Isolation and Expansion of Osteoarthritic Chondrocytes

Cartilage tissues were obtained from four patients with end-stage OA after total knee arthroplasty (ages 71, 72, 64, and 82 years; all female) at the University Hospital of Basel under ethical agreement (Ethikkommission beider Basel, Ref.Nr. EK: 78/07). The cells were isolated as described previously elsewhere [58 (link)]. In brief, after cutting the tissues with a scalpel into small pieces, they were digested overnight in 0.2% collagenase II (300 U/mg, Worthington Biochemical Corp, Lakewood, NJ, USA) on an orbital shaker at 37 °C. The isolated chondrocytes were expanded for three passages to 80% confluency in basal medium (BM, Dulbecco’s modified Eagle medium, high glucose (DMEM)), 1 mM sodium pyruvate, 10 mM HEPES, 1% penicillin/streptomycin (P/S), and 0.29 mg/mL glutamate (all from Gibco), supplemented with 10% fetal bovine serum (FBS), 1 ng/mL transforming growth factor (TGF)-β1, and 5 ng/mL fibroblast growth factor-2 (FGF-2) (both from Fitzgerald, Acton, MO, USA) in a humidified incubator (37 °C, 5% CO2).