Ciliary marginal zone of the developing human retina maintains retinal progenitor cells until late gestational stages

Non-mammalian vertebrates maintain a proliferative stem cell population at the far periphery of their retina called the ciliary marginal zone (CMZ), which gives rise to all retinal cell types and contributes to retinal regeneration upon injury. Humans do not maintain a proliferative CMZ into adulthood; however, it is not known how long in development this region continues to generate new neurons. Here, we identify a population of cells in the far peripheral retina of the fetal human that continues to proliferate long after the rest of the retina is quiescent. Single cell RNA-sequencing and EdU tracing at late time points in development reveal that this region has features of the non-mammalian CMZ, including the capacity to produce both early and late born cell types at late developmental stages, and a longer cell cycle than more centrally located retinal progenitor cells (RPCs). Moreover, while more central RPCs exit the cell cycle with the addition of a TGFb-inhibitor, we show that early RPCs within the CMZ do not. These findings define the late stages of neurogenesis in human retinal development, and present a unique model system to study the fetal CMZ in humans. Overall design: Retinospheres were generated from day 137 human fetal tissues. Retinospheres were sorted into CMZ containing retinospheres vs. those derived from the far periphery. Retinospheres from both groups were cultured for an additional 48 days, to an equivalent of 185 days fetal gestation. The CMZ of far-peripheral retinospheres was dissected and dissociated, and far peripheral retinospheres were also dissociated for comparison. An equivalent number of cells from each sample were then processed with the 10x Genomics single cell RNA-sequencing platform.