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Scientific Reports Aug 2017Tumour cell migration has an important impact on tumour metastasis. Magnetic manipulation is an ascendant method for guiding and patterning cells. Here, a unique...
Tumour cell migration has an important impact on tumour metastasis. Magnetic manipulation is an ascendant method for guiding and patterning cells. Here, a unique miniaturized microfluidic chip integrating cell isolation and migration assay was designed to isolate and investigate cell migration. The chip was fabricated and composed of a magnet adapter, a polytetrafluoroethylene(PDMS) microfluidic chip and six magnetic rings. This device was used to isolate MCF-7 cells from MDA-MB-231-RFP cells and evaluate the effects of TGF-β on MCF-7 cells. First, the two cell types were mixed and incubated with magnetic beads modified with an anti-EpCAM antibody. Then, they were slowly introduced into the chip. MCF-7 cells bond to the magnetic beads in a ring-shaped pattern, while MDA-MB-231-RFP cells were washed away by PBS. Cell viability was examined during culturing in the micro-channel. The effects of TGF-β on MCF-7 cells were evaluated by migration distance and protein expression. The integrated method presented here is novel, low-cost and easy for performing cell isolation and migration assay. The method could be beneficial for developing microfluidic device applications for cancer metastasis research and could provide a new method for biological experimentation.
Topics: Cell Movement; Epithelial Cells; Female; Humans; Immunomagnetic Separation; MCF-7 Cells; Microfluidics; Transforming Growth Factor beta
PubMed: 28827722
DOI: 10.1038/s41598-017-08661-z -
Cancer Investigation Sep 2017Using estrogen-dependent MCF-7 breast cancer cells and tamoxifen-resistant MCF-7/T subline we have shown that their co-cultivation lead to increase in tamoxifen...
Using estrogen-dependent MCF-7 breast cancer cells and tamoxifen-resistant MCF-7/T subline we have shown that their co-cultivation lead to increase in tamoxifen resistance in the parent MCF-7 cells. The proteome analysis of MCF-7/T cells and new-generated resistant cells revealed 21 common proteins differently expressed in both the resistant cell lines, among them - 6 proteins were associated with the drug or hormonal resistance. Both resistant lines were characterized with suppression of estrogen receptor and activation of SNAIL1-signaling - mesenchymal pathway playing an important role in the down-regulation of estrogen receptor and maintaining of the estrogen-independent phenotype.
Topics: Antineoplastic Agents, Hormonal; Breast Neoplasms; Cells, Cultured; Coculture Techniques; Drug Resistance, Neoplasm; Epithelial Cells; Epithelial-Mesenchymal Transition; Female; Gene Expression Regulation, Neoplastic; Humans; MCF-7 Cells; Mass Spectrometry; Proteome; Receptors, Estrogen; Signal Transduction; Snail Family Transcription Factors; Tamoxifen
PubMed: 28910558
DOI: 10.1080/07357907.2017.1368081 -
Journal of the Egyptian National Cancer... Dec 2022Mammosphere formation assay has become a versatile tool to quantify the activity of putative breast cancer stem cells in non-adherent in vitro cultures. However,...
BACKGROUND
Mammosphere formation assay has become a versatile tool to quantify the activity of putative breast cancer stem cells in non-adherent in vitro cultures. However, optimizing the suspension culture system is crucial to establish mammosphere cultures from primary breast tumors.
METHODS
This study aimed at determining the self-renewal and sphere-forming potential of breast cancer stem-like cells derived from human primary invasive ductal carcinoma and normal breast tissue samples, and MCF-7 breast cancer cell line using an optimal suspension culture system. Mammosphere-forming efficiency of the mammospheres generated from the tissue samples and cell line were compared. We evaluated the expression of CD44/CD24/ and CD49f/EpCAM/ phenotypes in the stem-like cells by flow cytometry. CK-18, CK-19, α-SMA, and EpCAM marker expression was assessed using immunohistochemical staining.
RESULTS
Breast epithelial cells isolated from the three samples formed two-dimensional spheroids in suspension cultures. Interestingly, mammospheres formed from patient-derived primary breast tumors were enriched in breast cancer stem-like cells with the phenotype CD44/CD24/ and exhibited a relatively more number of large spheres when compared to the normal breast stem cells. MCF-7-derived SCs were more aggressive and resulted in the formation of a significantly higher number of spheroids. The expression of CK-18/CK-19 and α-SMA/EpCAM proteins was confirmed in breast cancer tissues.
CONCLUSIONS
Thus, the use of primary tumor specimens and breast cancer cell lines as suitable models for elucidating the breast cancer stem cell activity was validated using mammosphere culture system.
