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. 2013 Nov 27;1(6):545–559. doi: 10.1016/j.stemcr.2013.10.006

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© 2013 The Authors

This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution and reproduction in any medium, provided the original author and source are credited.

PMC Copyright notice

A Single TF Induces Neurons from ESCs and Specification of Neural Types

(A) Smad7, Nr2f1, Sox11, Dmrt1, Sox9, Foxg1, Klf3, Pou5f1, Sox2, and Ascl1 are identified for ESC differentiation toward neural tissues/organs.

(B and C) Immunostaining of Ascl1-overexpressing ESCs cultured in the DMs for TUJ1, MAP2, NEUN, SYNAPSIN, TH, DAT, ISL1/ISL2, or GABA on day 7 of differentiation is presented. Scale bars, 200 μm.

(D) Representative traces show action potentials recorded from neurons induced by Ascl1 on day 11 of differentiation.

(E) Representative traces show postsynaptic currents recorded at −90 mV holding potential. Reversal potential that was calculated from traces was −25.5 mV.

(F) Immunostaining of Smad7- or Nr2f1-overexpressing ESCs for TUJ1 on day 7 of differentiation (top panels) is shown. Scale bars, 200 μm. FACS analysis on day 6 of differentiation for PSA-NCAM (neural progenitor marker)⁺ cell appearance used ESCs carrying Smad7 or Nr2f1 gene (bottom panels).

(G) Quantitative evaluation of PSA-NCAM⁺ cell percentages used ESCs carrying Smad7, Nr2f1, Sox11, Dmrt1, Sox9, Foxg1, Klf3, Pou5f1, Sox2, or Ascl1 gene by FACS analysis (three independent experiments, SEM; ^∗p<0.05 and ^∗∗p<0.01 versus Dox+).

(H) Quantitative evaluation of GABA⁺/TUJ1⁺, TH⁺/TUJ1⁺, and ISL1/ISL2⁺/TUJ1⁺ cell percentages used ESCs carrying Ascl1, Smad7, and Nr2f1 by immunostaining (three independent experiments, SEM; ^∗p < 0.05).

(I) Immunostaining of Smad7- or Nr2f1-overexpressing ESCs for TUJ1 (left panels) and GABA (middle panels) on day 7 of differentiation is shown. Scale bars, 200 μm.

See also Figures S1, S2, and S4.