title: Low oxygen atmosphere facilitates proliferation and maintains undifferentiated state of umbilical cord mesenchymal stem cells in an hypoxia inducible factor-dependent manner.
abstract: OBJECTIVE
  As we approach the era of mesenchymal stem cell (MSC) application in the medical clinic, the standarization of their culture conditions are of the particular importance. We re-evaluated the influences of oxygens concentration on proliferation, stemness and differentiation of human umbilical cord Wharton Jelly-derived MSCs (WJ-MSCs).
  
  
  METHODS
  Primary cultures growing in 21% oxygen were either transferred into 5% O2 or continued to grow under standard 21% oxygen conditions. Cell expansion was estimated by WST1/enzyme-linked immunosorbent assay or cell counting. After 2 or 4 weeks of culture, cell phenotypes were evaluated using microscopic, immunocytochemical, fluorescence-activated cell-sorting and molecular methods. Genes and proteins typical of mesenchymal cells, committed neural cells or more primitive stem/progenitors (Oct4A, Nanog, Rex1, Sox2) and hypoxia inducible factor (HIF)-1α-3α were evaluated.
  
  
  RESULTS
  Lowering O2 concentration from 21% to the physiologically relevant 5% level substantially affected cell characteristics, with induction of stemness-related-transcription-factor and stimulation of cell proliferative capacity, with increased colony-forming unit fibroblasts (CFU-F) centers exerting OCT4A, NANOG and HIF-1α and HIF-2α immunoreactivity. Moreover, the spontaneous and time-dependent ability of WJ-MSCs to differentiate into neural lineage under 21% O2 culture was blocked in the reduced oxygen condition. Importantly, treatment with trichostatin A (TSA, a histone deacetylase inhibitor) suppressed HIF-1α and HIF-2α expression, in addition to blockading the cellular effects of reduced oxygen concentration.
  
  
  CONCLUSIONS
  A physiologically relevant microenvironment of 5% O2 rejuvenates WJ-MSC culture toward less-differentiated, more primitive and faster-growing phenotypes with involvement of HIF-1α and HIF-2α-mediated and TSA-sensitive chromatin modification mechanisms. These observations add to the understanding of MSC responses to defined culture conditions, which is the most critical issue for adult stem cells translational applications.
authors.affiliation: NeuroRepair Department, Mossakowski Medical Research Centre, Polish Academy of Sciences, Warsaw, Poland.
authors.forename: Katarzyna
authors.initials: K
authors.lastname: Drela
authors.affiliation: NeuroRepair Department, Mossakowski Medical Research Centre, Polish Academy of Sciences, Warsaw, Poland.
authors.forename: Anna
authors.initials: A
authors.lastname: Sarnowska
authors.affiliation: NeuroRepair Department, Mossakowski Medical Research Centre, Polish Academy of Sciences, Warsaw, Poland.
authors.forename: Patrycja
authors.initials: P
authors.lastname: Siedlecka
authors.affiliation: NeuroRepair Department, Mossakowski Medical Research Centre, Polish Academy of Sciences, Warsaw, Poland.
authors.forename: Ilona
authors.initials: I
authors.lastname: Szablowska-Gadomska
authors.affiliation: First Department of Obstetrics and Gynecology, Medical University of Warsaw, Warsaw, Poland.
authors.forename: Miroslaw
authors.initials: M
authors.lastname: Wielgos
authors.affiliation: Cryo-Save Labs NV (The Cell Factory), Niel, Belgium.
authors.forename: Marcin
authors.initials: M
authors.lastname: Jurga
authors.affiliation: NeuroRepair Department, Mossakowski Medical Research Centre, Polish Academy of Sciences, Warsaw, Poland.
authors.forename: Barbara
authors.initials: B
authors.lastname: Lukomska
authors.affiliation: NeuroRepair Department, Mossakowski Medical Research Centre, Polish Academy of Sciences, Warsaw, Poland. Electronic address: krystyna.dj@gmail.com.
authors.forename: Krystyna
authors.initials: K
authors.lastname: Domanska-Janik
mesh_terms: D018929:Cell Culture Techniques; D049109:Cell Proliferation / Q000187:drug effects*; D036101:Cord Blood Stem Cell Transplantation / Q000379:methods; D006801:Humans; D006877:Hydroxamic Acids / Q000008:administration & dosage; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000378:metabolism*; D059630:Mesenchymal Stem Cells / Q000187:drug effects / Q000378:metabolism; D010100:Oxygen / Q000378:metabolism*; D044763:Ubiquitin-Conjugating Enzymes / Q000378:metabolism*; D014470:Umbilical Cord / Q000166:cytology*
chemical_list: D006877:Hydroxamic Acids; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; C012589:trichostatin A; C100879:UBE2K protein, human; D044763:Ubiquitin-Conjugating Enzymes; D010100:Oxygen
keywords: SRTF/HIF gene profiles; TSA effect; WJ MSC culture; cell development; differentiation; influence of oxygen concentration; proliferation