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Human feeders support prolonged undifferentiated growth of human inner cell masses and embryonic stem cells

Nature Biotechnology volume 20, pages 933–936 (2002)Cite this article

Abstract

Previous reports have demonstrated the growth of undifferentiated human embryonic stem (HES) cells on mouse embryonic fibroblast (MEF) feeders and on laminin- or Matrigel-coated plastic surfaces supplemented with MEF-conditioned medium1,2,3. These xenosupport systems run the risk of cross-transfer of animal pathogens from the animal feeder, matrix, or conditioned medium to the HES cells, thus compromising later clinical application. Here we show that human fetal and adult fibroblast feeders support prolonged undifferentiated HES cell growth of existing cell lines and are superior to cell-free matrices (collagen I, human extracellular matrix, Matrigel, and laminin) supplemented with human or MEF feeder–conditioned medium. Additionally, we report the derivation and establishment of a new HES cell line in completely animal-free conditions. Like HES cells cultured on MEF feeders, the HES cells grown on human feeders had normal karyotypes, tested positive for alkaline phosphatase activity, expressed Oct-4 and cell surface markers including SSEA-3, SSEA-4, Tra 1-60, and GCTM-2, formed teratomas in severely combined immunodeficient (SCID) mice, and retained all key morphological characteristics. Human feeder–supported HES cells should provide a safer alternative to existing HES cell lines in therapeutic applications.

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Figure 1: Morphology of HES cells grown on cell-free matrices and human feeder cells.
Figure 2: Characterization of new HES cell line (ten passages, 90 days) isolated and passaged from ICM stage onwards on FM fibroblasts and H2 medium containing 20% (vol/vol) human serum.

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Acknowledgements

We gratefully acknowledge the assistance of L. Retnam of the Animal Holding Unit, National University of Singapore for his help with the SCID mice work. This study was supported by a grant from Embryonic Stem Cell International (ESI).

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Authors and Affiliations

  1. Department of Obstetrics and Gynaecology, National University of Singapore, Kent Ridge, 119260, Singapore

    Mark Richards, Chui-Yee Fong, Peng-Cheang Wong & Ariff Bongso

  2. Department of Biological Sciences, National University of Singapore, Kent Ridge, 119260, Singapore

    Woon-Khiong Chan

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  1. Mark Richards
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Correspondence to Ariff Bongso.

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Competing interests

Financial support in terms of consumables and employment for M.R. was provided by ESI (Embryonic Stem Cell International) Pte Ltd.

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Richards, M., Fong, CY., Chan, WK. et al. Human feeders support prolonged undifferentiated growth of human inner cell masses and embryonic stem cells. Nat Biotechnol 20, 933–936 (2002). https://doi.org/10.1038/nbt726

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