Fig. 6

Light overresponse and photoreceptor cell death after light irradiation in RD retinal organoids. (A) Retinal organoids were exposed to white LED light on day 180. (B) Intracellular cGMP concentration was measured by ELISA. Percentage changes in cGMP concentration by light stimulation in control and RD retinal organoids are shown. Data represent mean ± SEM from independent experiments (n = 6). An unpaired, 2-tailed t test was used for statistical comparison (*P < 0.05). (C) Representative images of reactive oxygen species (ROS) stained with CellROX in control and RD retinal organoids. Twenty-four-hour light-stimulated (Bright) or dark-adapted (Dark) organoids on day 180 were evaluated. The lower panels are higher-magnification images of the dotted boxes in the upper panels. Scale bars: 100 μm. (D) Quantification of the data in C. The y axis indicates the ratio of CellROX-positive cells. Data represent mean ± SD from 3 retinal organoids. One-way ANOVA with Dunnett’s post hoc test was used for statistical comparison (*P < 0.05). (E) Representative immunofluorescence images of photoreceptor marker ARR1 and cleaved caspase-3 (Cl. CASP3) in retinal organoids after light exposure or dark adaptation. White arrowheads indicate cleaved caspase-3–positive cells. Scale bars: 50 μm. (F) Quantification of the data in E. The y axis indicates the number of cleaved caspase-3–positive cells per field. Data represent mean ± SEM from 3 retinal organoids. One-way ANOVA with Dunnett’s post hoc test was used for statistical comparison (*P < 0.05). (G) Representative immunofluorescence images of ARR1 and cleaved caspase-3 in control and EYS-KO retinal organoids after light exposure or dark adaptation. White arrowheads indicate cleaved caspase-3–positive cells. Scale bars: 50 μm. (H) Quantification of the data in G. The y axis indicates the number of cleaved caspase-3–positive cells per field. Data represent mean ± SEM from 3 retinal organoids. One-way ANOVA with Dunnett’s post hoc test was used for statistical comparison (*P < 0.05).