Abstract
Improved biomaterials are required for application in regenerative medicine, biosensing, and as medical devices. The response of cells to the chemistry of polymers cultured in media is generally regarded as being dominated by proteins adsorbed to the surface. Here we use mass spectrometry to identify proteins adsorbed from a complex mouse embryonic fibroblast (MEF) conditioned medium found to support pluripotent human embryonic stem cell (hESC) expansion on a plasma etched tissue culture polystyrene surface. A total of 71 proteins were identified, of which 14 uniquely correlated with the surface on which pluripotent stem cell expansion was achieved. We have developed a microarray combinatorial protein spotting approach to test the potential of these 14 proteins to support expansion of a hESC cell line (HUES-7) and a human induced pluripotent stem cell line (ReBl-PAT) on a novel polymer (N-(4-Hydroxyphenyl) methacrylamide). These proteins were spotted to form a primary array yielding several protein mixture 'hits' that enhanced cell attachment to the polymer. A second array was generated to test the function of a refined set of protein mixtures. We found that a combination of heat shock protein 90 and heat shock protein-1 encourage elevated adherence of pluripotent stem cells at a level comparable to fibronectin pre-treatment.
Highlights
Human embryonic stem cells are pluripotent cells derived from the inner cell mass (ICM) of a developing blastocyst embryo and have the capacity to differentiate into many cell types within the body.[4]
plasma etched tissue culture polystyrene (PE-TCPS) is known to be supportive for Human embryonic stem cells (hESC) culture whereas TCPS is not; the proteins adsorbed to each surface were identified by gel electrophoresis followed by liquid chromatography mass spectrometry (LC-MS) of the main bands
We focused on proteins unique to the PE-TCPS surface as we hypothesised that these may be responsible for the ability of the PE-TCPS surface to be able to support long term hESC culture using mouse embryonic fibroblast (MEF) conditioned media
Summary
Human embryonic stem cells (hESC) are pluripotent cells derived from the inner cell mass (ICM) of a developing blastocyst embryo and have the capacity to differentiate into many cell types within the body.[4]. This includes the removal of biologically-derived components often used for hESC culture, such as mouse embryonic fibroblast conditioned medium (MEF-CM)[5] or undefined surfaces, such aLaboratory of Biophysics and Surface Analysis, School of Pharmacy, University of Nottingham, Nottingham, NG7 2RD, UK. Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, MA, 02139, USA † Electronic supplementary information (ESI) available. Proteins have been identified within MEF-CM that maintain hESC pluripotency and are involved in cell signalling, cell–cell interaction, and cell adhesion.[11,12,13]
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