Fig. 1.

ihNSC and iPS derived neurons. (A) The cartoon illustrates the maintenance and differentiation of ihNSCs. Cells are grown as neurospheres (N) in medium containing EGF and FGF-2, and mechanically disaggregated for expansion. After 24 h (D0; D: days of differentiation), cells appear as doublets which are plated on laminin-coated surfaces and grown without EGF for 3 days, in order to induce progenitor differentiation. FGF-2 is then removed to promote the generation of mature neurons, oligodendrocytes and astrocytes. The different cell types are detected by IF using lineage-specific phenotypic markers. (B) A-T patient-derived skin fibroblasts are reprogrammed into iPS cells. iPS cells express the pluripotency markers Tra-1-81 and upon neutralization give rise to a self-renewing population of NPCs expressing Nestin. NPCs can be further differentiated into mature neurons expressing MAP2. A-T iPS cells are ATM-deficient and express the pluripotency factor Oct3/4 (western blot, left). Upon neuralization, NPCs downregulate Oct3/4 and express the neural markers Nestin and Sox2, which decrease from D0 to D14 whereas β-Tubulin III increases (western blot, right). Protein loading per lane was verified with anti-vinculin antibodies.