Fig. 6

RPE progressive disorganization after irradiation. (a) RPE/Choroid complex flat mounts stained with Phalloidin (in red) to visualize actin-F cytoskeleton. After a week, almost no changes were visible on the RPE integrity. After a month, large rounded cells (white arrows) and holes (blue arrows) within cells were observed. (b) Quantification of RPE cell surface. One-way ANOVA statistical test, followed by a Dunnett’s multiple comparison test. (c) Distribution of the cell area of RPE cells. The statistical analysis was performed using a Chi-square test comparing the distribution of the control (expected) with the distribution of the 1 month or 6 months (observed). (d) Evaluation of the RPE morphology according to their circularity. Hexagonal cells like RPE cells have a circularity of 0.8. The more cells become round shaped, the more their circularity tends towards 1. One-way ANOVA statistical test, followed by a Dunnett’s multiple comparison test. For all statistics, n = 4 eyes and between 1365 and 2360 cells per eye have been measured on 18 images representative of the whole RPE. **p < 0.005; ***p < 0.0005; ****p < 0.0001. For the control, 1-month and 6-months group, the mean circularity was 0,8012; 0,7985 and 0,8079 respectively with a standard deviation of 0,03860; 0,04185 and 0,04328. (e) RPE flat mounts stained with phalloidin (in red) and ZO-1 (in green) to localize tight-junctions, one week, one month and 6 months after irradiation. Scale bars represent 50 μm. (f) Enlargement of ZO-1 staining on RPE flat mounts, associated with the merge image with phalloidin (in red), ZO-1 (in green) and Dapi (in blue). Abnormalities are enlightened with white arrows. Scale bars represent 20 μm