A Facile Method for Generating Cerebral Organoids from Human Pluripotent Stem Cells

Human cerebral organoids (COs) provide a human-specific platform to study the cellular and molecular processes that underlie different neurological events. Over the last decade, significant progress has been made in establishing reliable protocols to produce neural organoids. However, the variable result obtained from in vitro differentiation of human pluripotent stem cells (hPSCs) toward neural cell lineages, which is mainly affected by different signaling pathways activity, has been stated as one of the main obstacles ahead. In this study, we considered some of the known sources of these variable signaling cues arising from cell culture media components and sought to remove or offset their effects by applying an optimized and complete combination of small molecules and growth factors during differentiation protocol for the COs generation. Histological analysis demonstrated that these COs can display a faithful recapitulation of the neural progenitor zone and early cortical layers organization, containing different types of neuronal and glial cells which were in accordance with single-nucleus transcriptome profiling results. Moreover, recording the electrophysiological activities indicated that the COs can behave as a functional neural network. Based on the obtained results, this optimized method proposes a facile and more affordable protocol for generating hPSC-derived COs that can faithfully mimic the features of their in vivo counterparts in the developing human brain.