Optimization of chondrocyte expansion in culture. Effect of TGF beta-2, bFGF and L-ascorbic acid on bovine articular chondrocytes.

In vitro multiplication of isolated chondrocytes is needed to repair articular cartilage defects with autologous material. In this study we used monolayer cultures of bovine articular chondrocytes. The effect of transforming growth factor beta-2, basic fibroblast growth factor or L-ascorbic acid on cell multiplication, in the presence of 10% fetal calf serum, was measured in primary culture, the third and tenth passage. TGF beta-2 stimulated the proliferation of chondrocytes in the primary culture and L-ascorbic acid stimulated in the third passage. On the basis of these results, we chose an optimal addition scheme in which TGF beta-2 was added in primary culture and first passage, followed by addition of L-ascorbic acid in the second and third passage; this resulted in a 7-fold increase in cell number compared to the control group, in about 4 weeks. Our findings stress the importance of adding the right growth factor at the right moment. Collagen type II expression was lost after the third passage, in the control as well as in the experimental condition. The ability to produce hyaline cartilage specific matrix components is essential, if multiplied cells are to be used to repair cartilage defects.