title: TGFβ inhibition during expansion phase increases the chondrogenic re-differentiation capacity of human articular chondrocytes.
abstract: OBJECTIVE
  Autologous chondrocyte implantation is a cell-based treatment to repair articular cartilage defects, relying on the availability of expanded (de-differentiated) chondrocytes. Unfortunately, the expansion process causes several phenotypical changes, requiring re-establishment of the native chondrogenic phenotype to sustain proper repair. Among other proteins, transforming growth factor-β (TGFβ) is known to influence the chondrogenic re-differentiation of human articular chondrocytes (HACs) and their matrix deposition. Thus we investigated the effects of TGFβ-depletion during the expansion phase.
  
  
  METHODS
  HACs were isolated from articular cartilage and expanded in the canonical serum-supplemented medium [fetal calf serum (FCS)] or in a chemically-defined (CD) medium, with or without anti-TGFβ antibody administration. The re-differentiation potential of the cells was assessed by pellet cultures, gene expression analysis and histology.
  
  
  RESULTS
  Cell proliferation proceeded more rapidly in CD-medium than in FCS-medium; it was not affected by the use of anti-TGFβ antibody but was further increased by addition of exogenous TGFβ1, via increased p-Smad1/5/8. Conversely, in FCS-medium, addition of anti-TGFβ antibody decreased both proliferation and p-Smad1/5/8 level. Challenging either FCS- or CD-medium with anti-TGFβ antibody during expansion enhanced chondrogenesis in the subsequent pellet cultures. Moreover, TGFβ-depletion during expansion in CD-medium inhibited mRNA expression of hypertrophic markers, collagen type-X (COL10) and matrix metalloproteinase-13 (MMP-13). Interestingly, the TGFβ1 level detected by enzyme-linked immunosorbent sandwich assay (ELISA) during cell expansion was correlated with COL10 mRNA expression after re-differentiation.
  
  
  CONCLUSIONS
  TGFβ-depletion during expansion improves the re-differentiation capacity of chondrocytes and inhibits hypertrophy. These results indicate the importance of the expansion medium composition to improve chondrogenic re-differentiation and to inhibit hypertrophy.
authors: Department of Orthopaedics, Erasmus MC, Rotterdam, The Netherlands. r.narcisi@erasmusmc.nl
authors: R
authors: R
authors: Narcisi
authors: L
authors: L
authors: Signorile
authors: J A N
authors: JA
authors: Verhaar
authors: P
authors: P
authors: Giannoni
authors: G J V M
authors: GJ
authors: van Osch
mesh_terms: D019138:Antibodies, Blocking / Q000494:pharmacology*; D001549:Benzamides / Q000494:pharmacology; D015415:Biomarkers / Q000378:metabolism; D002358:Cartilage, Articular / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D018929:Cell Culture Techniques; D002454:Cell Differentiation / Q000187:drug effects; D049108:Cell Enlargement / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D019902:Chondrocytes / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D003470:Culture Media / Q000737:chemistry*; D016895:Culture Media, Serum-Free; D006801:Humans; D006984:Hypertrophy / Q000139:chemically induced; D007718:Knee Injuries / Q000378:metabolism / Q000473:pathology / Q000601:surgery; D020370:Osteoarthritis, Knee / Q000378:metabolism / Q000473:pathology / Q000601:surgery; D011720:Pyrazoles / Q000494:pharmacology; D011743:Pyrimidines / Q000494:pharmacology; D051785:Smad Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D016212:Transforming Growth Factor beta / Q000037:antagonists & inhibitors / Q000276:immunology / Q000378:metabolism*
chemical_list: D019138:Antibodies, Blocking; D001549:Benzamides; D015415:Biomarkers; D003470:Culture Media; D016895:Culture Media, Serum-Free; D011720:Pyrazoles; D011743:Pyrimidines; D051785:Smad Proteins; D016212:Transforming Growth Factor beta; C516138:dorsomorphin