Role of RB1 in human embryonic stem cell-derived retinal organoids.

Three-dimensional (3D) organoid models derived from human pluripotent stem cells provide a platform for studying human development and understanding disease mechanisms. Most studies that examine biallelic inactivation of the cell cycle regulator Retinoblastoma 1 (RB1) and the link to retinoblastoma is in mice, however, less is known regarding the pathophysiological role of RB1 during human retinal development. To study the role of RB1 in early human retinal development and tumor formation, we generated retinal organoids from CRISPR/Cas9-derived RB1-null human embryonic stem cells (hESCs). We showed that RB is abundantly expressed in retinal progenitor cells in retinal organoids and loss of RB1 promotes S-phase entry. Furthermore, loss of RB1 resulted in widespread apoptosis and reduced the number of photoreceptor, ganglion, and bipolar cells. Interestingly, RB1 mutation in retinal organoids did not result in retinoblastoma formation in vitro or in the vitreous body of NOD/SCID immunodeficient mice. Together, our work identifies a crucial function for RB1 in human retinal development and suggests that RB1 deletion alone is not sufficient for tumor development, at least in human retinal organoids. A model of the role of RB1 during human retinal development. In wild-type cells, RB1 regulates entry into S-phase. While RB1 mutation in retinal organoids did not result in retinoblastoma formation, either in vitro or when injected into vitreous body of NOD/SCID mice, loss of RB1 promotes an accumulation of cells in S-phase, causes widespread apoptosis, and reduces the number of photoreceptor, ganglion and bipolar cells. Image 1 • RB expression is highest in progenitor cells and downregulated during maturation stages. • RB1 is required for S-phase entry and cell survival in retinal organoids. • RB1 is necessary for photoreceptor, ganglion and bipolar cell differentiation. • Loss of RB1 is not sufficient for retinoblastoma formation in retinal organoids. [ABSTRACT FROM AUTHOR]