Fig 2: Morphological characteristics of neurons generated via approaches A and B compared to NPC. (a) A schematic overview of approaches A and B differentiating hiPSC-derived NPC into MNs, created with http://BioRender.com. (b) Representative images of cells of approach A and B captured with the IncuCyte® showing MN-type clusters evolving from homogeneous NPC culture. (c) Neuronal identity was confirmed for both approaches via Bielschowsky silver staining, illustrated by brown-stained nuclei and axons. (d) Immunofluorescent staining for Tubb3 (beta-3-tubulin; red) revealed pronounced neuronal network formation via both approaches. Cell nuclei were counterstained with DAPI. All scale bars, 200 µm.

Fig 3: Quantitative real-time PCR analysis during ES cell differentiation.Time course of relative expression of (A) pluripotency genes (Zfp42, Oct4, Nanog) and differentiation marker (Fgf5), (B) cardiac specific genes (Myh6, Mlc2a) and mesodermal markers (Nkx2-5, Gata4 and T-bra), (C) catecholamine synthesis genes (TH, Ddc, Dbh, Pnmt), (D), endodermal marker genes (Sox17, Fox2a, Hnf4a, Alb, Afp), (E) ectodermal and neuronal markers (Map2, Tubb3, Nes). 14+P: d14 puromycin purified clusters; MEF: mouse embryonic fibroblasts. Values represent relative quantities (RQ) plotted on a log2 scale. Primers are listed in Table 1. (F) Semiquantitative reverse-transcription PCRs from ES cells (ES), control and reserpine-treated EBs during differentiation from day 2 to day 10 representative of the gene expression of a- and ß-AR subtypes (a1A, a1B, a1D, a2A, a2B, a2C, ß1, ß2, ß3; Gapdh was used as house-keeping control). L: DNA Ladder. Amplified fragment sizes are stated on the right of each pane.