Abstract
Purpose :
All-trans retinoic acid (ATRA) plays an essential role during human eye development, being temporally and spatially adjusted to create gradient concentrations. Perturbations in ATRA signaling can result in severe ocular developmental diseases including microphthalmia, anophthalmia, or coloboma. However, the effects of ATRA in the different ocular tissues during embryonic development of the eye remain unknown. Here we performed a preclinical study to test the effects of high and low ATRA concentrations on early development and maturation of ocular tissues using an in vitro model of human induced pluripotent stem cells (hiPSC) derived multiocular organoids.
Methods :
The differentiation protocol started by multizone ocular progenitor cells generation from hiPSC in 2D, and lifted at day 30, to generate 3D ocular organoids consisting of retina, retinal pigment epithelium (RPE) and cornea. Organoids in suspension were cultured in medium containing low or high ATRA concentrations from days 30 to 90. Histology, immunochemistry, PCR and WB were used to study gene and protein differential expression among groups. Unpaired two-tailed Student’s t-test was applied to determine statistical significance between high and low ATRA concentrations groups. For more than two groups, 2-way ANOVA with Bonferroni posttest was used.
Results :
Significant differences were observed between tested groups revealing some effects of ATRA during early eye development. The presence of high ATRA concentration promotes transparent corneal organoids and neuroretinal development in retinal organoids while inhibiting the pigmentation of RPE organoids and the maturation of photoreceptors (PR). By contrast, low ATRA concentrations enhanced pigmentation of RPE organoids, the opacity of corneal organoids - due to an increase of collagen type IV in the stroma– and PR maturation in retinal organoids.
Conclusions :
ATRA modulates the corneal epithelial integrity and transparency, promotes PR development and maturation of the RPE in a dose-dependent manner. These results demonstrate the relevance of ATRA during ocular tissue development and suggest a potential new strategy to better modulate the development and maturation of ocular organoids by controlling ATRA concentrations temporally and spatially.
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.