S8022

AC16 human cardiomyocyte cell lines were purchased from EMD Millipore (cod. SCC109). Following the manufacturer's instructions, the cell line was tested and authenticated for mycoplasma contamination, which was negative. Cells were cultured in Dulbecco's Modified Eagle's Medium (DMEM)/F12 (cod. AL215A, Microgem) containing 12.5% fetal bovine serum (FBS) (cod. ECS0180L, Euroclone), 1% antibiotics penicillin-streptomycin (cod. ECB3001D, Euroclone), and 1% of L-glutamine (cod. ECB3000D, Euroclone). The cell line was maintained in the incubator at 37 °C and 5% CO2. The cells were grown between 5 and 7 passages, and experiments were performed in triplicate. AC16 were exposed to 33 mmol/L D glucose (cod. G8644, EMD Millipore) for 7 days and treated with EMPA at a concentration of 0.5 µM (cod. S8022, BI 10773, Selleckchem) [28 (link)]. The medium was changed every 48 h. Normal glucose (NG), considered the control, are cells exposed to normal glucose concentration (5.5 mmol/L) and cultured for 7 days.

AC16 human cardiomyocyte cell lines were purchased from EMD Millipore (cod. SCC109). Following the manufacturer’s instructions, the cell line was tested and authenticated for mycoplasma contamination, which was negative. Cells were cultured in Dulbecco’s Modified Eagle's Medium (DMEM)/F12 (cod. AL215A, Microgem) containing 12.5% fetal bovine serum (FBS) (cod. ECS0180L, Euroclone), 1% antibiotics penicillin–streptomycin (cod. ECB3001D, Euroclone), and 1% of l-glutamine (cod. ECB3000D, Euroclone). The cell line was maintained in the incubator at 37 °C and 5% CO₂. The cells were grown between 4 and 6 passages, and experiments were performed six times. AC16 were exposed to 33 mmol/L D-glucose (cod. G8644, EMD Millipore) for 2 days and treated with EMPA at a concentration of 0.5 µM (cod. S8022, BI 10773, Selleckchem) according to a previous protocol [25 (link), 26 (link)]. Normal glucose (NG), considered the control, are cells exposed to normal glucose concentration (5.5 mmol/L) and cultured for 2 days.