Rt112

Human bladder cancer cells T24, 5637, J82, RT112, EJ, TCCSUP, UM-UC-3, and human bladder immortalized epithelium cell line (SV-HUC-1) were purchased from Procell (Procell Life Science& Technology Co., Ltd). SV-HUC-1 cells were cultured in Ham’s F-12K medium. MEM mixed with 10% FBS (Excell) was used to cultivate the UM-UC-3 and the J82 line. RPMI1640 (Invitrogen) mixed with 10% FBS (Excell) was used to cultivate T24, 5637, RT112, EJ, TCCSUP, and those cell lines were all cultured in a water-saturated atmosphere with 5% CO2 at 37°C.

The human lung adenocarcinoma cell line Calu-3 (FGFR1-low expression) was kindly provided by the Stem Cell Bank, Chinese Academy of Sciences (China) and maintained in MEM (BI, Israel) containing 10% fetal bovine serum (FBS) (BI, Israel). RPMI 1640 medium (BI, Israel) supplemented with 10% FBS was used to culture the gastric carcinoma cell line SNU-16 (FGFR1-low expression) purchased from KeyGEN BioTECH (China), the lung adenocarcinoma cell line A549 (FGFR1-low expression) and the bladder cancer cell line RT-112 (FGFR1-high expression), purchased from Procell (China). All cell lines were identified by short tandem repeat (STR) profiling before purchase and Mycoplasma testing was negative. The cells were then cultured under standard conditions (37°C in a 5% CO₂ atmosphere). The growth rate and morphology of all cell lines were determined by inverted microscopy with phase contrast. After trypsin-EDTA solution (0.02% EDTA, 0.25% trypsin; BI, Israel) digestion of the adherent cells, the cells were collected. All experiments were performed at the first five passages of the cells.

Human ureteral epithelial cells SV-HUC-1, BC cell lines including UM-UC-1, UM-UC-3, RT4, RT112 and T24 were purchased from Procell Life Sciences (Wuhan, China). SV-HUC-1 cells were cultured in Ham’s F-12 K medium (Macgene, China) supplemented with 10% fetal bovine serum (FBS) (Gibco, MA, USA). UM-UC-1, UM-UC-3, RT4, RT112 and T24 cells were cultured in RPMI-1640 medium (Gibco, MA, USA) supplemented with 10% FBS. All cells were cultured in 5% CO₂ at 37 °C.
The si-PFK-1 and PFK-1 overexppressing plasmids and their negative control were synthesized by Gemma Shanghai. UM-UC-1 and RT112 cells were grown at 90% fusion. Transfection was performed using Lipofectamine 3000 regent according to the instructions. The culture medium was changed 6 h after transfection, and PCR was performed to detect transfection efficiency at 24–48 h after transfection.