Mito stress kits

Embryonic ventricular rat heart derived H9c2 cardiomyoblasts (CRL-1446) were from American Type Culture Collection (Manassas, USA); Dulbecco's Modified Eagle's Medium (DMEM), Penicillin-Streptomycin, Phosphate Buffered Saline (DPBS), and fetal bovine serum (FBS) were from Carlo Erba (Milano, Italy); Bradford and RC DC protein assay kit was from Bio-Rad Laboratories (Hercules, USA); 2', 7'-dichlorfluorescein diacetate (DCFH-DA) and MitoSOX fluorescent dyes were from ThermoFisher Scientific (Waltham, USA); ViaCount was from Merck-Millipore (Darmstadt, Germany). Seahorse XF96 microplate plates, Seahorse XF Assay media, Seahorse XF base media without phenol red and Mito Stress Kits were from Agilent (Santa Clare, USA). All other chemicals including palmitate and 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide) (MTT) were obtained from Sigma-Aldrich (St. Louis, USA).

Metabolic flux analyses were performed using the Agilent glycolysis stress and mito-stress kits 21 (Supplementary Methods). Metabolomics was performed on primary human MDMs differentiated in the presence and absence of glucose-6-phosphate transporter inhibitor S3483 21 . Quantification of phosphorylation of S6 ribosomal protein (pS6) was performed by intracellular staining and FACS (Supplementary Methods).