HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text Full Text (PDF) An addition or correction has been published All Versions of this Article: 73/6/1147    most recent biolreprod.104.036673v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Khoo, M. L.M. Articles by Tuch, B. E. Search for Related Content PubMed PubMed Citation Articles by Khoo, M. L.M. Articles by Tuch, B. E. Agricola Articles by Khoo, M. L.M. Articles by Tuch, B. E.

Growth and Differentiation of Embryoid Bodies Derived from Human Embryonic Stem Cells: Effect of Glucose and Basic Fibroblast Growth Factor¹

Diabetes Transplant Unit, Prince of Wales Hospital and The University of New South Wales, Sydney, New South Wales 2031, Australia

ABSTRACT

Differentiation of embryonic stem (ES) cells generally occurs after formation of three-dimensional cell aggregates, known as embryoid bodies (EBs). This differentiation occurs following suspension culturing of EBs in media containing a high (25 mM) glucose concentration. Although high-glucose-containing media is used for maintenance and proliferation of ES cells, it has not been demonstrated whether this is a necessary requirement for EB development. To address this, we examined the growth and differentiation of EBs established in 0-mM, 5.5-mM (physiological), and 25-mM (high) glucose concentrations, through morphometric analysis and examination of gene and protein expression. The effect on EB development of supplementation with basic fibroblast growth factor (FGF2) was also studied. We report that the greatest rate of EB growth occurs in 5.5 mM glucose media. A morphological study of EBs over 104 days duration under glucose-containing conditions demonstrated the development of all three major embryonic cell types. The difference from normal human development was obvious in the lack of rostrocaudal control by the notochord. In the latest stages of development, the main tissue observed appeared to be cartilage and cells of a mesodermal lineage. We conclude that physiological glucose concentrations are suitable for the culturing of EBs, that the addition of FGF2 enhances the temporal expression of genes including POU5F1, nestin, FOXA2, ONECUT1, NEUROD1, PAX6, and insulin, and that EBs can be cultured in vitro for long periods, allowing for further examination of developmental processes.

developmental biology, early development, embryo, gene regulation

¹ Support was provided by a project grant from the Sydney Foundation for Medical Research. M.K. was the recipient of The University of New South Wales Faculty of Science Scholarship. J.L. was the recipient of the Home Wilkinson Lowry Diabetes Stem Cell Scholarship at The University of New South Wales.

² Correspondence: Bernard E. Tuch, Diabetes Transplant Unit, Prince of Wales Hospital, Randwick, N.S.W. 2031 Australia. FAX: 61 2 93824826; b.tuch{at}unsw.edu.au

This article has been cited by other articles:

				G. M. Hoben, E. J. Koay, and K. A. Athanasiou Fibrochondrogenesis in Two Embryonic Stem Cell Lines: Effects of Differentiation Timelines Stem Cells, February 1, 2008; 26(2): 422 - 430. [Abstract] [Full Text] [PDF]