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Figure 3 | BMC Cell Biology

Figure 3

From: All-trans retinoic acid promotes neural lineage entry by pluripotent embryonic stem cells via multiple pathways

Figure 3

Requirement for RA signaling for efficient neural specification of ESCs. A, RT-PCR for RARs, RXRs and RA metabolism-relative enzymes. RNA was isolated from ESCs (day 0), monolayer cultures (days 1, 3, 5 and 10) and adult mouse brain. Expression of RA nuclear receptors and two RA-metabolizing enzymes (CYP26a1 and Raldh2) were analyzed by RT-PCR; B, C, Q-PCR for RARα and CYP26a1 during monolayer differentiation; D-G, Monolayer differentiation of 46C ESCs in RA-treated cultures (N2B27 + 10-8 M RA) exposed to 2 μM RARα inhibitor Ro 415253 or to equivalent amounts of DMSO diluents; D, Cultures of monolayer differentiation, shown in phase contrast or stained for Sox1GFP; E, Percentage of Sox1GFP+ cells scored by FACS. #, significant difference between inhibitor-treated cultures and DMSO-treated cultures (p < 0.001); F, Cultures of monolayer differentiation, shown in stained for Oct4 or combination of Oct4 and Hoechst; G, Western blot analysis of Oct4 at day 5 under monolayer cultures; H, I, Monolayer differentiation of 46C ESCs in cultures without RA (N2B27) exposed to 2 μM RARα inhibitor Ro 415253 or to equivalent amounts of DMSO diluents at Day6; H, Cultures of monolayer differentiation, shown in phase contrast or stained for Sox1GFP; I, Percentage of Sox1GFP+ cells scored by FACS; Scale bars: D, 40 μm; F, H, 80 μm.

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