In this study, we have expanded cerebral organoids as an experimental platform to analyze the early human cortical development in healthy and disease states and to investigate disease mechanisms. Using the protocol of Lancaster et al.,40 in which the first step, an efficient generation of EBs, was modified as in Brennand et al.,33 we successfully generated cerebral organoids from hESCs (H9 and HUES8 lines) and from seven human iPSC lines. We observed time-dependent generation and development of organoids, recapitulating the inside-out pattern of human cortical development. The VZ in the center of the rosettes contained proliferating GFAP+ radial glia, the IZ contained migrating doublecortin+ neuroblasts and immature βIII-tubulin+ neurons, and the CZ incorporated pioneer neurons (TBR1+), and built cortical-like layers of the βIII-tubulin+ and Pan-Neu+ neurons, and calretinin+ interneurons. We employed quantitative cell counting, image intensity measurements, and computational tools to quantify and analyze normal and pathological tissue samples.
