Novolin n

ASC spheroids were produced using micro-molded non-adhesive hydrogel (2% agarose in NaCl 0.9%) with 800 μm diameter in each of the 81 circular recesses (3D Petri Dish®, MicroTissues Inc) following the manufacturer’s protocol. Approximately 2 × 10⁶ cells were seeded into the chamber of the non-adhesive hydrogel resuspended in a medium composed of DMEM supplemented with 50 ug/mL ascorbic acid (Sigma Aldrich, USA), 1.25 ug/mL human albumin (Farma Biagini SPA, Brazil), 100 ug/ml penicillin and 100 ug/ml streptomycin, and ITS 1X (Lonza). For adipogenic induction, ASC spheroids were maintained in DMEM supplemented with 50 ug/mL ascorbic acid (Sigma Aldrich, USA), 1.25 ug/mL human albumin (Farma Biagini SPA), 100 U/ml penicillin and 100 ug-ml streptomycin, and ITS 1X (Lonza, USA), and the components of adipogenic induction were 10 μM insulin (Novolin® N, Novo Nordisk, Brazil) 0.5 mM isobutylmethylxanthine, 1 μM dexamethasone, and 200 μM indomethacin (Sigma Aldrich, USA). At week 1 of adipogenic induction, the supplementation of the medium was changed, and only 10 μM insulin (Novolin® N, Novo Nordisk, Brazil) was maintained as a component of adipogenic induction for up to 5 weeks. A part of ASC spheroids induced for adipogenesis was maintained under 0.5 ug/mL lipopolysaccharide (LPS) (Sigma-Aldrich, USA).