title: Lowering oxygen tension enhances the differentiation of mouse embryonic stem cells into neuronal cells.
abstract: Embryonic stem cells (ESC) are capable of proliferating indefinitely in vitro whilst retaining their ability to differentiate into cells of every adult lineage. Efficient, high yield processes, which direct differentiation of ESC to specific lineages, will underpin the development of cost-effective drug screening and cell therapy products. The aim of this study was to investigate whether laboratory oxygen tension currently used for the neuronal differentiation of ESC was suboptimal resulting in inefficient process yields. An adherent monolayer protocol for the neuronal differentiation of mouse ESC (mESC) was performed in oxygen controlled chambers using a chemically defined media over an 8 day period of culture. When exposed to oxygen tensions more appropriate to in vivo neuronal development (2% O(2)), there was a 34-fold increase in the yield of viable cells from the differentiation process. Low oxygen tension inhibited cell death during an early phase (48 to 96 h) and toward the end (120 to 192 h) of the process. The percentage of cells expressing neuronal markers was determined by flow cytometry, revealing a small rise in the betaIII tubulin and a threefold increase in the MAP2 populations at 2% O(2). The total increase in the yield of viable cells expressing neuronal markers was shown to be 55-fold for betaIII tubulin and 114-fold for MAP2. In conclusion, this study revealed that low oxygen tension can be used to enhance the yield of neuronal cells derived from ESCs and has implications for the development of efficient, cost-effective production processes.
authors.affiliation: The Advanced Centre for Biochemical Engineering, Dept. of Biochemical Engineering, University College London, UK.
authors.forename: Paul
authors.initials: P
authors.lastname: Mondragon-Teran
authors.forename: Gary J
authors.initials: GJ
authors.lastname: Lye
authors.forename: Farlan S
authors.initials: FS
authors.lastname: Veraitch
mesh_terms: D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods; D002470:Cell Survival; D053595:Embryonic Stem Cells / Q000166:cytology / Q000378:metabolism / Q000502:physiology*; D005434:Flow Cytometry; D007150:Immunohistochemistry; D007381:Intermediate Filament Proteins / Q000378:metabolism; D051379:Mice; D008858:Microscopy, Phase-Contrast; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000378:metabolism; D064231:Nestin; D055495:Neurogenesis / Q000502:physiology*; D009474:Neurons / Q000166:cytology / Q000378:metabolism*; D010100:Oxygen / Q000737:chemistry*; D012397:Rosette Formation; D014404:Tubulin / Q000378:metabolism
chemical_list: D007381:Intermediate Filament Proteins; D008869:Microtubule-Associated Proteins; C079240:Mtap2 protein, mouse; D009419:Nerve Tissue Proteins; C577485:Nes protein, mouse; D064231:Nestin; D014404:Tubulin; D010100:Oxygen