Rat Chondrocyte Isolation and IL-1β Treatment

Knee joint tissues were collected from 6-day-old Sprague Dawley rats. Primary rat articular chondrocytes were isolated from the knee joint tissues as described previously, with minor modifications (Gosset et al., 2008 (link); Zhou et al., 2018 (link)). Briefly, bilateral knee joint cartilage was rinsed twice with PBS and then digested with trypsin at 37°C for 30 min in a thermal incubator supplied with 5% CO₂. Then the tissue fragments were incubated overnight in a 0.1–0.2% collagenase II solution at 37°C (Jiang et al., 2016 (link)). After filtering and centrifuging at 400 × g for 10 min, rat articular chondrocytes were seeded and cultured in GIBCO DMEM/F12 (1:1) medium (No.: C11330500BT), supplemented with 10% fetal bovine plasma (FBS; Gibco) and 1% penicillin and streptomycin (Sigma Aldrich). Cells grew on 60 mm culture plates at 37°C in a humidified incubator supplied with 5% CO₂. Cells were passaged when they reached 80–90% confluent. Chondrocytes within P3 generation were used in this study.
Rat chondrocytes were treated with recombinant rat IL-1β (cat. No. I2393; Sigma Aldrich; 10 ng/ml) for 12 or 24 h to establish an in vitro OA cellular model as previously described (Huang et al., 2019 (link)). Chondrocytes cultured under normal conditions without IL-1β were set as normal control (NC) group.