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A feeder-free, human plasma-derived hydrogel for maintenance of a human embryonic stem cell phenotype in vitro

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Author(s): Lewis Fiona C | Bryan Nicholas | Hunt John A

Journal: Cell Regeneration
ISSN 2045-9769

Volume: 1;
Issue: 1;
Start page: 6;
Date: 2012;
Original page

Keywords: Embryonic stem cells | Hydrogel | Pluripotent stem cells | Cell culture

ABSTRACT
Abstract Background Human embryonic stem cells (hESCs) represent a tremendous resource for cell therapies and the study of human development; however to maintain their undifferentiated state in vitro they routinely require the use of mouse embryonic fibroblast (MEF) feeder-layers and exogenous protein media supplementation. Results These well established requirements can be overcome and in this study, it will be demonstrated that phenotypic stability of hESCs can be maintained using a novel, human plasma protein-based hydrogel as an extracellular culture matrix without the use of feeder cell co-culture. hESCs were resuspended in human platelet poor plasma (PPP), which was gelled by the addition of calcium containing DMEM-based hESC culture medium. Phenotypic and genomic expression of the pluripotency markers OCT4, NANOG and SOX2 were measured using immunohistochemistry and qRT-PCR respectively. Typical hESC morphology was demonstrated throughout in vitro culture and both viability and phenotypic stability were maintained throughout extended culture, up to 25 passages. Conclusions PPP-derived hydrogel has demonstrated to be an efficacious alternative to MEF co-culture with its hydrophilicity allowing for this substrate to be delivered via minimally invasive procedures in a liquid phase with polymerization ensuing in situ. Together this provides a novel technique for the study of this unique group of stem cells in either 2D or 3D both in vitro and in vivo.
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