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. Author manuscript; available in PMC: 2013 May 7.
Published in final edited form as: J Neurosci. 2012 Nov 7;32(45):15934–15945. doi: 10.1523/JNEUROSCI.3227-12.2012

Fig. 6. BMP4 sufficiency to induce CPEC markers from hESC-derived neural progenitors.

Fig. 6

(A) Schematic of the differentiation method. The hESCs form multiple and less uniform SFEBq aggregates than mESCs. (B) Temporal analysis of neural induction in hSFEBq aggregates (9000 hESCs/well, RT-qPCR normalized to hESCs). Dramatic increases in forebrain (LHX2, FOXG1) and generic neural progenitor gene expression (SOX1, NESTIN) occur by 6 DIV and are stably maintained thereafter. (C) ICC of 9000 cells/well hSFEBq aggregates at 21 DIV for PAX6 and NESTIN demonstrate efficient neural induction. (D) Temporal- and BMP4 concentration-dependent induction of CPEC genes in hSFEBq aggregates (9000 cells/well, RT-qPCR normalized to no BMP4 controls). 18-day aggregates show marked BMP4-mediated upregulation of TTR and LMX1A, while little to no induction is seen in 9-, 12-, or 15-day aggregates. TTR and LMX1A baselines also increased with aggregation time (~64X for TTR, 4X for LMX1A), but these baseline changes were much less than the BMP4-mediated changes in 18-day aggregates (>1000X for TTR, >100X for LMX1A). (E) CPEC and neural progenitor markers, 18-day aggregates (RT-qPCR normalized to no BMP4 controls). BMP4 induces CPEC markers TTR, LMX1A, and OTX2 at the expense of neural progenitor markers FOXG1 and NESTIN. Scale bars: 50 um.