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Growth requirements of low‐density rabbit costal chondrocyte cultures maintained in serum‐free medium

The factors required for the active proliferation of low‐density rabbit costal chondrocytes exposed to 9:1 (v/v) mixture of Dulbecco's modified Eagle's medium and Ham's F12 medium have been defined. Low‐density primary cultures of rabbit costal chondrocytes proliferated actively when the medium was supplemented with high‐density lipoprotein (300 μg/ml), transferrin (60 μg/ml), fibroblast growth factor (FGF) (1 ng/ml), hydrocortisone (10−6 M), and epidermal growth factor (EGF) (30 ng/ml). Insulin, although it slightly decreased the final cell density, was required for reexpression of the cartilage phenotype at confluence. Optimal proliferation of low‐density chondrocyte cultures was only observed when dishes were coated with an extracellular matrix (ECM) produced by cultured corneal endothelial cells, but not on plastic. Furthermore, serum‐free chondrocyte cultures seeded at low density and maintained on ECM‐coated dishes gave rise to a homogeneous cartilage‐like tissue composed of spherical cells. These chondrocytes therefore seem to provide a good experimental system for analyzing factors involved in supporting proliferation of chondrocytes and their phenotypic expression.