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. 2016 Jul 14;7(2):149–157. doi: 10.1016/j.stemcr.2016.06.007

Figure 1.

Figure 1

Differentiation Protocol Schematic, and Expression of Anterior Neural Progenitor Markers after Neural Induction

(A) Schematic representation of the protocol. For the first 10 days the cells were treated with dual SMAD inhibition (dSMADi) using dorsomorphin and SB431542. FGF8 was added at d11, and Notch inhibitor DAPT was added at d21.

(B) Real-time qPCR results at d10 showing an increased expression of pan-neural marker SOX1 and forebrain- and olfactory placode-associated genes EMX2 and FOXG1. Preplacodal markers SIX1 and EYA1 remained low. Expression levels are relative to d0 hPSCs (HEL11.4 and H9 representative experiments, n = 6, mean ± SEM).

(C) Relative expression of anterior neural progenitor (EMX2, FOXG1, DLX2, and DLX5), ventral forebrain (NKX2.1), and caudal CNS markers (PAX5 and GBX2) in FGF8-treated cells confirming anterior neural identity. HEL11.4-derived cells, n = 3 (mean + SEM). Corresponding data from H9-derived cells are shown in Figure S1B.

(D) Immunocytochemistry at d21 revealed an abundant expression of SOX2, FOXG1, and Ki-67 within neural rosettes. TUJ1 was found mainly outside the neural rosette structures. HEL11.4-derived cells. Scale bars, 100 μm.