Single-cell-resolution analysis of RPE flatmounts revealed only minor changes to cellular morphology.

(a) Ift20+/+;Tyrp2-Cre and Ift20null;Tyrp2-Cre RPE flatmounts stained with Phalloidin (red), cell borders were automatically analyzed, segmented, and visualized by color-coded images. (b) Representative images showing Phalloidin staining (red) at P11, P29, and 3M of Ift20+/+;Tyrp2-Cre and Ift20null;Tyrp2-Cre RPE flatmounts. Scale bar: 20 μm. (c) Cell area data binned at intervals of 50 μm2. Binned profiles look remarkably similar between control and mutant across all 3 ages examined. (d–g) Quantification of the cell size, AR, number of neighbors, and hexagonality of Ift20+/+;Tyrp2-Cre and Ift20null;Tyrp2-Cre RPE flatmounts. Analysis of cell size revealed a modest increase in cell size in the mutant starting from P29 onwards, while AR was decreasing. No changes were detected in number of neighbors and hexagonality. Median: cell area P11 Ift20+/+;Tyrp2-Cre 208.67 μm2, Ift20null;Tyrp2-Cre 218.08 μm2. P29: Ift20+/+;Tyrp2-Cre 249.7 μm2, Ift20null;Tyrp2-Cre 271.84 μm2. 3m: Ift20+/+;Tyrp2-Cre 287.82 μm2, Ift20null;Tyrp2-Cre 303.72 μm2. Median: aspect ratio P11 Ift20+/+;Tyrp2-Cre 1.375, Ift20null;Tyrp2-Cre 1.365. P29: Ift20+/+;Tyrp2-Cre 1.331, Ift20null;Tyrp2-Cre 1.335. 3m: Ift20+/+;Tyrp2-Cre 1.375, Ift20null;Tyrp2-Cre 1.363. Median: neighbors P11 Ift20+/+;Tyrp2-Cre 5.61, Ift20null;Tyrp2-Cre 5.68. P29: Ift20+/+;Tyrp2-Cre 5.51, Ift20null;Tyrp2-Cre 5.59. 3m: Ift20+/+;Tyrp2-Cre 5.34, Ift20null;Tyrp2-Cre 5.43. Median: hexagonality P11 Ift20+/+;Tyrp2-Cre 8.70, Ift20null;Tyrp2-Cre 8.73. P29: Ift20+/+;Tyrp2-Cre 8.88, Ift20null;Tyrp2-Cre 8.86. 3m: Ift20+/+;Tyrp2-Cre 8.60, Ift20null;Tyrp2-Cre 8.64 μm2. n numbers P11: Ift20+/+;Tyrp2-Cre (7 flat mounts, 163,530 cells); Ift20null;Tyrp2-Cre (14 flat mounts 295,059 cells), P29: Ift20+/+;Tyrp2-Cre (11 flat mounts, 274,428 cells); Ift20null;Tyrp2-Cre (9 flat mounts, 227,064 cells), 3m: Ift20+/+;Tyrp2-Cre (13 flat mounts, 510,473 cells); Ift20null;Tyrp2-Cre (18 flat mounts 675,748 cells). Significance levels: >0.05 not significant (ns), S6 Table. AR, aspect ratio; RPE, retinal pigment epithelium.