Si gas5

miR-23a mimics (miR-23a), control mimics (miR-control), siRNA against GAS5 (si-GAS5), siRNA control (si-control), pcDNA-GAS5, and pcDNA empty control (pcDNA-control) were purchased from GenePharma (Shanghai, P.R. China). Cultured A549 and H157 cells were transfected with miR-23a, si-GAS5, pcDNA-GAS5, pcDNA-GAS5 + miR-23a, or their corresponding control at approximately 60%–70% confluence using Lipofectamine 2000 (Invitrogen). Transfection efficiency was checked by quantitative real-time polymerase chain reaction (qRT-PCR). Subsequent experiments were performed 48 h after transfection.

CCA cell lines, including RBE, CCLP1, HuCCT1 and HCCC-9810 cholangiocarcinoma cell lines and a normal human intrahepatic biliary epithelial cell (HIBEC) line were purchased from ATCC. Cells were cultured in the atmosphere with 5% CO2 at 37 °C. The medium of cell culture is Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% fetal bovine serum. si-GAS5 and hsa-miR-1297 inhibitor were purchased from GenePharma (Shanghai China). Lipofectamine 3000 reagent (Invitrogen, USA) was used for cell transfection. The transfection of cell with si-GAS5 and hsa-miR-1297 inhibitor were performed according to the manufacturer’s instructions. The PCR primers were miR-1297-forward: 5′-TCGGCAGGTTCAAGTAATT-3′; miR-1297-reverse: 5′-CTCAACTGGTGTCGTGGA-3′. The sequence of siRNA targeting GAS5 was 5′-UCUUCAAUCAUGAAUUCUGAG-3′. The negative control nontargeting sequence was 5′-ACGUGACACGUUCGGAGAATT-3′.