Fig. 1

Differentiation of WTC-11 induced pluripotent stem cells (iPSC) into motor neurons (MN). a Schematic diagram of the overall experimental design. Shown are the small molecule stimuli for each developmental stage and the corresponding morphological changes in the course of iPSC conversion into MN. The scale bar for the phase contrast images is 400 μm; (b) Representative immunofluorescence images of key pluripotency (Oct4) and pan-neuronal (Nestin, Pax6, and β-III tubulin) biomarkers indicated at efficient iPSC differentiation into neuronal stem cells (NSC) at day 7th post-induction (accumulation of Nestin is shown in orange). Upregulation of Pax6 and β-III tubulin (shown in green) at day 13 of iPSC induction marked the formation of motor neuron progenitor (MNP) cells. c Immunostaining of neuronal structural proteins (beta-III-tubulin in green, MAP2 in orange), motor neuron specific markers (HB9 and ChAT in green), and synaptic vesicles protein (Synaptophysin in green) showed a pure population of mature MN at day 28 post-stimulation of iPSC. d Image quantification of Pax6- and HB9-labeled cells exemplified the percentage of cells that transitioned from NSC to MN. e Immunocytochemistry with anti-ISL1 antibody and quantitative analysis of biomarker positive cells at D28 of the differentiation protocol. The graphs show average of 6 cultures with > 300 cells in random fields for each culture. Scale bar: 100 μm