Mir 23a mimic

Hepa1–6 and Hca-P cells were cultured in six-well plates for 12–24 h before transfection. miR-23a mimic, miR-23a inhibitor (antisense oligonucleotide) and scrambled miRNA oligonucleotide pairs were purchased from Ribo-bio (Guangzhou, China). Runx2 siRNAs were designed and synthesized by GenePharma (Shanghai, China). Both cell lines were transfected with 100 nM miR-23a mimic, miR-23a inhibitor, scrambled miRNA or Runx2 siRNA using riboFECT™ CP transfection reagent (Ribo-bio, China) according to the manufacturer’s instructions. The eukaryotic expression plasmid pCMV-Runx2 was obtained from the Karsenty Lab (Columbia University) and was transfected into Hepa1–6 and Hca-P cells using Lipofectamine 2000 (Invitrogen, USA) according to the standard protocol. Cells were collected 48 h after transfection for further assays.

HEC-1-A cells (human endometrial carcinoma cells-1-A) are human endometrium adenocarcinoma cell line and were obtained from American Type Culture Collection (Manassas, VA, USA) and were cultured in McCoy’s 5A modified medium supplemented with 10 % fetal bovine serum and 1 % penicillin/streptomycin (Sigma-Aldrich, St Louis, MO, USA). HEC-1-A cells are used as an accepted model to study endometrial cancer [15 (link)]. MiR-23a mimic, inhibitor, agomir or antagomir were purchased from RiboBio Inc (GuangZhou, China). Human recombinant TGFβ1 was obtained from R & D Systems Inc (Minneapolis, MN, USA).

Cells were cultured with RPMI-1640 medium containing 10% FBS overnight, and then 50 nmol/L miR-23a mimic, miR-23a inhibitor and their respective negative control (RiboBio, Guangzhou, China) were transfected into cells using riboFect™ CP Transfection Kit (Ribobio) according to the manufacturer’s instruction, respectively. 24h after transfection, the experiments were performed on the transfected cells.