Cr 12800

MSCs were planted in high-density (5 × 10⁵ cells) micromasses in a chondrogenic medium. The chondrogenic medium was composed of high-glucose DMEM (Cienry, CR-12800), 1% ITS Premix (Corning, 354351), 1% penicillin‒streptomycin, 50 μM ascorbic acid, 10 ng/ml recombinant human transforming growth factor (TGF) beta 3 (rhTGF-β3; R&D, 243-B3-010), 1 mM sodium pyruvate (Sigma‒Aldrich, P5280), and 0.1 μM dexamethasone. The cells were cultured in a 37 °C, 5% CO₂ incubator, and the medium was exchanged every 2 to 3 days. After 21 days of induction, the chondrogenic cells were processed for Alcian blue staining (Sigma‒Aldrich, 66011) according to the manufacturer’s protocol.

Approximately, 5 × 10⁵ MSCs were first centrifuged at 600 g for 5 min to form pellets. Then, the MSCs were seeded as high-density pellets (5 × 10⁵ cells) in serum-free chondrogenic medium composed of high-glucose DMEM (Cienry, CR-12800) supplemented with 1% ITS-Premix (Corning, 354351), 50 mg/L ascorbic acid, 1 mM sodium pyruvate (Sigma, P5280), 100 nM dexamethasone and 10 ng/mL recombinant human transforming growth factor (TGF)-β3 (R&D system, 243-B3-010) for 21 days. Toluidine blue staining was used to detect the chondrogenic differentiation potential.