Induction of Differentiation into Endoderm, Insulin-Secreting Cells from Mouse Embryonic Stem Cells Using Activin A and Retinoic Acid

抄録

Embryonic stem (ES) cells have been known to differentiate into various progenitor cells. In this study, we investigated the differentiation capacity of mouse ES cells into pancreatic hormone-secreting cells, and insulin-secreting cells. ES cells were cultured in Dulbecco’s modified Eagle medium (DMEM) after removal of leukemia inhibitory factor (LIF) for 3 days and then transferred in DMEM supplemented with retinoic acid or activin A. When the culture of embryoid bodies (EBs) derived from ES cells in DMEM added with activin A (2 × 10^-9 M or 2 × 10^-10 M) for 5 days was performed, endoderm marker genes, GATA4, Sox17 and Foxa2 were expressed in the EBs. However, at 6 days of culture, expression of Sox17 was not observed. When EBs were cultured with activin A (2 × 10^-9 M) for 6 days, and followed by 6 days of culture with retinoic acid (10^-6 M), expression of the pancreatic cell marker genes, GATA4 and Fxa2, were continued, and pancreatic islet genes, insulins 1 and 2, glucagons and somatostatin, were expressed from 5 days of culture. Immunohistochemistrical analysis gave results similar to RT-PCR. We consider this differentiation method for definitive endoderm production and pancreatic hormone—secreting cells, by using activin A and, retinoic acid is considered to be effective.