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　 The generation of specific lineages of the definitive endoderm from embryonic stem (ES) cells is an important issue in developmental biology as well as in regenerative medicine. This study demonstrates that ES cells are induced sequentially into regional specific gut endoderm lineages, such as pancreatic, hepatic and other cell lineages, when they are cultured directly on a monolayer of mesoderm-derived supporting cells. A detailed chronological analysis revealed that Activin, FGF or BMP signals are critical at various steps, and that additional short range signals are required for differentiation into Pdx1-expressing cells. Under selective culture conditions, a high efficiency of definitive endoderm (47%) or Pdx1-posititive pancreatic progenitors (30%) are yielded. When transplanted under the kidney capsule, the Pdx1-positive cells further differentiated into all three pancreatic lineages, namely endocrine, exocrine and duct cells.

 

Figure 1: M15, a mesoderm-derived cell line, directs the differentiation of ES cells into Pdx1-expressing regional specific definitive endoderm cells.
ES cells grown on M15, without (left panel) or with (right panel) growth factors, differentiate into Pdx1 /GFP-expressing cells (green) on d8. A merged image of fluorescent and phase contrast image is shown. Numbers indicate the proportion of Pdx1 /GFP-expressing cells within total ES cell culture. Bar indicates 200 μm.

Figure 2: A schematic drawing of the signaling events in ES differentiation into Pdx1 + cells. Signaling events involved in the inductive processes observed in this study are shown in blue. Act, Activin; Ect, ectoderm; End, endoderm; ICM, inner cell mass; ME, mesendoderm; Mes, mesoderm; Epi, epiblast; p38, p38 MAP kinase, RA, retinoic acid.