title: Stimulation of Activin A/Nodal signaling is insufficient to induce definitive endoderm formation of cord blood-derived unrestricted somatic stem cells.
abstract: BACKGROUND
  Unrestricted somatic stem cells (USSC) derived from umbilical cord blood are an attractive alternative to human embryonic stem cells (hESC) for cellular therapy. USSC are capable of forming cells representative of all three germ line layers. The aim of this study was to determine the potential of USSC to form definitive endoderm following induction with Activin A, a protein known to specify definitive endoderm formation of hESC.
  
  
  METHODS
  USSC were cultured for (1) three days with or without 100 ng/ml Activin A in either serum-free, low-serum or serum-containing media, (2) three days with or without 100 ng/ml Activin A in combination with 10 ng/ml FGF4 in pre-induction medium, or (3) four days with or without small molecules Induce Definitive Endoderm (IDE1, 100 nM; IDE2, 200 nM) in serum-free media. Formation of definitive endoderm was assessed using RT-PCR for gene markers of endoderm (Sox17, FOXA2 and TTF1) and lung epithelium (surfactant protein C; SPC) and cystic fibrosis transmembrane conductance regulator; CFTR). The differentiation capacity of Activin A treated USSC was also assessed.
  
  
  RESULTS
  Activin A or IDE1/2 induced formation of Sox17+ definitive endoderm from hESC but not from USSC. Activin A treated USSC retained their capacity to form cells of the ectoderm (nerve), mesoderm (bone) and endoderm (lung). Activin A in combination with FGF4 did not induce formation of Sox17+ definitive endoderm from USSC. USSC express both Activin A receptor subunits at the mRNA and protein level, indicating that these cells are capable of binding Activin A.
  
  
  CONCLUSIONS
  Stimulation of the Nodal signaling pathway with Activin A or IDE1/2 is insufficient to induce definitive endoderm formation from USSC, indicating that USSC differ in their stem cell potential from hESC.
authors.affiliation: Early Development and Disease, Murdoch Childrens' Research Institute, Royal Children's Hospital, Flemington Rd, Parkville, VIC 3052, Australia. filby@wehi.EDU.AU
authors.forename: Caitlin E
authors.initials: CE
authors.lastname: Filby
authors.forename: Robert
authors.initials: R
authors.lastname: Williamson
authors.forename: Peter
authors.initials: P
authors.lastname: van Kooy
authors.forename: Alice
authors.initials: A
authors.lastname: Pébay
authors.forename: Mirella
authors.initials: M
authors.lastname: Dottori
authors.forename: Ngaire J
authors.initials: NJ
authors.lastname: Elwood
authors.forename: Faten
authors.initials: F
authors.lastname: Zaibak
mesh_terms: D029404:Activin Receptors / Q000235:genetics / Q000378:metabolism; D028341:Activins / Q000494:pharmacology*; D002454:Cell Differentiation / Q000187:drug effects; D016895:Culture Media, Serum-Free; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D004707:Endoderm / Q000378:metabolism*; D005312:Fetal Blood / Q000166:cytology*; D051519:Fibroblast Growth Factor 4 / Q000494:pharmacology; D051542:Hepatocyte Nuclear Factor 3-beta / Q000235:genetics / Q000378:metabolism; D006801:Humans; D055457:Nodal Protein / Q000378:metabolism*; D055760:SOXF Transcription Factors / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D013234:Stem Cells / Q000166:cytology* / Q000187:drug effects; D014157:Transcription Factors
chemical_list: D016895:Culture Media, Serum-Free; D004268:DNA-Binding Proteins; C496502:FOXA2 protein, human; D051519:Fibroblast Growth Factor 4; C526007:NODAL protein, human; D055457:Nodal Protein; C109188:SOX17 protein, human; D055760:SOXF Transcription Factors; C504003:TTF1 protein, human; D014157:Transcription Factors; C422405:activin A; D028341:Activins; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D051542:Hepatocyte Nuclear Factor 3-beta; D029404:Activin Receptors