FGF2 Secreting Human Fibroblast Feeder Cells:A Novel Culture System for Human Embryonic Stem Cells

Saxena, S and Deb, K and Sharma, V and Totey, S and Hanwate, M (2008) FGF2 Secreting Human Fibroblast Feeder Cells:A Novel Culture System for Human Embryonic Stem Cells. Molecular Reproduction and Development, 75 (10). pp. 1523-1532. ISSN 1098-2795

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Abstract

Human embryonic stem cell (hESC) lines are traditionally derived and maintained on mouse embryonic fibroblasts (MEF) which are xenogeneic and enter senescence rapidly. In view of the clinical implications of hESCs, the use of human fibroblast as feeders has been suggested as a plausible alternative. However, use of fibroblast cells from varying sources leads to culture variations along with the need to add FGF2 in cultures to sustain ES cell pluripotency. In this study we report the derivation of FGF2 expressing germ layer derived fibroblast cells (GLDF) from hESC lines. These feeders could support the pluripotency, karyotypes and proliferation of hESCs with or without FGF2 in prolonged cultures as efficiently as that on MEF. GLDF cells were derived from embryoid bodies and characterized for expression of fibroblast markers by RT-PCR, Immunofluorescence and by flow cytometry for CD marker expression. The expression and secretion of FGF2 was confirmed by RT-PCR, Western blot, and ELISA. The hESC lines cultured on MEF and GLDF were analyzed for various stemness markers. These feeder cells with fibroblast cells like properties maintained the properties of hESCs in prolonged culture over 30 passages. Proliferation and pluripotency of hESCs on GLDF was comparable to that on mouse feeders. Further we discovered that these GLDF cells could secrete FGF2 and maintained pluripotency of hESC cultures even in the absence of supplemental FGF2. To our knowledge, this is the first study reporting a novel hESC culture system which does not warrant FGF2 supplementation, thereby reducing the cost of hESC cultures.

Item Type: Article
Additional Information: Copyright of this article is belongs to John Wiley and Sons.
Uncontrolled Keywords: embryonic stem cells;feeder cells;pluripotency.
Subjects: Regenerative Medicine > MIRM Bangalore
Depositing User: MCON Library
Date Deposited: 20 Mar 2012 06:48
Last Modified: 20 Mar 2012 06:48
URI: http://eprints.manipal.edu/id/eprint/3547

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