Figure 1.

Combination of dual SMAD inhibition with WNT activation converts human iPSCs into caudal/ventral NSCs in 6 days

(A) Schematic diagram of the MN differentiation protocol from iPSCs.

(B) Representative images of NESTIN⁺/PAX6⁺ and SOX2⁺/OCT4^– NSCs generated after 6 days of SB + DMH1 + CHIR treatment.

(C) The proportion of NESTIN⁺, PAX6⁺, and SOX2⁺ cells on day 6 of differentiation, N = 7 cell lines, n = 3 replicates.

(D) RT-qPCR analysis of NESTIN, PAX6, SOX2 and OCT4 mRNA expression after 6 days of SB + DMH1+CHIR treatment, N = 2, n = 3.

(E) CHIR-induced WNT activation conferred a caudal identity of the NSCs expressing fore/midbrain marker OTX2 and hindbrain/spinal markers of HOXB4 and HOXC5. N = 4, n = 3.

(F) The NSCs were positively immunostained for a ventral marker NKX6.1 after 6 days of SB + DMH1+CHIR treatment.

(G) RT-qPCR analysis showed activation of NKX6.1 and OLIG2 mRNA by addition of CHIR, N = 1, n = 3. All representative images were from the control line of ALSH84C5, Scale bar, 100 μM.

See also Figure S2.