Control sirna sicontrol

A series of RCC cell lines (769-P, 786-O, A498, Caki-1, and ACHN) and an immortalized human embryonic kidney cell line - HEK293T were used for this study. Dulbecco's Modified Eagle Medium (DMEM) supplemented with 10% fetal bovine serum and 1% penicillin/streptomycin was used for cell culture. Cells were cultured in DMEM in a humidified chamber maintained at 37°C and 5% CO₂. OSR1-short interfering RNA (siOSR1) and control siRNA (siControl) were purchased from Santa Cruz Biotechnology (Dallas, TX, USA). Transfection was carried out according to the manufacturer's instruction using RNAiMAX transfection reagent (Invitrogen, Eugene, OR, USA).

Tubular epithelial HK2 cells were obtained from ATCC and cultured in Dulbecco’s modified Eagle medium with 10% foetal bovine serum and 1% penicillin‐streptomycin solution (HyClone) at 37°C and 5% CO₂. When the cell density reached 80%, the cells were passaged. For the down‐regulation of CIRBP and ELAVL1, siRNA (si‐CIRBP, SC‐97329), ELAVL1 siRNA (sc‐35619, si‐ELAVL1) and control siRNA (si‐control) were obtained from Santa Cruz Biotechnology. The CIRBP overexpression vector pCDNA3.1‐CIRBP (NM_001300829.2) was constructed by Hanbio. Cells were transfected with Lipofectamine 3000 (Invitrogen) according to the manufacturer’s instructions. In HR experiments, HK2 cells were exposed to 6/12 hours of hypoxia (5% CO₂, 1% O₂ and 94% N₂) followed by reoxygenation (5% CO₂, 21% O₂ and 74% N₂). For erastin treatment, cells were treated with erastin (0, 2, 4, 6, 8, 10 and 15 μmol/L) for 24 hours.