Derivation, characterization and differentiation of human embryonic stem cells: Comparing serum-containing versus serum-free media and evidence of germ cell differentiation

H. F. Chen, H. C. Kuo, C. L. Chien, C. T. Shun, Y. L. Yao, P. L. Ip, C. Y. Chuang, C. C. Wang, Y. S. Yang, H. N. Ho

Research output: Contribution to journalArticle

113 Citations (Scopus)

Abstract

Background: This study was designed to establish human embryonic stem cell (hESC) lines, to identify the differences when maintained in serum-containing versus serum-free medium and to test their potential of in vitro differentiation. Methods: Procedures including immunosurgery were performed on 11 donated human blastocysts to establish hESC lines. The cell lines were characterized and maintained using either serum-free or serum-containing media to compare their morphology, Oct-4 expression, apoptosis and growth speed. Differentiation of these lines was evaluated by the morphology and the expression of genes belonging to the three embryonic germ layers and the germ cell lineage. Results: Three hESC lines were established, and they grew at similar speed in both media (serum-containing or serum-free), but hESC cultured in serum-containing medium yielded significantly higher percentages of morphologically good colonies and cells expressing Oct-4. These cell lines differentiated spontaneously in vitro into cells expressing markers belonging to all three embryonic germ layers and germ cell markers, including c-Kit, STELLA, VASA and growth differentiation factor 9 (GDF9), in directly adherent culture. Conclusions: Three hESC lines with Taiwanese ancestry have been established, and they retain the in vitro differentiation potential with or without embryoid body (EB) formation. The data support that hESC may be capable of differentiation into germ cells although further confirmation is needed. It is also suggested that strategies such as stepwise adaptation will be needed before implementing a serum-free culture condition for hESC lines that have previously been derived in a medium containing serum.

Original languageEnglish
Pages (from-to)567-577
Number of pages11
JournalHuman Reproduction
Volume22
Issue number2
DOIs
Publication statusPublished - Jan 1 2007
Externally publishedYes

Fingerprint

Serum-Free Culture Media
Germ Cells
Cell Differentiation
Cell Line
Serum
Germ Layers
Growth Differentiation Factor 9
Embryoid Bodies
Human Embryonic Stem Cells
Blastocyst
Cell Lineage
Apoptosis
Gene Expression
Growth

Keywords

  • Differentiation
  • Embryoid body
  • Embryonic stem cells
  • Germ cell
  • Serum-free medium

ASJC Scopus subject areas

  • Reproductive Medicine
  • Obstetrics and Gynaecology

Cite this

Derivation, characterization and differentiation of human embryonic stem cells : Comparing serum-containing versus serum-free media and evidence of germ cell differentiation. / Chen, H. F.; Kuo, H. C.; Chien, C. L.; Shun, C. T.; Yao, Y. L.; Ip, P. L.; Chuang, C. Y.; Wang, C. C.; Yang, Y. S.; Ho, H. N.

In: Human Reproduction, Vol. 22, No. 2, 01.01.2007, p. 567-577.

Research output: Contribution to journalArticle

Chen, H. F. ; Kuo, H. C. ; Chien, C. L. ; Shun, C. T. ; Yao, Y. L. ; Ip, P. L. ; Chuang, C. Y. ; Wang, C. C. ; Yang, Y. S. ; Ho, H. N. / Derivation, characterization and differentiation of human embryonic stem cells : Comparing serum-containing versus serum-free media and evidence of germ cell differentiation. In: Human Reproduction. 2007 ; Vol. 22, No. 2. pp. 567-577.
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AU - Shun, C. T.

AU - Yao, Y. L.

AU - Ip, P. L.

AU - Chuang, C. Y.

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AB - Background: This study was designed to establish human embryonic stem cell (hESC) lines, to identify the differences when maintained in serum-containing versus serum-free medium and to test their potential of in vitro differentiation. Methods: Procedures including immunosurgery were performed on 11 donated human blastocysts to establish hESC lines. The cell lines were characterized and maintained using either serum-free or serum-containing media to compare their morphology, Oct-4 expression, apoptosis and growth speed. Differentiation of these lines was evaluated by the morphology and the expression of genes belonging to the three embryonic germ layers and the germ cell lineage. Results: Three hESC lines were established, and they grew at similar speed in both media (serum-containing or serum-free), but hESC cultured in serum-containing medium yielded significantly higher percentages of morphologically good colonies and cells expressing Oct-4. These cell lines differentiated spontaneously in vitro into cells expressing markers belonging to all three embryonic germ layers and germ cell markers, including c-Kit, STELLA, VASA and growth differentiation factor 9 (GDF9), in directly adherent culture. Conclusions: Three hESC lines with Taiwanese ancestry have been established, and they retain the in vitro differentiation potential with or without embryoid body (EB) formation. The data support that hESC may be capable of differentiation into germ cells although further confirmation is needed. It is also suggested that strategies such as stepwise adaptation will be needed before implementing a serum-free culture condition for hESC lines that have previously been derived in a medium containing serum.

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