**2.2 3D culture of ADSC and hACh in RAD16-I composites scaffolds**

ADSC and hACh 3D cultures were maintained under control or chondrogenic conditions. Control medium was prepared with DMEM High Glucose, GlutaMAX (61965, Gibco), ITS + Premix 100x (354352, BDBioscience), 100 U/mL Penicillin/100 μg/mL Streptomycin (P11-010, PAA), 40 μg/mL l-Proline (P5607, Sigma) and 1 mM Sodium Pyruvate (11360, Life Technologies). Cultures for chondrogenic differentiation were induced at day 2 with chondrogenic medium (control medium supplemented with 10 ng/mL TGFβ1 (GF111, Millipore), 25 μg/mL l-ascorbic acid 2-phosphate (A8960; Sigma) and 100 nM Dexamethasone (D8893; Sigma)). Chondrocytes were also cultured in expansion medium (see Section 2.1). 3D cell cultures were maintained in the incubator at 37°C and 5% CO2, and medium was changed every other day. Cultures were maintained for 4 weeks in the described serum-free media under control or chondrogenic conditions (in the presence of stimulating factors to induce chondrogenic differentiation) [33, 34]. After 4 weeks, 3D constructs were analyzed for morphology, gene and protein expression, glycosaminoglycans production and mechanical properties.