Topics: Humans; Female; MCF-7 Cells; Breast; Breast Neoplasms; Neoplastic Stem Cells
PubMed: 36504339
DOI: 10.1186/s43046-022-00152-1 -
International Journal of Radiation... May 2018Breast cancer is one of the most common malignant tumors in women all over the world. Many of these women resist the common treatments. Therefore, it is important to...
PURPOSE
Breast cancer is one of the most common malignant tumors in women all over the world. Many of these women resist the common treatments. Therefore, it is important to find new products to increase the efficacy of the treatment process. Legume beans, with their various pharmacological properties, can be regarded as a sensitizer when they are combined with radiation. The present study strove to survey the radio-sensitivity effect of proteins isolated from mung bean aqueous extract on the human breast adenocarcinoma cell line (MCF-7), human cervical cancer cells (Hela) and the human dermal fibroblast cell line.
MATERIALS AND METHODS
The mung bean aqueous extract was partially purified by ammonium sulfate. At first, various concentrations of the extracts were used to evaluate the inhibitory activity by MTT cell proliferation assay.
RESULTS
The results showed that MCF-7 cells and Hela cells were inhibited by an IC value of less than 250 and 411 µg/ml, respectively, but it proved to have a proliferation effect on the fibroblast cells. Then, the cells were incubated with 250 µg/ml extract and exposed to 2, 4, and 6 Gy of X-ray radiation. The percentage of the cell survival was investigated through MTT and the clonogenic assay. Apoptosis was measured using acridine orange/ethidium bromide staining. The results demonstrated that the treated MCF-7 cells and Hela cells had significant radio-sensitivity compared with the results of the control group in radiation dose manner in all MTT, clonogenic, and apoptosis assays. In contrast, the treated fibroblast showed a protective effect against radiation.
CONCLUSION
The results suggest that mung bean proteins have the capacity to be regarded as a radio-sensitizer for breast cancer. Our results also indicated that it could be worth to investigate on mung bean proteins further and they should be tested in animal models for being treated in radiotherapy.
Topics: Apoptosis; Cell Line; Cell Proliferation; Cell Survival; Dose-Response Relationship, Radiation; Female; Fibroblasts; HeLa Cells; Humans; Inhibitory Concentration 50; MCF-7 Cells; Plant Extracts; Radiation Tolerance; Skin; Vigna
PubMed: 29482484
DOI: 10.1080/09553002.2018.1446226 -
Molecular and Cellular Biochemistry Jan 2017Calyptranthes tricona is a species (Myrtaceae) native to South Brazil. Plants belonging to this family are folkloric used for analgesia, inflammation, and infectious...
Calyptranthes tricona is a species (Myrtaceae) native to South Brazil. Plants belonging to this family are folkloric used for analgesia, inflammation, and infectious diseases. However, little is known about the toxic potential of C. tricona. The present study aimed to evaluate the antioxidant activity of C. tricona ethanol and hexane leaf extracts, as well as verify their effect on human lymphocytes and MCF-7 cells. The extracts were subjected to preliminary phytochemical screening, antioxidant activity using DPPH and ORAC methods. Genotoxic and mutagenic effects in cultured human lymphocytes were assessed using the comet assay and the micronucleus assay, respectively. In addition, cell viability by MTT assay and fluorometric analysis of mitochondrial potential and caspases-9 activity were performed in order to verify the possible effects of both extracts on HO-induced cell death of MCF-7 cells. Our findings revealed that the phenol content and the antioxidant activity were only present in the ethanol extract. Also, the phytochemical screening presented steroids, triterpenoids, condensed tannins, and flavones as the main compounds. However, both extracts were capable of inducing concentration-dependent DNA damage in human lymphocytes. When treating MCF-7 cells with the extracts, both of them inhibited MCF-7 cell death in response to oxidative stress through a decrease of mitochondrial depolarization and caspases-9 activity. Thus, our results need to be considered in future in vitro and in vivo studies of C. tricona effects. In the meanwhile, we recommend caution in the acute/chronic use of this homemade preparation for medicinal purpose.
Topics: Cell Death; DNA Damage; Female; Humans; Hydrogen Peroxide; Lymphocytes; MCF-7 Cells; Myrtaceae; Oxidative Stress; Plant Extracts
PubMed: 27704465
DOI: 10.1007/s11010-016-2840-9 -
The Journal of Steroid Biochemistry and... 1997Stable aromatase-expressing MCF-7 and T-47D cell lines (i.e. MCF-7aro and T-47Daro) have been prepared by aromatase cDNA transfection and G418 (neomycin) selection....
