNA (green) and anti-S-opsin (red) antibody; nuclei were counterstained with dapi (blue). Magnifications of white square show length of cone outer segment with S-opsin distribution. Scale bar: 10 μm. (H) Retinal cryosections stained with TUNEL (green). Nuclear layers were stained with DAPI (blue). Scale bar: 25 μm. (I) Percentage of TUNEL⁺ cells in ONL cryosections of the retina. In (A and E) unpaired t-test was performed. *P < 0.05, **P < 0.01 and ***P < 0.001 vs illuminated group (n = 3-4 mice/group). In I, one-way ANOVA test with Newman-Keuls for multiple comparison was performed. ^#P < 0.05 and ^###P < 0.001 vs no illumination group. Ψ P < 0.05 vs illuminated group (n = 3-4 mice/group). Data are shown as mean ± S.E.M. RGC: Retinal Ganglion Cell. INL: Inner Nuclear Layer. ONL: Outer Nuclear Layer. RPE: Retinal Pigment Epithelium. CHR: Choroid.