Mcf 7

The normal breast tissue cell line, MCF-10A, and the MCF7, T47D and MDA-MB-231 breast cancer cell lines were obtained from the American Type Culture Collection (Manassas, VA, USA). MCF-7 and T47D cells were cultured in Dulbecco's modified Eagle's medium (DMEM; Gibco; Thermo Fisher Scientific, Inc., Waltham, MA, USA), and MDA-MB-231 cells were cultured in RPMI-1640 supplemented with 10% fetal bovine serum (FBS; both Invitrogen; Thermo Fisher Scientific, Inc.). MCF-10A cells were cultured in DMEM-F12 supplemented with 5% horse serum (Invitrogen; Thermo Fisher Scientific, Inc.). The cells were incubated in an atmosphere containing 5% CO₂ at 37°C. Lipofectamine^® 2000 (Invitrogen; Thermo Fisher Scientific, Inc.) was used for transfection to transfected into MCF-7 or MDA-MB-231 cells. The relative small interfering (si)RNAs targeting SOX5 (si-SOX5-1 and si-SOX5-2) or enhancer of zeste 2 polycomb repressive complex 2 subunit (EZH2) or negative control and G418 were purchased from Sigma-Aldrich (Merck KGaA, Darmstadt, Germany). SOX5 and vector plasmid were purchased from Genepharma (Shanghai, China). The 70% confluence of MCF-7 or MDA-MB-231 cells were achieved overnight prior to transfection. In each group, 2 µg oligonucleotide were used for transfection. At 48 h following transfection, the cells were harvested for experimentation.

Breast cancer cells (MCF-7; drug sensitive, DS) and TAX-resistant breast cancer cells (MCF-7/TAX; drug resistant, DR) were purchased from The Cell Bank of Type Culture Collection of The Chinese Academy of Sciences and Procell Life Science & Technology Co., Ltd., respectively. The MCF-7 and MCF-7/TAX cells were cultured in Dulbecco's modified Eagle's medium (DMEM; Thermo Fisher Scientific, Inc.) containing 10% fetal bovine serum (FBS; Thermo Fisher Scientific, Inc.) and 100 U/ml penicillin/100 µg/ml streptomycin (Thermo Fisher Scientific, Inc.), and maintained in an incubator containing 5% CO₂ at 37°C.

Human breast cancer cell lines MDA-MB-468, MDA-MB-231 and normal epithelial cell line MCF-10A were purchased from the Cell Type Culture Collection of the Chinese Academy of Sciences (Shanghai, China) and routinely maintained in RPMI-1640 medium (Gibco; Thermo Fisher Scientific, Inc., Waltham, MA, USA) with 10% fetal bovine serum (FBS; Gibco; Thermo Fisher Scientific, Inc.) and human breast cancer cell line MCF-7 was cultured in RPMI 1640 medium. All cells were incubated in a humidified 5% CO₂ atmosphere at 37°C. The miR-135 mimic, miR-135 inhibitor or scrambled miRNA control which expressed green fluorescent protein (GFP) were transfected to the MDA-MB-468 and MCF-7 cells with Lipofectamine 2000 (Invitrogen; Thermo Fisher Scientific, Inc.) according to the manufacturer’s protocol. The sequences were as follows: miR-135 mimic forward, 5′-UAU GGC UUU UCA UUC CUA UGU GA-3′ and reverse, 5′-ACA UAG GAA UGA AAA GCC AUA UU-3′; miR-135 inhibitor forward, 5′-UCA CAU AGG AAU GAA AAG CCA UA-3′ and reverse, 5′-CAG UAC UUU UGU GUA GUA CAA-3′; and scrambled miRNA control forward, 5′-UUC UCC GAA CGU GUC ACG UTT-3′ and reverse, 5′-ACG UGA CAC GUU CGG AGA ATT-3′. A total of 2 days later, cells were collected and stored at −80°C.