Skip to main content
. 2020 Oct 19;29(20):3373–3387. doi: 10.1093/hmg/ddaa228

Figure 1.

Figure 1

Nf2 disruption in ocular tissues causes coloboma in the embryonic mouse eye. (A) Control embryonic head at E13.5. (B) Nf2CKO embryos exhibit cleft lip and palate (arrowhead). (C) Higher magnification of E13.5 control eye. (D) In Nf2CKO embryos at E13.5, the ventral eye shows an OF closure defect with a narrow gap (arrowhead) and the pigment appears slightly darker. (E) Immunolabeling for the basement marker laminin in the E13.5 ventral optic cup of controls confirms absence of laminin and successful fusion of the OF region (arrowhead). Sagittal view with nasal orientation to the left. (F) Laminin is persistent in the ventral optic cup in E13.5 Nf2CKO embryos indicating failed OF fusion (arrowhead). (G) Brightfield image of the ventral optic cup in control showing pigmentation of the RPE in the fused OF (arrowhead). (H) In the ventral optic cup of E13.5 Nf2CKO embryos, pigmentation extends into the unfused OF (arrowhead) and the RPE layer appears slightly increased in height. Insets in G, H show magnified regions outlined in boxed area, revealing increased thickness in mutant RPE (H). (I) During OF closure at E11.5, YAP1 protein expression can be localized both to the cytoplasm and nucleus in the RPE (arrowheads point to RPE nuclei). (J) In Nf2CKO eyes, nuclear YAP1 expression can be increased in retina and RPE (arrowheads display nuclear Yap localization in RPE). RPE in I and J is indicated by dotted lines and sections were counterstained with 2-(4-amidinophenyl)-1H-indole-6-carboxamide (Dapi, blue). Insets in I and J show enlarged greyscale images of individual cells in the RPE (boxed area). I′ and J′ display the Dapi channel, I″ and J″ display the YAP1 channel. Individual nuclei in the insets are outlined by dotted lines. (K) YAP1 average integrated density per RPE nuclei shows a significant increase in Nf2CKO compared with control nuclei. (L, M) Cyclin D1 (CCND1)-labeled cells are significantly increased in Nf2CKO ventral RPE (M; arrowheads; N). Asterisks in I, J, L, M indicate OF. Data are shown as means ± SD, and P-values are indicated above the horizontal lines in each graph (significant < 0.05). Statistical analysis was performed using unpaired, two-tailed Student T-test (Welch’s correction included for K). Scale bars: 100 μm in E, L; 20 μm in I, J.