Stem Cell Reviews and Reports

, Volume 5, Issue 4, pp 353–368 | Cite as

Isolation and Differentiation of Chondrocytic Cells Derived from Human Embryonic Stem Cells Using dlk1/FA1 as a Novel Surface Marker

  • Linda Harkness
  • Hanna Taipaleenmaki
  • Amer Mahmood
  • Ulrik Frandsen
  • Anna-Marja Saamanen
  • Moustapha Kassem
  • Basem M. Abdallah
Article

Abstract

Few surface markers are available to monitor lineage differentiation during chondrogenesis. Recently, delta-like1/fetal antigen1 (dlk1/FA1), a transmembrane protein of the Notch/Delta/Serrata family, was shown to be essential for inducing early chondrogenesis. Thus, we investigated the possible use of dlk1/FA1 as a novel surface marker for chondroprogenitor cells during hESC differentiation. We found that, Dlk1/FA1 is expressed specifically in cells undergoing transition from proliferating to prehypertrophic chondrocytes during endochondral ossification of the mouse limb. In hESC cells, dlk1/FA1 was not expressed by undifferentiated hESC, but expressed during in vitro embryoid bodies (hEBs) formation upon down-regulation of undifferentiated markers e.g. Oct 3/4. Similarly, dlk1/FA1 was expressed in chondrocytic cells during in vivo teratoma formation. Interestingly, treatment of hEBs with Activin B, a member of TGF-ß family, markedly increased Dlk1 expression in association with up-regulation of the mesoderm-specific markers (e.g. FOXF1, KDR and VE-cadherin) and SOX9. dlk1/FA1+ cells isolated by fluorescence activated cell sorting (FACS) were capable of differentiating into chondrocytic cells when cultured as micromass pellets in a xeno-free system containing TGFβ1. In conclusion, we identified dlk1/FA1 as a novel marker of chondroprogenitor cells that undergo embryonic lineage progression from proliferation to the prehypertrophic stage. Tracking dlk1/FA1 expression as a mesoderm/chondroprogenitor surface marker provides a novel strategy for designing clinically relevant protocols to direct the differentiation of hESC into chondrocytes.

Keywords

Dlk1 FA1 Pref-1 Chondrogenesis hESC Bone development Surface marker 

Supplementary material

12015_2009_9099_Fig7_ESM.jpg (515 kb)
Figure S1

Immunolocalization of dlk1/FA1 in developing articular cartilage. Immunolocalization of dlk1 (FA1) is compared to those of prechondrogenic type IIA procollagen (COLIIA), and type X collagen (COLX) in adjacent sagital paraffin sections from knee femoral epiphyses of 10-day-old (top panels) and 20-day-old (bottom panels) mice. Secondary HRP-linked antibody was detected by DAB staining (FA1 and COLIIA) or AP-linked antibody by Fast Red (COLX) and sections were counterstained with haematoxylin. Bar equals to 100 μm. (JPEG 514 kb)

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Copyright information

© Humana Press 2009

Authors and Affiliations

  • Linda Harkness
    • 1
  • Hanna Taipaleenmaki
    • 2
  • Amer Mahmood
    • 1
  • Ulrik Frandsen
    • 1
  • Anna-Marja Saamanen
    • 2
  • Moustapha Kassem
    • 1
    • 3
  • Basem M. Abdallah
    • 1
  1. 1.Molecular Endocrinology laboratory (KMEB), Medical Biotechnology Centre, Odense University HospitalUniversity of South DenmarkOdense CDenmark
  2. 2.Department of Medical Biochemistry and Molecular BiologyUniversity of TurkuTurkuFinland
  3. 3.Stem Cell Unit, Department of AnatomyKing Saud UniveristyRiyadhSaudi Arabia

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