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Hum. Reprod. Advance Access originally published online on June 25, 2009
Human Reproduction 2009 24(10):2567-2581; doi:10.1093/humrep/dep232
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© The Author 2009. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oxfordjournals.org

Immortalized human skin fibroblast feeder cells support growth and maintenance of both human embryonic and induced pluripotent stem cells

Christian Unger1,{dagger}, Shuping Gao2,{dagger}, Marie Cohen2, Marisa Jaconi3, Rosita Bergstrom1, Frida Holm1, Amparo Galan4, Eva Sanchez4, Olivier Irion5, Jean Bernard Dubuisson5, Marc Giry-Laterriere1, Patrick Salmon6, Carlos Simon4, Outi Hovatta1,2,{dagger} and Anis Feki2,5,7,{dagger}

1 Division of Obstetrics and Gynaecology, Department of Clinical Science, Intervention and Technology, Karolinska Institutet, Stockholm, Sweden 2 Laboratory of Stem Cell Research, Geneva University Hospitals, 30, Bld de la Cluse, CH-1211 Geneva, Switzerland 3 Department of Pathology and Immunology, Faculty of Medicine, Geneva University, Geneva, Switzerland 4 Spanish Stem Cell Bank, Comunidad Valenciana Node, Prince Felipe Research Centre (CIPF), Valencia, Spain 5 Department of Obstetrics and Gynaecology, Geneva University Hospitals, Geneva, Switzerland 6 Department of Neurosciences, Faculty of Medicine, Geneva University, Geneva, Switzerland

7 Correspondence address. Tel: +41-22-38-24-392; Fax: +41-22-38-24-310; E-mail: anis.feki{at}hcuge.ch

BACKGROUND: Feeder cells are frequently used for the early-stage of derivation and culture of human embryonic stem cell (hESC) lines.

METHODS: We established a conditionally immortalized human foreskin fibroblast line that secreted basic fibroblast growth factor (bFGF). These cells were used as feeder cells for hESC culture and induced pluripotent stem (iPS) cell derivation and expansion. This conditional immortalization was performed using lentiviral vector (LV) mediated transduction of Bmi-1 and human telomerase reverse transcriptase genes and the resulting cell line was further modified by LV-mediated transduction of a secreted form of bFGF gene product. Three different laboratories have tested whether this feeder cell line could support the maintenance of four different hESC lines.

RESULTS: Immortalized fibroblasts secreting stable amounts of bFGF supported the growth of all hESC lines, which remained pluripotent and had a normal karyotype for at least 10 passages. Even at high passage (p56), these modified cells, when used as feeders, could support iPS derivation and propagation. Derived iPS cells expressed pluripotency markers, had hESC morphology and produced tissue components of the three germ layers when differentiated in vitro.

CONCLUSION: These modified fibroblasts are useful as a genetically-defined feeder cell line for reproducible and cost-effective culture of both hESC and iPS cells.

Key words: feeder cells/human embryonic stem cells/induced pluripotent stem cells/human fibroblasts/immortalization

{dagger} These authors contributed equally to this work.

Submitted on April 10, 2009; resubmitted on May 27, 2009; accepted on June 3, 2009.


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