Stable aromatase-expressing MCF-7 and T-47D cell lines (i.e. MCF-7aro and T-47Daro) have been prepared by aromatase cDNA transfection and G418 (neomycin) selection. MCF-7aro was further subjected to a clonal purification. Aromatase activity in the transfected MCF-7 and T-47D cell lines was determined to be 73 +/- 6 pmol/mg/h and 48 +/- 4 pmol/mg/h, respectively. It is thought that these cell lines express aromatase in a stable manner, as demonstrated by a steady expression of the enzyme during culture in the absence of G418. The growth of these cells could be stimulated by androgens (1-10 nM) as demonstrated through a spheroid culture method. The androgen-stimulated growth could be suppressed by 4-hydroxyandrostenedione (4-OHA) (0.01-0.1 mM) or tamoxifen (50 nM-1 microM). In order to test the hypothesis that tumor aromatase can affect breast tumor growth in a paracrine manner, we have carried out cell culture experiments by co-culturing MCF-7 cells with either MCF-7aro or T-47Daro cells. Testosterone (1 nM) increased cell growth to a similar degree for MCF-7/MCF-7aro co-culture (0.75 x 10(6) cells each type) as with MCF-7aro only (2- to 3-fold). In addition, the enzyme activities remained unchanged for MCF-7/MCF-7aro co-culture samples with and without androgen treatment, indicating that estrogen produced by transfected cells can also stimulate the growth of untransfected cells. The androgen response could be inhibited by an addition of 4-OHA (0.01-0.1 mM). For MCF-7/T-47Daro co-culture experiments, a clear induction of cell growth by androgen was observed, and the level of the increase was similar to that on T-47Daro only. However, for either culture with T-47D only or with MCF-7/T-47Daro co-culture, the aromatase activity was found to increase significantly after testosterone treatment. T-47Daro cells were not subjected to a clonal purification, and it is therefore thought that the androgen treatment may selectively stimulate the growth of high aromatase-expressing T-47Daro cells. These results indicate that estrogen synthesized by tumor aromatase can stimulate breast tumor growth in both an autocrine and a paracrine manner.
Topics: Androgens; Anti-Bacterial Agents; Aromatase; Breast Neoplasms; Cell Communication; Genetic Vectors; Gentamicins; Humans; Neoplasms, Hormone-Dependent; Spheroids, Cellular; Transfection; Tumor Cells, Cultured
PubMed: 9449203
DOI: 10.1016/s0960-0760(97)00068-x -
International Journal of Molecular... Jul 2023In healthy tissues, cells are in mechanical homeostasis. During cancer progression, this equilibrium is disrupted. Cancer cells alter their mechanical phenotype to a...
In healthy tissues, cells are in mechanical homeostasis. During cancer progression, this equilibrium is disrupted. Cancer cells alter their mechanical phenotype to a softer and more fluid-like one than that of healthy cells. This is connected to cytoskeletal remodeling, changed adhesion properties, faster cell proliferation and increased cell motility. In this work, we investigated the mechanical properties of breast cancer cells representative of different breast cancer subtypes, using MCF-7, tamoxifen-resistant MCF-7, MCF10A and MDA-MB-231 cells. We derived viscoelastic properties from atomic force microscopy force spectroscopy measurements and showed that the mechanical properties of the cells are associated with cancer cell malignancy. MCF10A are the stiffest and least fluid-like cells, while tamoxifen-resistant MCF-7 cells are the softest ones. MCF-7 and MDA-MB-231 show an intermediate mechanical phenotype. Confocal fluorescence microscopy on cytoskeletal elements shows differences in actin network organization, as well as changes in focal adhesion localization. These findings provide further evidence of distinct changes in the mechanical properties of cancer cells compared to healthy cells and add to the present understanding of the complex alterations involved in tumorigenesis.
Topics: Humans; Female; Cell Line, Tumor; Cytoskeleton; MCF-7 Cells; Actins; Tamoxifen; Breast Neoplasms; Microscopy, Atomic Force
PubMed: 37569585
DOI: 10.3390/ijms241512208 -
Natural Product Research Jun 2024In the present study, we derivatized several hydroxycinnamic and hydroxybenzoic acids to phenolic amides (PAMs) one step BOP mediated amide coupling reactions. Fifteen...
In the present study, we derivatized several hydroxycinnamic and hydroxybenzoic acids to phenolic amides (PAMs) one step BOP mediated amide coupling reactions. Fifteen PAMs were synthesized in >40% yields and were screened for their cytotoxic activities against four cancer cell lines: THP-1 (leukaemia), HeLa (cervical), HepG2 (liver), and MCF-7 (breast), in comparison to 5-flurouracil (5-FU). Four amides showed IC ranging from 5 to 55 µM against all four cell lines. In contrast, tetradecyl-gallic-amide () affected only THP-1 leukaemia cells with IC of 3.08 µM. The activities of these compounds support the promise of phenolic amides as anticancer agents.
