Construction of HEK293-SGLT2 cell lines
HEK293 cells (CLS Cat# 300192/p777_HEK293, RRID: CVCL_0045) were seeded into a 6-well plate at the density of 0.5 × 106cells/well and incubated overnight. To construct a cell line with over-expression of SGLT2, the plasmid plvx-puro-SGLT2 (containing SGLT2 gene sequence with lentivirus) were transfected into HEK293 cells using lipofectamine 3000 (Invitrogen, Cat: L3000-015, USA). After six-hour of infection, the culture medium was replaced with fresh Dulbecco’s modified Eagle’s medium (DMEM). After 48 hours, the cells were treated with puromycin (Thermo Fisher Scientific, USA, CID: 439530) at the final concentration of 2 μg/mL, and the complete culture medium (with 2 μg/mL of puromycin) was replaced every 48 hours. RT-PCR and Western Blot were performed to determine the expression level of SGLT2.