TGFβ inhibition during expansion phase increases the chondrogenic re-differentiation capacity of human articular chondrocytes

• Published in: Osteoarthritis and cartilage Vol. 20; no. 10; pp. 1152 - 1160
• Main Authors: Narcisi, R, Signorile, L, Verhaar, J.A.N, Giannoni, P, van Osch, G.J.V.M
• Format: Journal Article
• Language: English
• Published: England Elsevier Ltd 01.10.2012
• Subjects: Antibodies, Blocking - pharmacology; Benzamides - pharmacology; Biomarkers - metabolism; Cartilage, Articular - cytology; Cartilage, Articular - drug effects; Cartilage, Articular - metabolism; Cell Culture Techniques; Cell Differentiation - drug effects; Cell Enlargement - drug effects; Cell Proliferation - drug effects; Cells, Cultured; Chondrocyte; Chondrocytes - cytology; Chondrocytes - drug effects; Chondrocytes - metabolism; Chondrogenesis; Culture Media - chemistry; Culture Media, Serum-Free; Expansion culture; Humans; Hypertrophy; Hypertrophy - chemically induced; Knee Injuries - metabolism; Knee Injuries - pathology; Knee Injuries - surgery; Osteoarthritis, Knee - metabolism; Osteoarthritis, Knee - pathology; Osteoarthritis, Knee - surgery; Proliferation; Pyrazoles - pharmacology; Pyrimidines - pharmacology; Rheumatology; Smad Proteins - antagonists & inhibitors; Smad Proteins - metabolism; TGFβ; Transforming Growth Factor beta - antagonists & inhibitors; Transforming Growth Factor beta - immunology; Transforming Growth Factor beta - metabolism; Expansion culture; TGFβ; Proliferation; Chondrocyte; Chondrogenesis; Hypertrophy
• ISSN: 1063-4584; 1522-9653
• DOI: 10.1016/j.joca.2012.06.010

Summary 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. Design 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. Conclusion 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.