title: Biological impact of xeno-free chemically defined cryopreservation medium on amniotic epithelial cells.
abstract: BACKGROUND
  Amnion-derived stem cells have been proposed for cell replacement therapy and tissue regeneration. An easily accessible cell source, the placenta, allows us to potentially establish a bio-bank of cells for immunotype matched clinical applications. Several xeno-free (XF) cryopreservation media are currently available for pluripotent stem cells, however, these media have not yet been evaluated for the cryopreservation of amnion-derived stem cells.
  
  
  METHODS
  Human amniotic epithelial cells were collected using standard protocols, and stored at -160 °C in one of five commercially available media. Cells frozen in standard media containing fetal bovine serum served as controls. Cells were then thawed, and evaluated for viability, mitochondrial membrane stability, and senescence status. Quantitative real time PCR was utilized to assess for expression of stem cell genes, and flow cytometry was used to identify the stem cell surface markers.
  
  
  RESULTS
  Cell recovery and repopulation assays indicated no significant difference between XF media versus standard cryopreservation medium. In addition, no impact was observed on the senescence status, the cytostructural or mitochondrial morphology between the tested cryopreservation media. Differences were observed on the expression of stem cell marker genes (OCT4, SOX2, and NANOG) and a cell surface marker (TRA1-60) following cryopreservation in different chemically defined XF media, however, these were not statistically significant.
  
  
  CONCLUSIONS
  Xeno-free cryopreservation of human amnion-derived stem cells is feasible and can be standardized to establish a bio-bank with human amnion-derived stem cells for future clinical application. Optimization of this media may allow for improved preservation of stem cell-like characteristics.
authors.affiliation: Department of Biochemistry and Molecular Biology, Keck School of Medicine, University of Southern California, Los Angeles, CA, USA. toshiomi@usc.edu.|Eli and Edythe Broad Center for Regenerative Medicine and Stem Cell Research at USC, Keck School of Medicine, University of Southern California, Los Angeles, CA, USA. toshiomi@usc.edu.
authors.forename: Toshio
authors.initials: T
authors.lastname: Miki
authors.affiliation: Eli and Edythe Broad Center for Regenerative Medicine and Stem Cell Research at USC, Keck School of Medicine, University of Southern California, Los Angeles, CA, USA. wisiaw@hotmail.com.
authors.forename: Wisia
authors.initials: W
authors.lastname: Wong
authors.affiliation: Eli and Edythe Broad Center for Regenerative Medicine and Stem Cell Research at USC, Keck School of Medicine, University of Southern California, Los Angeles, CA, USA. ezhou3.14@gmail.com.
authors.forename: Elton
authors.initials: E
authors.lastname: Zhou
authors.affiliation: Eli and Edythe Broad Center for Regenerative Medicine and Stem Cell Research at USC, Keck School of Medicine, University of Southern California, Los Angeles, CA, USA. anthonyg369@yahoo.com.
authors.forename: Anthony
authors.initials: A
authors.lastname: Gonzalez
authors.affiliation: Eli and Edythe Broad Center for Regenerative Medicine and Stem Cell Research at USC, Keck School of Medicine, University of Southern California, Los Angeles, CA, USA. imgarcia@usc.edu.
authors.forename: Irving
authors.initials: I
authors.lastname: Garcia
authors.affiliation: Department of Obstetrics and Gynecology, Keck School of Medicine, University of Southern California, Los Angeles, CA, USA. Brendan.Grubbs@med.usc.edu.
authors.forename: Brendan H
authors.initials: BH
authors.lastname: Grubbs
mesh_terms: D000650:Amnion; D015415:Biomarkers / Q000378:metabolism; D002462:Cell Membrane / Q000187:drug effects / Q000648:ultrastructure; D002470:Cell Survival; D016922:Cellular Senescence; D015925:Cryopreservation; D003451:Cryoprotective Agents / Q000494:pharmacology*; D004847:Epithelial Cells / Q000187:drug effects / Q000502:physiology*; D006801:Humans; D008247:Lysosomes / Q000201:enzymology; D008928:Mitochondria / Q000187:drug effects / Q000648:ultrastructure; D039904:Pluripotent Stem Cells / Q000187:drug effects / Q000502:physiology; D001616:beta-Galactosidase / Q000378:metabolism
chemical_list: D015415:Biomarkers; D003451:Cryoprotective Agents; D001616:beta-Galactosidase