Bt 474

CAFs were isolated as previously described.^{45 (link)} Briefly, tissue from early-stage IDC (stage 1) that was less than 10 mm in diameter was sliced and then digested overnight with a collagenase preparation (ISU ABXIS; Seoul, South Korea). Digested tissue was filtered through a 70 μm cell strainer (SPL Life Science; Pocheon-si, South Korea). Cells were separated by Ficoll gradients, washed with PBS, resuspended with DMEM/F12 cell culture medium containing 20% (v/v) fetal bovine serum (FBS), 100 IU/ml penicillin, and 100 μg/ml streptomycin (Gibco BRL; Grand Island, NY, USA) and cultured at 37 °C in a humidified incubator containing 5% CO₂. The fibrotic nature of the isolated cells was confirmed by microscopic determination of morphology and immunofluorescence characterization using the antibodies against vimentin (Abcam; Cambridge, UK), cytokeratin (Dako; Glostrup, Denmark) and cytokeratin 5 (Novocastra; Newcastle upon Tyne, UK). Breast cancer cell lines (BT-474, MCF7, SK-BR-3, MDA-MB-231) were purchased from Korean Cell Line Bank (Seoul, South Korea) (authenticated using morphology and STR profiling) and cultured with DMEM cell culture medium containing 10% (v/v) fetal bovine serum (FBS), 100 IU/ml penicillin, and 100 μg/ml streptomycin at 37 °C in a humidified incubator containing 5% CO₂.

The human BRC cell lines BT549, BT20, T47D, ZR75-1, SKBR3, BT474, and MDA-MB453 were purchased from the Korean Cell Line Bank. Other cell lines (MDA-MB231 and MCF7) were obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA). Cells from the ATCC were certified by the results of a short tandem repeat (STR) DNA profiling assay, a cytochrome C oxidase I assay, and a mycoplasma contamination assay. The nine BRC cell lines were characterized by the short tandem repeat (STR)-polymerase chain reaction (PCR) method and screened for mycoplasma contamination.
The BRC cells used in this study were cultured in Roswell Park Memorial Institute (RPMI)-1640 medium (HyClone, Logan, UT, USA) supplemented with 10% fetal bovine serum (FBS, HyClone) and 1% penicillin/streptomycin (HyClone) (hereafter referred to as complete RPMI1640 medium) at 37 °C in 5% CO₂.

MCF10A, ZR75-30, MDA-MB-453, BT20, JIMT-1, MDA-MB-231, and HEK293T cells were obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA). MCF7, T47D, ZR75-1, SKBR3, BT474, HCC-1954, HCC1419, and HS578T cells were purchased from the Korean Cell Line Bank. MCF7, T47D, MDA-MB-231, and HEK293T cells were cultured in DMEM (Welgene, Daegu, Republic of Korea) supplemented with 10% FBS, and SKBR3, BT474, HCC-1954, HCC-1419, JIMT-1, and HS5788T cells were cultured in RPMI (Welgene) supplemented with 10% FBS at 37 °C in a 5% CO2 atmosphere. Trastuzumab was obtained from Roche (Basel, Switzerland). The β-Catenin/TCF inhibitor FH-535 was purchased from Sigma-Aldrich (St. Louis, MO, USA).

The MCF7 and BT474 cell lines (Korean Cell Line Bank, Seoul, Korea) were used to determine the effect of S. aureus EVs and tamoxifen in breast cancer cells. A total of 5 × 10⁵ cells were incubated in Dulbecco’s modified eagle medium supplemented with 10% fetal bovine serum. PBS was used to wash the cells, and fresh medium was added after 24 h of incubation. The EVs were then titered and diluted with distilled water. Cells in the control group were treated with distilled water, while those in the experimental group were treated with EVs at 10 ng/mL, 100 ng/mL, and 1 μg/mL concentrations for 72 h. S. aureus EVs (1 μg/mL) were used in co-treatment experiments with tamoxifen. Cells were treated with 1 μg/mL S. aureus EVs, 10 μM tamoxifen, or 10 μM tamoxifen plus 1 μg/mL S. aureus EVs for 72 h [31 (link),32 (link)]. A trypan blue viability assay was performed to measure relative cell viability.

The human MCF-7, MDA-MB-231, HCC-1143, HCC-1187, BT20, BT-474, and T47D breast cancer cell lines were purchased from the Korean Cell Line Bank (Seoul, Korea). Cells were cultured in RPMI 1640 media supplemented with 10% heat-inactivated fetal bovine serum (FBS) (Welgene, Korea) and were maintained at 37 °C in a humidified 5% CO₂ atmosphere. Dexamethasone (DXM), Dexamethasone 21-phosphate (DXM-21-P), betamethasone 21-phosphate (BTM-21-P), mifepristone (Sigma Aldrich, St. Louis, MO, USA), Wortmannin, QNZ, MK2206, idasanutlin, AS1842856, XMU-MP-1 and p38 MAPK inhibitor (Selleckchem, Houston, TX, USA) were used in this study.