From confluent human iPS cells to self-forming neural retina and retinal pigmented epithelium

Significance Human induced pluripotent stem cells (hiPSCs) could be used as an unlimited source of retinal cells for the treatment of retinal degenerative diseases. The production of retinal cells from hiPSCs for personalized therapeutic approaches must comply with certain criteria, such as safety, efficiency, reproducibility, and low production cost. Here, we report a simple and scalable retinal differentiation process for the generation of retinal pigmented epithelial cells and neural retinal tissues containing retinal progenitor cells. These progenitors can be differentiated into all retinal cell types, including retinal ganglion cells and precursors of photoreceptors, which could find important applications in regenerative medicine. This method also provides an accessible in vitro model to investigate mechanisms involved in human retinogenesis and retinal diseases. Progress in retinal-cell therapy derived from human pluripotent stem cells currently faces technical challenges that require the development of easy and standardized protocols. Here, we developed a simple retinal differentiation method, based on confluent human induced pluripotent stem cells (hiPSC), bypassing embryoid body formation and the use of exogenous molecules, coating, or Matrigel. In 2 wk, we generated both retinal pigmented epithelial cells and self-forming neural retina (NR)-like structures containing retinal progenitor cells (RPCs). We report sequential differentiation from RPCs to the seven neuroretinal cell types in maturated NR-like structures as floating cultures, thereby revealing the multipotency of RPCs generated from integration-free hiPSCs. Furthermore, Notch pathway inhibition boosted the generation of photoreceptor precursor cells, crucial in establishing cell therapy strategies. This innovative process proposed here provides a readily efficient and scalable approach to produce retinal cells for regenerative medicine and for drug-screening purposes, as well as an in vitro model of human retinal development and disease.