Vukasin M. Jovanovic,1,∗ Claire Weber,1 Jaroslav Slamecka,1 Seungmi Ryu,1 Pei-Hsuan Chu,1 Chaitali Sen,1 Jason Inman,1 Juliana Ferreira De Sousa,1 Elena Barnaeva,1 Marissa Hirst,2 David Galbraith,2 Pinar Ormanoglu,1 Yogita Jethmalani,1 Jennifer Colon Mercado,3 Sam Michael,1 Michael E. Ward,3 Anton Simeonov,1 Ty C. Voss,1 Carlos A. Tristan,1 and Ilyas Singeç1,∗∗
1National Center for Advancing Translational Sciences (NCATS), Division of Preclinical Innovation, Stem Cell Translation Laboratory (SCTL), National Institutes of Health, Rockville, MD 20850, USA
2Rancho Biosciences, San Diego, CA 92127, USA
3Inherited Neurodegenerative Disease Unit, National Institute of Neurological Disorders and Stroke (NINDS), National Institutes of Health, Bethesda, MD 20892, USA
Human gliogenesis remains poorly understood, and derivation of astrocytes from human pluripotent stem cells (hPSCs) is inefficient and cumbersome. Here, we report controlled glial differentiation from hPSCs that bypasses neurogenesis, which otherwise precedes astrogliogenesis during brain development and in vitro differentiation. hPSCs were first differentiated into radial glial cells (RGCs) resembling resident RGCs of the fetal telencephalon, and modulation of specific cell signaling pathways resulted in direct and stepwise induction of key astroglial markers (NFIA, NFIB, SOX9, CD44, S100B, glial fibrillary acidic protein [GFAP]). Transcriptomic and genome-wide epigenetic mapping and single-cell analysis confirmed RGC-to-astrocyte differentiation, obviating neurogenesis and the gliogenic switch. Detailed molecular and cellular characterization experiments uncovered new mechanisms and markers for human RGCs and astrocytes. In summary, establishment of a glia-exclusive neural lineage progression model serves as a unique serum-free platform of manufacturing large numbers of RGCs and astrocytes for neuroscience, disease modeling (e.g., Alexander disease), and regenerative medicine.