Human breast cancer cell lines, including HCC1937, Bcap-37, MDA-MB-231, MCF7, and SK-BR-3, and one normal breast epithelial cell line (Hs 578Bst) were purchased from American Type Culture Collection (ATCC). Hs 578Bst cells were maintained in Hybri-Care Medium (ATCC), supplemented with 30 ng/ml EGF and 10% fetal bovine serum (FBS; Gibco, MA, USA). HCC1937 and Bcap-37 cells were grown in medium containing 90% RPMI (Roswell Park Memorial Institute)-1640 medium (Gibco) and 10% FBS. MDA-MB-231 and MCF7 cells were maintained in DMEM-H medium (Gibco) with 10% FBS. SK-BR-3 cells were incubated in 90% McCoy's 5a medium (Gibco) and 10% FBS.
Hs 578bst
Manufactured by American Type Culture Collection
Sourced in United States
The Hs 578Bst is a cell line derived from human breast tissue. It is a commonly used model for studying breast cancer research and development. The Hs 578Bst cell line is available for purchase through the American Type Culture Collection.
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Lab products found in correlation
Mcf 7, by American Type Culture Collection (12 mentions)
Mda mb 231, by American Type Culture Collection (10 mentions)
Skbr3, by American Type Culture Collection (7 mentions)
Hybri care medium, by American Type Culture Collection (6 mentions)
Hs578t, by American Type Culture Collection (5 mentions)
Mcf 10a, by American Type Culture Collection (4 mentions)
Hcc1937, by American Type Culture Collection (4 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (4 mentions)
Mda mb 468, by American Type Culture Collection (3 mentions)
Bt474, by American Type Culture Collection (3 mentions)
Mda mb 453, by American Type Culture Collection (3 mentions)
1640 medium, by Thermo Fisher Scientific (2 mentions)
Sk br 3, by Thermo Fisher Scientific (2 mentions)
Dmem h medium, by Thermo Fisher Scientific (2 mentions)
Mda mb 231, by Thermo Fisher Scientific (2 mentions)
Mcf 7, by Thermo Fisher Scientific (2 mentions)
Mccoy s 5a medium, by Thermo Fisher Scientific (2 mentions)
Bcap37, by American Type Culture Collection (2 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (2 mentions)
Au565, by American Type Culture Collection (2 mentions)
14 protocols using hs 578bst
1
Culturing Diverse Breast Cell Lines
2
Culturing Breast Cell Lines for Research
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The human breast cancer cell lines MCF-7, T47-D, normal human breast cell (Hs578-Bst), and normal human rectal mucosal cell (FHC) were purchased from American Type Culture Collection (ATCC, Manassas, VA, USA). Normal human bronchial epithelial cell of lung (HBE) was purchased from Mingzhou Biology Company (Ningbo, China). MCF-7, T47-D, and HBE cell lines were cultured with high-glucose DMEM (Hyclone, SH30022.01, USA) medium supplemented with 10% fetal bovine serum (FBS, Gibco, 10,100,147, Australia origin). Hs578-Bst was cultured in ATCC Hybri-Care Medium (Catalog No. 46-X) supplemented with 1.5 g/L sodium bicarbonate, 30 ng/mL mouse epidermal growth factor (EGF), and 10% FBS. FHC was cultured in DMEM:F12 (Gibco, 11,330,032, USA) medium with 10% FBS, 10 ng/mL cholera toxin, 0.005 mg/mL insulin, 0.005 mg/mL transferrin, and 100 ng/mL hydrocortisone. Cells were cultivated in an incubator with a humidified atmosphere of 5% CO2 at 37°C. Both MCF7 and T47-D cell lines are estrogen receptor (ER), progesterone receptor (PR) positive, and human epidermal growth factor receptor-2 (HER2) negative.
