Characterization of Local Retinal Damage during Postnatal Development of Pax6 Small-eye Mice

Abstract

Purpose : Aniridia is a panocular developmental disorder, which is often accompanied by a broad range of phenotypes. Approximately 1 in every 100,000 will develop aniridia, with most cases being inherited from parents. Roughly 90% of aniridia cases arise from mutations in the PAX6 gene, a transcription factor necessary for brain and eye development. Our previous studies found that Pax6 haploinsufficiency results in localized regions of retinal damage known as “hotspots”. Much remains unknown about how layers and retinal subtypes that contribute to this damage and how it progresses with age.

Methods : We utilized a small-eye mouse model with C57Bl/6 and 129S1/Svlmj background for in vivo and in vitro assessments of retinal health. We measured the following metrics at four ages (P20, P30, P60, and P90): (1) optomotor assessments with a qOMR system; and (2) intraocular pressure (IOP) using rebound tonometry. At P90 and P30, retinas were prepared for immunolabeling by markers such as Rbpms (retinal ganglion cells), ChAT (cholinergic amacrine cells), AP2 (amacrine cells), Iba1 (microglia), and GFAP (Müller glia).

Results : Mean IOP of C57Bl/6 Pax6^Sey IOP reached 19.4±0.58 mmHg by P30, while the WT IOP was 14.8±0.26 mmHg (p < 0.0001, P30, ANOVA with post-hoc Tukey test). The mean IOP of 129s Pax6^Sey did not show significantly different IOPs until P60 (17.7±0.86 mmHg) compared to littermate WT (14.7±0.29 mmHg, p=0.002). Optomotor assessments showed a significant decline in both groups. At P20, C57Bl/6 Pax6^Sey exhibited a low acuity (0.15±0.02 cyc/deg; WT:0.35±0.006 cyc/deg, p<0.0001), as did129 Pax6^Sey at this age (0.32±0.01 cyc/deg; WT:0.43±0.007 cyc/deg, p=0.034). IHC studies showed a trend toward a reduction of Rbmps+ RGCs and AP2+ amacrine cells in the Sey mice of both groups. Whole mount studies showed a significant increase in Iba1+ of the C57Bl/6 Pax6^Sey (90.4±8.2 per 90000µm²) as compared to WT (54.1±2.3 per 90000µm², p=0.0034). 129S1 WT and Pax6^Sey showed no significant difference (p=0.87).

Conclusions : Our findings indicate that retinal damage in aniridia varied among individuals with different genetic backgrounds. Increased IOP may contribute to the development of local retinal damage with age.. Our studies give us a fuller understanding of retina’s developmental defects in aniridia and build the foundation for developing new treatments for retinal damage and vision loss for aniridia patients.

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.