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In Vitro Differentiation of Mouse Embryonic Stem Cells: Enrichment of Endodermal Cells in the Embryoid Body

^a Department of Surgery, Stem Cell Therapy Center, Soonchunhyang University Hospital, Seoul, Korea;
^b Division of Genome and Proteome Research, National Genome Research Institute, NIH Korea, Seoul, Korea;
^c Division of Genetic Disease, Department of Biomedical Science, National Institute of Health, Seoul, Korea;
^d Department of Pathology, Hanyang University Hospital, Seoul, Korea;
^e Department of Biochemistry, College of Medicine, Ewha Womans University, Seoul, Korea;
^f Department of Animal Biotechnology, Graduate School of Bio & Information Technology, Hankyong National University, Ansung, Korea;
^g Department of Surgery, Hanyang University Hospital, Seoul, Korea

Key Words. Embryonic stem cell • Embryoid body • Differentiation • Endoderm

Correspondence: Bermseok Oh, Ph.D., Nokbun-Dong 5, Eunpyung-Gu, Seoul, 122-701, Korea. Telephone: 82-2-380-1523; Fax: 82-2-354-1063; e-mail: ohbs{at}nih.go.kr

Embryonic stem (ES) cells have the potential to differentiate into all three germ layers, providing new perspectives not only for embryonic development but also for the application in cell replacement therapies. Even though the formation of an embryoid body (EB) in a suspension culture has been the most popular method to differentiate ES cells into a wide range of cells, not much is known about the characteristics of EB cells. To this end, we investigated the process of EB formation in the suspension culture of ES cells at weekly intervals for up to 6 weeks. We observed that the central apoptotic area is most active in the first week of EB formation and that the cell adhesion molecules, except ß-catenin, are highly expressed throughout the examination period. The sequential expression of endodermal genes in EBs during the 6-week culture correlated closely with that of normal embryo development. The outer surface of EBs stained positive for -fetoprotein and GATA-4. When isolated from the 2-week-old EB by trypsin treatment, these endodermal lineage cells matured in vitro into hepatocytes upon stimulation with various hepatotrophic factors. In conclusion, our results demonstrate that endodermal cells can be retrieved from EBs and matured into specific cell types, opening new therapeutic usage of these in vitro differentiated cells in the cell replacement therapy of various diseases.

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