3
Cell Culture Protocols for Cancer Research
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Stable transfected human T-cell leukemia Jurkat cell lines Jneo and JBcl2 were kindly provided by Dr. Chris Bleackley (University of Alberta), JR cells were kindly provided by Dr. Hannah Rabinowich (University of Pittsburgh). The human normal breast epithelial cell lines MCF-10A was obtained from ATCC and maintained as previously described [42 (link)]. B16-F0 (ATCC, CRL-6322), B16-F10 (ATCC, CRL-6475), Hs578BST (ATCC, HTB-125) and Hs578T (ATCC, HTB-126) cells were obtained from ATCC (Manassas, VA, USA). B16-F0, B16-F10 and Hs578T were maintained in Dulbecco's Modified Eagle's Medium, high glucose, with 10% FCS; Hs578BST was maintained in Dulbecco's Modified Eagle's Medium, high glucose, with 10% FCS and 30ng/ml EGF. Z-VAD-fmk and z-DEVD-fmk were purchased from BD Pharmingen (Mississauga, ON, Canada), SYTOX Green death stain and Alexa Fluor 647 Annexin V conjugate were all obtained from Invitrogen (Carlsbad, CA, USA). All chemicals were purchased from Sigma-Aldrich (St Louis, MO, USA) unless indicated otherwise.
4
Cytotoxicity Screening of Cell Lines
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A549 human lung adenocarcinoma cells, NCM460 human normal colon epithelial cells, WI-38 human embryonic lung fibroblast cells, H9C2 rat cardiomyoblast cells, normal human hepatic cell line L02, and human normal breast cell line HS578BST were purchased from ATCC (Manassas, VA) and cultured in DMEM supplemented with 10% FBS (FBS was inactivated in water bath at 56 ℃ for 30 min before used) following the suppliers instructions (at 37 ℃, in an atmosphere of 5% CO2). This culture condition is also applied for the cell lines experiments. NCI-H460 human lung large cell carcinoma cells were kindly provided by the Stem Cell Bank, Chinese Academy of Sciences. The effects of chemotherapy agents on cell viability were determined by MTT assay. In brief, cancer cells (3 × 104 cells/mL) or normal cells (10 × 104 cells/mL) were plated in 96-well plates and allowed to attach for 24 h at 37 °C in an atmosphere with 5% CO2. Different concentrations of agents were added and incubated for the indicated times, then 25 μL of MTT (5 mg/mL in 1 × PBS (10 mM, pH = 7.4)) was added per well and incubated for 5 h. Absorbance at 570 nm was recorded and cell viability was calculated as percentage of untreated groups.
5
TNBC Cell Culture Protocol
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Human TNBC cell lines MDA-MB-231 and MDA-MB-468, and human normal breast epithelial cell line Hs578Bst were purchased from the ATCC (Manassas, VA, USA). The cells were cultured in Dulbecco's modified Eagle's medium (DMEM; Thermo Fisher Scientific, Inc., Waltham, MA, USA) supplemented with 10% fetal bovine serum (FBS; Zeta Life, Inc., San Francisco, CA, USA) at 37 °C 5% CO2.
6
Maintenance of Breast Cancer Cell Lines
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The Breast cancer cell lines including MCF-7, MDA-231, SKBR3, SUM102, ZR75B and BT474 were originally purchased from American Type Culture Collection (ATCC, Manassas, VA, USA) and were maintained in Dulbecco’s Modified Eagle’s Medium containing 10% fetal bovine serum, 100 units/mL penicillin, and 100 μg/mL streptomycin. Non-cancerous human mammary epithelial cells MCF10A were originally purchased from ATCC and were maintained in Dulbecco’s modified Eagle’s medium containing 10% fetal bovine serum, 100 ng/ml EGF, 50 ng/ml Insulin, 100 units/mL penicillin, and 100 μg/mL streptomycin. Fibroblasts Hs578Bst were obtained from ATCC and maintained in Hybri-Care Medium (ATCC, Manassas, VA, USA) with 30 ng/ml EGF, 100 units/mL penicillin, and 100 μg/mL streptomycin. Fibroblasts 1068SK were maintained in Dulbecco’s Modified Eagle’s medium containing 10% fetal bovine serum, 2 mmol/L glutamine, 100 units/mL penicillin, and 100 μg/mL streptomycin. All the cell lines were cultured in a humidified atmosphere of 95% air and 5% CO2 at 37 °C.
