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Fig. 3. Differentiation of mesendoderm cells into definitive endoderm and mesoderm
lineages. (A) RT-PCR analysis of gene expression in
GFP+ECD+ and GFP+ECD- cells on day
5. (B) Multicolour immunofluorescent staining of cells in day 5 cultures.
Gsc+ECD+Foxa2+ cells are within the
epithelial sheet, whereas Gsc+ECD-Foxa2-
cells disperse around the sheet. (C) RT-PCR analysis of gene expression in
differentiated Gsc+ECD+ cells.
Gsc+ECD+ cells were purified from cultures with activin
on day 6 and re-cultured in SCM. Expression level of Gsc, Mixl1 and
Afp changes during culture. Gsc is positive only in the day
5 culture (A), Mixl1 is in the day 5 (A) and 12 cultures, and
AFP is only in the day 12 culture. (D) Albumin-producing cells are
generated from Gsc+ECD+ population. (i) Gsc+
cells were induced in SFM containing activin. On day 6,
Gsc+ECD+ cells were purified by cell-sorter and
re-cultured for 3 days on collagen I-coated dish in the presence of Egf, Bmp4,
acid Fgf and basic Fgf. (ii) The expression of albumin in the
Gsc+ECD+-derived cells detected by immunostaining. (iii)
The expression of albumin in the Gsc+ECD+-derived cells
detected by RT-PCR. Hnf6, a marker of precursor of liver and
pancreatic cells, was stimulated to be expressed.
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