Waves of cell death modulate the abundance and maturation of human retinal ganglion cells

Retinal ganglion cells (RGCs) are the projection neurons that transmit visual information from the retina to the brain via the optic nerve. A substantial proportion of RGCs are eliminated by programmed cell death during development to regulate their final number, yet how cell death impacts RGC development in human retinas remains poorly understood. Here, we characterized the timing and cell-type-specificity of cell death in human fetal retinas and retinal organoids. We identified two waves of apoptosis: an early wave targeting retinal neuronal precursors and a late wave affecting RGCs and other neurons. In addition to these two waves of apoptosis, organoids displayed a distinct wave of necrosis in the innermost core region that eliminates nearly all RGCs during long-term culture. To investigate the functions of the apoptotic waves during retinal development, we differentiated BAX/BAK double mutant organoids that are deficient for apoptosis. In these mutant organoids, RGC survival was improved, RGC neurogenesis and maturation were delayed, and RGCs were lost through a compensatory increase in necrosis. Our data suggest that the waves of developmental apoptosis promote human RGC neurogenesis and maturation. Together, our results highlight the roles of cell death in human RGC development and the challenges in retinal organoid design. Overcoming these obstacles would improve the utility of retinal organoids for modeling optic neuropathies like glaucoma Overall design: Wildtype or BAX/BAK double mutant organoids human retinal organoids at different ages were lysed into a single nuclei suspension and run on the 10x Genomics Chromium Single Cell System platform using the Single Cell Multiome ATAC + Gene Expression chemistry. Libraries were sequenced on Illumina NovaSeq X and processed through the Cell Ranger pipeline using the GRCh38 reference genome.