maintenance of beige adipocytes 1

Beige adipocyte lines were established by immortalizing SVFs from the inguinal WAT B6 WT mice according to the cell immortalization protocol using SV40 large T antigen. Preadipocytes were cultured to confluence and adipocyte differentiation was induced with DMEM/F-12 medium (with phenol red, Wako) containing 10% FBS, 1% penicillin-streptomycin, 5 µg/ml insulin, 1 nM T3, 0.25 mM IBMX, 0.125 mM indomethacin, 2 µg/ml dexamethasone. Two days after induction, cells were switched to the maintenance medium containing 10% FBS, 5 µg/ml insulin, and 1 nM T3 for an additional for 4 days in the presence of rosiglitazone at a dose of 1 µM. After the induction, Rosiglitazone was continued (Rosi) or discontinued (Rosi-Off) and cells were cultured for 3 more days.