title: Discrete adipose-derived stem cell subpopulations may display differential functionality after in vitro expansion despite convergence to a common phenotype distribution.
abstract: Complex immunophenotypic repertoires defining discrete adipose-derived stem cell (ASC) subpopulations may hold a key toward identifying predictors of clinical utility. To this end, we sorted out of the freshly established ASCs four subpopulations (SPs) according to a specific pattern of co-expression of six surface markers, the CD34, CD73, CD90, CD105, CD146, and CD271, using polychromatic flow cytometry.
  
  
  
  Using flow cytometry-associated cell sorting and analysis, gating parameters were set to select for a CD73+CD90+CD105+phenotype plus one of the four following combinations, CD34-CD146-CD271-(SP1), CD34-CD146+CD271-(SP2), CD34+CD146+CD271-(SP3), and CD34-CD146+CD271+(SP4). The SPs were expanded 700- to 1000-fold, and their surface repertoire, trilineage differentiation, and clonogenic potential, and the capacity to support wound healing were assayed.
  
  
  
  Upon culturing, the co-expression of major epitopes, the CD73, CD90, and CD105 was maintained, while regarding the minor markers, all SPs reverted to resemble the pre-sorted population with CD34-CD146-CD271-and CD34-CD146+CD271-representing the most prevalent combinations, followed by less frequent CD34+CD146-CD271-and CD34+CD146+CD271-variants. There was no difference in the efficiency of adipo-, osteo-, or chondrogenesis by cytochemistry and real-time RT-PCR or the CFU capacity between the individual SPs, however, the SP2CD73+90+105+34-146+271-outperformed others in terms of wound healing.
  
  
  
  Our study shows that ASCs upon culturing inherently maintain a stable distribution of immunophenotype variants, which may potentially disguise specific functional properties of particular downstream lines. Furthermore, the outlined approach suggests a paradigm whereby discrete subpopulations could be identified to provide for a therapeutically most relevant cell product.
authors.affiliation: Laboratory for Stem Cell Research, Department for Health Science and Technology, Aalborg University, Aalborg, Denmark.
authors.forename: Frederik Mølgaard
authors.initials: FM
authors.lastname: Nielsen
authors.affiliation: Laboratory for Stem Cell Research, Department for Health Science and Technology, Aalborg University, Aalborg, Denmark.
authors.forename: Simone Elkjær
authors.initials: SE
authors.lastname: Riis
authors.affiliation: Laboratory for Stem Cell Research, Department for Health Science and Technology, Aalborg University, Aalborg, Denmark.
authors.forename: Jens Isak
authors.initials: JI
authors.lastname: Andersen
authors.affiliation: Department of Bioengineering, Polytech Nice-Sophia Engineering School, Nice, France.
authors.forename: Raphaëlle
authors.initials: R
authors.lastname: Lesage
authors.affiliation: Laboratory for Stem Cell Research, Department for Health Science and Technology, Aalborg University, Aalborg, Denmark.
authors.forename: Trine
authors.initials: T
authors.lastname: Fink
authors.affiliation: Laboratory for Stem Cell Research, Department for Health Science and Technology, Aalborg University, Aalborg, Denmark.
authors.forename: Cristian Pablo
authors.initials: CP
authors.lastname: Pennisi
authors.affiliation: Laboratory for Stem Cell Research, Department for Health Science and Technology, Aalborg University, Aalborg, Denmark. vlaz@hst.aau.dk.
authors.forename: Vladimir
authors.initials: V
authors.lastname: Zachar
mesh_terms: D017667:Adipocytes / Q000166:cytology* / Q000378:metabolism; D000328:Adult; D015703:Antigens, CD / Q000235:genetics / Q000378:metabolism; D001681:Biological Assay; D002454:Cell Differentiation; D019070:Cell Lineage / Q000235:genetics; D019902:Chondrocytes / Q000166:cytology* / Q000378:metabolism; D005434:Flow Cytometry; D015870:Gene Expression; D006801:Humans; D016130:Immunophenotyping; D008297:Male; D010006:Osteoblasts / Q000166:cytology* / Q000378:metabolism; D010641:Phenotype; D061251:Primary Cell Culture; D013234:Stem Cells / Q000145:classification* / Q000166:cytology* / Q000378:metabolism; D050507:Subcutaneous Fat, Abdominal / Q000166:cytology / Q000378:metabolism
chemical_list: D015703:Antigens, CD
keywords: Adipose-derived stem cells; Antigenic repertoire; FACS; Phenotype; Polychromatic cell sorting; Scratch assay; Subpopulations