Tissue Engineering of Articular Cartilage under the Influence of Collagen I/III Membranes and Low Oxygen Tension

Abstract

The objective of this study was to study the matrix production and phenotype stability of articular chondrocytes cultured on collagen I/III membranes (CM) under the influence of low oxygen tension (PO₂). Primary bovine and osteoarthritic human chondrocytes were cultured for 2 weeks under 5–21% PO₂ on CM, in alginate, or as monolayers. Dedifferentiated cells were produced by 2-week monolayer culture under 21% PO₂. Collagen (Coll) type II and I expression was demonstrated immunohistochemically, by Western blotting (Coll II), and by semiquantitative RT-PCR; proteoglycan synthesis was demonstrated histochemically (toluidine blue); and biosynthetic activity was indicated by radiolabel incorporation ([³H]proline and [³⁵S]sulfate). Bovine chondrocytes on CM showed an increase in Coll II expression and proteoglycan synthesis under low PO₂ conditions, whereas Coll I decreased. This oxygen-dependent phenotype-stabilizing effect was even more pronounced in alginate cultures. Biosynthesis of bovine and human chondrocytes was also increased by low PO₂, except for proline incorporation, which decreased in bovine CM cultures (low-oxygen effects were significantly higher in alginate than in CM cultures). Dedifferentiated chondrocytes reexpressed Coll II protein when cultured under low PO₂ on CM or in alginate only, but not under high PO₂ or in monolayer culture. We conclude that CM and, even more, alginate foster phenotype stability and cartilage-specific matrix production of bovine chondrocytes, especially when cultured under in vivo-like oxygen conditions.