title: Effect of culture medium on propagation and phenotype of corneal stroma-derived stem cells.
abstract: OBJECTIVE
  The limbal area of the corneal stroma has been identified as a source of mesenchymal-like stem cells, which have potential for exploitation as a cell therapy. However, the optimal culture conditions are disputed and few direct media comparisons have been performed. In this report, we evaluated several media types to identify the optimal for inducing an in vitro stem cell phenotype.
  
  
  METHODS
  Primary human corneal stroma-derived stem cells (CSSCs) were extracted from corneoscleral rims. Culture in seven different media types was compared: Dulbecco's modified Eagle's medium (DMEM) with 10% fetal bovine serum (FBS); M199 with 20% FBS; DMEM-F12 with 20% serum replacement, basic fibroblast growth factor and leukemia inhibitory factor (SCM); endothelial growth medium (EGM); semi-solid MethoCult; serum-free keratinocyte medium (K-SFM); and StemPro-34. Effects on proliferation, morphology, protein and messenger RNA expression were evaluated.
  
  
  RESULTS
  All media supported proliferation of CSSCs with the exception of K-SFM and StemPro-34. Morphology differed between media: DMEM produced large cells, whereas EGM produced very small cells. Culture in M199 produced a typical mesenchymal stromal cell phenotype with high expression of CD105, CD90 and CD73 but not CD34. Culture in SCM produced a phenotype more reminiscent of a progenitor cell type with expression of CD34, ABCG2, SSEA-4 and PAX6.
  
  
  CONCLUSIONS
  Culture medium can significantly influence CSSC phenotype. SCM produced a cell phenotype closest to that of a pluripotent stem cell, and we consider it to be the most appropriate for development as a clinical-grade medium for the production of CSSC phenotypes suitable for cell therapy.
authors.affiliation: Academic Ophthalmology, Division of Clinical Neuroscience, Queen's Medical Centre Campus, University of Nottingham, Nottingham, United Kingdom. Electronic address: laura.sidney@nottingham.ac.uk.
authors.forename: Laura E
authors.initials: LE
authors.lastname: Sidney
authors.affiliation: Academic Ophthalmology, Division of Clinical Neuroscience, Queen's Medical Centre Campus, University of Nottingham, Nottingham, United Kingdom.
authors.forename: Matthew J
authors.initials: MJ
authors.lastname: Branch
authors.affiliation: Academic Ophthalmology, Division of Clinical Neuroscience, Queen's Medical Centre Campus, University of Nottingham, Nottingham, United Kingdom.
authors.forename: Harminder S
authors.initials: HS
authors.lastname: Dua
authors.affiliation: Academic Ophthalmology, Division of Clinical Neuroscience, Queen's Medical Centre Campus, University of Nottingham, Nottingham, United Kingdom.
authors.forename: Andrew
authors.initials: A
authors.lastname: Hopkinson
mesh_terms: D000818:Animals; D018952:Antigens, CD34 / Q000378:metabolism; D002417:Cattle; D018929:Cell Culture Techniques; D002454:Cell Differentiation / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects*; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D003319:Corneal Stroma / Q000166:cytology*; D003470:Culture Media / Q000378:metabolism / Q000494:pharmacology*; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000166:cytology*; D010641:Phenotype; D039904:Pluripotent Stem Cells / Q000166:cytology*; D055746:Stage-Specific Embryonic Antigens / Q000378:metabolism
chemical_list: D018952:Antigens, CD34; D003470:Culture Media; D055746:Stage-Specific Embryonic Antigens; C066584:stage-specific embryonic antigen-4
keywords: cornea; corneal stroma; corneal stromal stem cells; culture medium; keratocytes; mesenchymal stromal cells