Si pdcd4

Empty pcDNA3.1 vector (Vector) was obtained from Genepharma (Shanghai, China). TUG1 or PDCD4 overexpressing vector pcDNA3.1-TUG1 or pcDNA3.1-PDCD4 (TUG1 or PDCD4), small interfering RNAs against TUG1 (si-TUG1 #1 or si-TUG1 #2) or PDCD4 (si-PDCD4) and their scramble negative control (si-con) were chemically synthesized by Genepharma (Shanghai, China). All cell transfections were performed using the Lipofectamine 2000 (Invitrogen, Carlsbad, CA, USA).

An anti-miR-21 oligonucleotide (AMO-miR-21) was designed according to sequence complementary to mature miRNA-21: AMO-miR-21, 5′-ATAAGCTA-3′ (8 bp). A control scramble AMO (SCR) 5′ -TCATACTA-3′ (8 bp) was also synthesized (Additional file 1: Figure S2). All oligodeoxynucleotides were chemically synthesized and modified with phosphorothioate and/or fluorescein isothiocyanate (FITC) by the Shanghai Sangon Bio-engineering Company, China. The siRNA sequence of PDCD4 (siPDCD4) was 5′-AAGGUGGCUGGAACAUCUAUU-3′. The RNA duplexes were synthesized and purified by Shanghai GenePharma Company, China.

Normal human liver L02 cells and the HCC cell lines (Huh-6, Huh-7, SNU-449 and Li-7) were purchased from the Institute of Biochemistry and Cell Biology of the Chinese Academy of Sciences (Shanghai, China). All the cells were cultured in DMEM medium (Gibco, Grand Island, NY, U.S.A.) containing 10% fetal bovine serum (Gibco). Cell transfection was performed using Lipofectamine 2000 (Invitrogen, Carlsbad, CA, U.S.A.) according to the manufacturer’s instructions. Negative control inhibitor (NC inhibitor), miR-183-5p inhibitor, miR-183-5p mimics and small interfering RNA specifically targeting PDCD4 (si-PDCD4) were synthesized by GenePharma Co. Ltd (Shanghai, China). When cells were allowed to grow to 70–90% confluence, cell transfection was performed. NC inhibitor or miR-183-5p inhibitor at a final concentration of 50 nM was transfected into HCC cells. For the rescue assay, si-PDCD4 at a final concentration of 50 nM was transfected into miR-183-5p inhibitor-treated HCC cells. The sequence information of NC inhibitor, miR-183-5p inhibitor, si-PDCD4, negative control mimics and miR-183-5p mimics was listed as follows: NC inhibitor, 5′-UUCUCCGAUCUGGCUACAGU-3′; miR-183-5p inhibitor, 5′-UUCUGACCAUCUUAAGUGAU-3′; si-PDCD4, 5′-GGAGGUGGAUGUGAAAGAU-3′; negative control mimics, 5′-ACUACUGAGUGACAGUAGA-3′; miR-183-5p mimics, 5′-UAUGGCACUGGUAGAAUUCACU-′3.