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. 2022 Apr 26;9(2):ENEURO.0442-21.2022. doi: 10.1523/ENEURO.0442-21.2022

Figure 6.

Figure 6.

Characterization of neuronal differentiation in SAG-treated ES-Hypo. A–G, Serial sections from SFEBq-cultured mESC aggregates (with SAG) on days 7 and 13. The sections were immunostained for Nkx2.1/Nkx2.2 (#1), Nkx2.1/HuC/D (#2), or Nkx2.1/Sox1 (#3). The day-13 aggregate contains a HuC/D+ neuron-dense area (B–D) and a Sox1+ rosette structure (E–G). Scale bars: 100 μm (A) and 50 μm (B–G). H, Representative images of SAG-treated mESC aggregates immunostained for MCH/Nkx2.1 (top) and MCH/Nkx2.2 (bottom) on day 22. Arrows indicate double-positive cells. Scale bar: 20 μm. I, The percentage of MCH-ir cells expressing Nkx2.1 or Nkx2.2 on day 22. n = 4–5 aggregates per marker. J, Triple immunostaing of SAG-treated mESC aggregates for MCH/CART/Nkx2.2 on day 30. Representative images of MCH+CART+ (left) and MCH+CART− (right) cell clusters are shown, and MCH+Nkx2.2+ cells are indicated by arrows. Scale bar: 20 μm.