Characterization of late proliferative zone in developing human retina

Abstract

Purpose : Early stages of human retinal development have remained elusive due to low tissue availability and a lack of trackable experimental systems. Small pieces of human fetal retina form spheres (retinospheres) much like pluripotent stem cell-derived organoids, and can be maintained in vitro for many months. We used this system to determine when the process of neurogenesis is complete in the human retina.

Methods : Human fetal tissue was isolated and the far periphery of the retina was dissected to produce retinospheres. Retinospheres were made from central retina, peripheral retina and far peripheral retina, containing the ciliary epithelium. The retinospheres were generated from a range of different fetal ages and maintained for up to 245 days equivalent gestation. Proliferation was tracked by addition of EDU at several timepoints in development. We further compared retinospheres taken from peripheral or far peripheral retina using single-cell-RNA-sequencing. Transcriptomes of the late replicating cells were compared to replicating cells at different time points in human retinal development. Additionally, we characterized the cell cycle length of the proliferating cells in the far periphery of the retina.

Results : We show there is a small zone of cells between the laminated retina and the ciliary epithelium that retains cells that incorporate EdU long after the rest of the retina is mitotically quiescent. The cells of this zone have other similarities to the CMZ found in a similar region of non-mammalian vertebrates. The transcriptome of these late proliferating cells in the far peripheral retinal sample contains markers for early retinal progenitor cells including FGF19 and YAP1.

Conclusions : In conclusion, we have shown that there exists a late proliferating zone in the human retina that continues to proliferate for at least 245 days of development. This region continues to shrink and becomes localized to the far periphery of the retinosphere, closest to the ciliary epithelium. Single cell analysis indicates that these late replicating cells are similar to early progenitor cells, containing markers for early retinal progenitor cells including FGF19 and YAP1. These data suggest that human retina may have a transient CMZ-like zone of cells at the retinal margin, similar to the stem cell zone of non-mammalian vertebrates.

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