7
Cell Line Authentication and Characterization
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The Research Resource Identifiers (RRIDs) for all cell lines used are provided in Table S2 . HMEC (human mammary epithelial cells) were a gift from M. R. Stampfer [39 ]. All other cell lines were purchased from DSMZ (“Deutsche Sammlung von Mikro-Organismen und Zellkulturen,” Braunschweig, Germany): CAL-51, HCC1143, HCC1937, and KPL-1, or ATCC (American Type Culture Collection, Manassas, USA): AU565, BT-474, CAMA-1, Hs 578T, Hs 578Bst, MCF-7, MCF-10A, MCF-10F, MDA-MB-231, MDA-MB-453, MDA-MB-468, SK-BR-3, T-47D, and ZR-75-1. Cell culture conditions of all cell lines were described previously [40 (link)]. DSMZ and ATCC authenticate all cell lines by STR profiling before distribution. Genomic DNA and total RNA were isolated from all cell lines immediately after receipt, i.e., within three to eight passages [40 (link),41 (link)].
8
Breast Cancer Tissue Acquisition and Murine Xenograft Model
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A total of 30 breast cancer tissues and adjacent non-tumor tissues were obtained from patients at Huaihe Hospital (Kaifeng, China). All patients (age range, 37-70 years; no smoking history available) had a non-specific type of invasive breast cancer (10 cases of grade II and 20 cases of grade III breast cancer; 13 cases of non-lymph node metastasis; and 17 cases of lymph node metastasis). Balb/c nude mice (specific pathogen-free; female; 4-6 weeks) weighing 18-22 g were supplied by Beijing Weitong Lihua Experimental Animal Technology Co., Ltd. The mice were allowed free access to food and water, and were housed in an environment with constant temperature (24±1°C), humidity (50-70%), ventilation rate (10-20 times/h), noise (<40 db) and working illumination (250 lx) under a 12-h light/dark cycle. Drinking water, food and experimental supplies were sterilized and disinfected. The present study was approved by the Ethics Committee at the Medical School, Henan University (Kaifeng, China). Early metastatic breast cancer cells (MDA-MB-231), non-metastatic breast cancer cells (MCF7) and mammary epithelial cells (Hs578Bst) were obtained from the American Type Culture Collection, and cultured in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% fetal bovine serum (Zeta-life) in a 37°C incubator with 5% CO2.
9
Breast Cancer Samples and Cell Lines Study
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Seventy breast cancer samples and twenty-nine normal breast tissues were collected from the department of radiotherapy, Changzhou Tumor Hospital, Soochow University. Pathological diagnosis of breast cancer was performed according to World Health Organization criteria (Hanby et al., 2004 ). Informed written consent was obtained from all patients and donors. The four non-aggressive human breast cancer cell lines MCF-7, MDA-MB-453, T47D, SKBR3 and the normal breast cell line HS578Bst were purchased from the American Type Culture Collection (ATCC). All cell lines were grown in 5% CO2 at 37 °C incubator and maintained according to providers' recommendations. This study protocol was approved by the Medical Ethics Committee of Changzhou Tumor Hospital, Soochow University. The clinic pathological features of 70 patients are summarized in Table 1 .Table 1
The clinical characteristics of the breast cancer patients.
| Variable | Number of patients |
|---|---|
| Age (years) | |
| <50 | 32 |
| ≥50 | 38 |
| Tumor size (cm) | |
| <2 | 41 |
| ≥2 | 29 |
| Clinical stages | |
| I/II | 33 |
| III/IV | 37 |
| Lymph nodes metastasis | |
| Negative (N) | 43 |
| Positive (P) | 27 |
| Histological subtype | |
| Ductal | 62 |
| Lobular | 6 |
| Lobular and ductal mixed forms | 2 |
10
Evaluation of Deguelin and FGFR Inhibitor in Breast Cancer Cells
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Human breast cancer cell lines MCF‐7, MDA‐MB‐231 and normal mammary epithelial cell line Hs 578Bst were purchased from American Type Culture Collection (ATCC) and cultured as suggested. RPMI 1640, F‐15 medium, fetal bovine serum (FBS), and Penicillin Streptomycin were purchased from Gibco (Guangzhou, China). Hybri‐Care Medium was from ATCC. Deguelin (Sigma, Shanghai, China) was dissolved in dimethyl sulfoxide (DMSO) at a 1 mmol/L stock concentration and stored at −80°C. FGFR inhibitor SU5402 (Sigma) was dissolved to 1 mol/L as a stock solution. Antibody against FGFR4, Phospho‐AKT (S473), Phospho‐p44/p42 MAPK, p44/p42 MAPK, Phospho‐Histone H3, β‐actin, and HRP‐conjugated secondary antibody were all obtain from Cell Signaling Technology (Guangzhou, China).
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