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) All Versions of this Article: 2005-0112v1 23/9/1333    most recent Alert me when this article is cited Alert me if a correction is posted Citation Map Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints/Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Fok, E. Y.L. Articles by Zandstra, P. W. Search for Related Content PubMed PubMed Citation Articles by Fok, E. Y.L. Articles by Zandstra, P. W.

Shear-Controlled Single-Step Mouse Embryonic Stem Cell Expansion and Embryoid Body–Based Differentiation

Department of Chemical Engineering and Applied Chemistry, Institute of Biomaterials and Biomedical Engineering, University of Toronto, Toronto, Ontario, Canada

Key Words. Embryonic stem cells • Embryoid bodies • Scalable bioreactor • Stirred-suspension culture • Microcarriers • Aggregates • E-cadherin

Correspondence: Peter W. Zandstra, Ph.D., Institute of Biomaterials and Biomedical Engineering, Room 407, Roseburgh Building, 4 Taddle Creek Road, Toronto, Ontario, Canada M5S 3G9. Telephone: 416-978-8888; Fax: 416-978-4317; e-mail: peter.zandstra{at}utoronto.ca

To facilitate the exploitation of embryonic stem cells (ESCs) and ESC-derived cells, scale-up of cell production and optimization of culture conditions are necessary. Conventional ESC culture methods are impractical for large-scale cell production and lack robust microenvironmental control. We developed two stirred-suspension culture systems for the propagation of undifferentiated ESCs—microcarrier and aggregate cultures—and compared them with tissue-culture flask and Petri dish controls. ESCs cultured on glass microcarriers had population doubling times (~14–17 hours) comparable to tissue-culture flask controls. ESC growth could be elicited in shear-controlled stirred-suspension culture, with population doubling times ranging between 24 and 39 hours at 100 rpm impeller speed. Upon removal of leukemia inhibitory factor, the size-controlled ESC aggregates developed into embryoid bodies (EBs) capable of multilineage differentiation. A comprehensive analysis of ESC developmental potential, including flow cytometry for Oct-4, SSEA-1, and E-cadherinprotein expression, reverse transcription–polymerase chain reaction for Flk-1, HNF3-ß, MHC, and Sox-1 gene expression, and EB differentiation analysis, demonstrated that the suspension-cultured ESCs retained the developmental potential of the starting cell population. Analysis of E-cadherin^–/– and E-cadherin^+/– cells using both systems provided insight into the mechanisms behind the role of cell aggregation control, which is fundamental to these observations. These cell-culture tools should prove useful for both the production of ESCs and ESC-derived cells and for investigations into adhesion, survival, and differentiation phenomena during ESC propagation and differentiation.

This article has been cited by other articles:

				R. L. Carpenedo, C. Y. Sargent, and T. C. McDevitt Rotary Suspension Culture Enhances the Efficiency, Yield, and Homogeneity of Embryoid Body Differentiation Stem Cells, September 1, 2007; 25(9): 2224 - 2234. [Abstract] [Full Text] [PDF]

				A. Ouyang, R. Ng, and S.-T. Yang Long-Term Culturing of Undifferentiated Embryonic Stem Cells in Conditioned Media and Three-Dimensional Fibrous Matrices Without Extracellular Matrix Coating Stem Cells, February 1, 2007; 25(2): 447 - 454. [Abstract] [Full Text] [PDF]

				K. Kim, P. Lerou, A. Yabuuchi, C. Lengerke, K. Ng, J. West, A. Kirby, M. J. Daly, and G. Q. Daley Histocompatible Embryonic Stem Cells by Parthenogenesis Science, January 26, 2007; 315(5811): 482 - 486. [Abstract] [Full Text] [PDF]