Topics: Humans; Amides; Phenols; HeLa Cells; MCF-7 Cells; Antineoplastic Agents; Hep G2 Cells; Molecular Structure; Cell Line, Tumor; Drug Screening Assays, Antitumor; Inhibitory Concentration 50
PubMed: 37526601
DOI: 10.1080/14786419.2023.2241971 -
European Review For Medical and... May 2020To study the effectiveness of natural killer cell-derived exosome (NK-Exos)-entrapped paclitaxel (PTX-NK-Exos) in enhancing its anti-tumor effect.
OBJECTIVE
To study the effectiveness of natural killer cell-derived exosome (NK-Exos)-entrapped paclitaxel (PTX-NK-Exos) in enhancing its anti-tumor effect.
MATERIALS AND METHODS
The NK-Exos were isolated through ultra-high-speed centrifugation, and the PTX-NK-Exos system was constructed via electroporation. The morphology, particle size, Zeta potential and entrapment rate of PTX-NK-Exos were evaluated using transmission electron microscope (TEM), dynamic light scattering (DLS), Western blotting and high-performance liquid chromatography (HPLC), respectively. The uptake of Exos in human breast cancer MCF-7 cells was observed under a laser confocal microscope. Moreover, the effect of PTX-NK-Exos on MCF-7 cell viability was determined through methyl thiazolyl tetrazolium (MTT) assay, flow cytometry and 4',6-diamidino-2-phenylindole (DAPI) staining. The effects of PTX-NK-Exos on messenger ribonucleic acid (mRNA) and protein expressions of B-cell lymphoma-2 (Bcl-2), Bcl-2 associated X protein (Bax) and Caspase-3 in MCF-7 cells were detected using quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and Western blotting, respectively.
RESULTS
The NK-Exos were successfully isolated via ultra-high-speed centrifugation, and they had uniform particle size and high expression of markers for Exos. MCF-7 cells could take up Exos. The PTX-NK-Exos drug delivery system was successfully prepared using electroporation. In PTX group and NK-Exos group, the proliferation of MCF-7 cells declined, the nuclear apoptosis was evident and the apoptosis rate of MCF-7 cells rose compared with those in Control group. In PTX group and PTX-NK-Exos group, the migration of MCF-7 cells declined compared with that in Control group. According to the results of qRT-PCR and Western blotting, PTX-NK-Exos exerted an anti-tumor effect through inducing the up-regulation of Bax and Caspase-3 in the apoptotic signaling pathway in tumor cells.
CONCLUSIONS
Exos isolated through ultra-high-speed centrifugation can be used to prepare the PTX-NK-Exos drug delivery system through electroporation. Drug-loaded Exos can effectively inhibit proliferation and induce apoptosis of tumor cells, thereby exerting an anti-tumor effect.
Topics: Antineoplastic Agents, Phytogenic; Apoptosis; Cell Movement; Cell Proliferation; Cells, Cultured; Drug Screening Assays, Antitumor; Exosomes; Humans; Killer Cells, Natural; Paclitaxel
PubMed: 32495906
DOI: 10.26355/eurrev_202005_21362 -
Biomedicine & Pharmacotherapy =... Sep 2017We have recently shown that olmesartan could induce toxicity in HeLa and MCF-7 cell lines. In this study we investigated toxicity mechanism of olmesartan in HeLa and...
We have recently shown that olmesartan could induce toxicity in HeLa and MCF-7 cell lines. In this study we investigated toxicity mechanism of olmesartan in HeLa and MCF-7 cell lines. HeLa and MCF-7 cells were cultured in DMEM in optimum conditions. Cells were pretreated with rutin as an antioxidant and treated with olmesartan as a cytotoxic agent. Cell proliferation was determined by MTT assay. The role of ROS was determined using DCFH-DA by flow cytometry analysis. Also, cells were treated with olmesartan (5mM) and Bay 11-7-82 (25μM) for 24h, then expression of apoptotic proteins including Bax, caspase3 and IκB were investigated in both cell lines by western blotting. Cell viability decreased with olmesartan in malignant cell lines. Kinetic of ROS assay showed increment of ROS generation starting at 2h which peaked at 4h after treatment. Pretreatment with antioxidant rutin decreased ROS increment which was consistent with improved viability of olmesartan-treated cells. Apoptosis results showed that olmesartan and Bay 11-7082 increased expression of apoptotic proteins such as Bax, caspase3 and IκB. Results proposed ROS increment and apoptosis could be involving mechanisms in olmesartan-induced toxicity in HeLa and MCF-7 cell lines.
Topics: Apoptosis; Cell Survival; HeLa Cells; Humans; Imidazoles; Intracellular Space; MCF-7 Cells; NF-kappa B; Nitriles; Protective Agents; Reactive Oxygen Species; Rutin; Signal Transduction; Sulfones; Tetrazoles
PubMed: 28666209
DOI: 10.1016/j.biopha.2017.06.074