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. Author manuscript; available in PMC: 2021 Jan 16.
Published in final edited form as: FASEB J. 2020 Jul 28;34(9):12502–12520. doi: 10.1096/fj.202000612R

Fig. 1. Phenotypic Characterization of RPE-Specific Tsc1 KO Mice.

Fig. 1.

(A) Western blot analysis of RPE tissue from 4 ~ 6 weeks old RPE_Tsc1 KO and Tsc1 flox littermate. Actin was used as the loading control. The data are representative of three different experiments. (B) Immunostaining of Cre recombinase on cryosections prepared from the posterior eye of RPE Tsc1_KO at the age of 4 ~ 6 weeks. Red: Cre; Blue: Nuclei. The immunostaining was repeated with cryosections prepared from three different animals. (C) Immunostaining of flat-mounted RPE/choroid tissue with an antibody specific for Cre. Green: Cre; Red: F-actin; Blue: Nuclei. The data are representative of three different experiments. Animals at the age of 6 ~ 8 weeks were used. (D) OCT scan of mice at 8 ~ 11 months. RPE_Tsc1 KO displayed hyper-reflective spots (arrowheads) resembling subretinal drusenoid deposits (SDD). (E-J) H&E-stained histology sections of the posterior eyes from Tsc1 flox littermates and Tsc1 RPE-specific knockouts at 8 ~ 11 months. The RPE_Tsc1 KO displayed outer retina pathology, including subretinal cell infiltration (blue arrows in Fig. 1F and I), OS disorganization (white arrows in Fig. 1G), RPE cyst and lysis (asterisk in Fig. 1F), hypopigmentation (Fig. 1J black arrows), hyperpigmentation (Fig. 1J white arrows), hypertrophy (blue arrowheads in Fig. 1G) and atrophy of the RPE (black arrowheads in Fig. 1G), anuclear material accumulation on the apical face of the RPE (Fig. 1H), and choroid thinning (Fig. 1E, G and H). (K) Quantification of the numbers of rows of nuclei in the outer nuclei layer (n=5 ~ 6 per genotype). Values are presented as mean ± SEM, * indicates a statistically significant difference between different genotypes (*p < 0.05, ** p < 0.01, two-way ANOVA followed by Sidak’s multiple comparison tests). Scale bar: 50 μm (B, H-J); 20 μm (C); 100 μm (E-G). ELM: external limiting membrane; IS/OS: inner segments/outer segments; RPE: retinal pigment epithelium; GCL: ganglion cell layer; IPL: inner plexiform layer; INL: inner nuclei layer; OPL: outer plexiform layer; ONL: outer nuclei layer; CH: choroid.