MCF 10A cells are a non tumorigenic mammary epithelial cell line isolated from fibrocystic breast tissue of a 36 year old white female in 1984. These cells exhibit characteristics of luminal ductal cells, but do not show characteristics of myoepithelial cells, and respond to insulin, glucocorticoids, cholera enterotoxin, and epidermal growth factor (EGF).
MCF 10A cells exhibit three-dimensional growth ability during the cultivation process, forming a dome shaped structure in the collagen matrix without showing signs of terminal differentiation or aging. These cells are widely used to study the function of normal breast cells, gene function analysis of breast cancer, and cell interaction in breast development.
However, there is some controversy regarding whether MCF 10A cells can reliably represent normal human breast cells. Some studies have shown that under different culture conditions, MCF 10A cells may not fully represent the phenotype of normal breast epithelial cells, especially in terms of basal origin and stem cell characteristics [2]. In addition, studies have shown that MCF 10A cells exhibit a basal like phenotype under certain conditions, but share many similarities with mesenchymal carcinoma cell lines, further questioning their applicability as a normal breast epithelial model [2].
In general, MCF 10A cell line has become an important tool in breast cancer research because of its non tumorigenicity and three-dimensional growth ability, but whether it can accurately reflect the characteristics of normal breast epithelial cells still needs careful evaluation [2].
What are the phenotypic changes of MCF 10A cells under different culture conditions?
The phenotypic changes of MCF-10A cells under different culture conditions are mainly reflected in significant differences in their morphology and structure. These changes can be observed through two-dimensional culture, three-dimensional culture, and specific treatment conditions such as oxidative stress, hormone deprivation, etc.
Under two-dimensional culture conditions, MCF-10A cells typically form a cuboidal epithelial morphology, exhibiting expression of basal/myoepithelial and papillary cell markers such as actin, alpha smooth muscle actin, and N-cadherin [2]. These cells have basal origin and exhibit strong CK5 staining and moderate levels of CK17 expression under certain conditions [2].
Under three-dimensional culture conditions, MCF-10A cells exhibit more complex morphological changes. For example, in Matrigel, MCF-10A cells form spherical structures, while in mixed Matrigel/collagen I gel, cells grow branch and acinar structures [6]. This three-dimensional culture environment enables MCF-10A cells to form tissue like structures [4].
In addition, MCF-10A cells also exhibit significant morphological changes under different oxidative stress conditions. For example, under long-term exposure to H2O2, MCF-10A cells become larger and more extensive, and transformation foci begin to appear after 6 weeks [5]. These transformation foci are related to the oxidative microenvironment and indicate the phenotypic changes of MCF-10A cells under oxidative stress [5].
Under hormone deprivation conditions, the morphology of MCF-10A cells also underwent significant changes. For example, in - HC/- EGF medium, the appearance of the transformation focus of MCF-10A cells is affected, exhibiting different growth characteristics [3]. In addition, MCF-10A cells exhibit a unique fibrous cell morphology in media lacking HC and EGF [3].
The phenotypic changes of MCF-10A cells under different culture conditions include a transition from cuboidal epithelial morphology to spherical structure, branching and acinar structure, as well as significant morphological changes under oxidative stress and hormone deprivation conditions.
What are the differences in gene expression and function between MCF 10A cells and normal breast epithelial cells?
The differences in gene expression and function between MCF10A cells and normal breast epithelial cells are mainly reflected in the following aspects:
1. Gene expression pattern:
MCF10A cells are similar in gene expression to primary mammary epithelial cells (HMEC), but there are some significant differences. For example, MCF10A cells exhibit higher levels of expression of certain genes, such as IL-1 α, IL-1 β, CXCL8, IL-32, CXCL1, CXCL2, and CCL20 [7]. The upregulation of these genes may reflect the gene expression changes that occur in MCF10A cells during in vitro culture.
· After co culture with breast cancer cells, MCF10A cells showed a significant increase in S100A8 and S100A9 genes, while the expression of CDH1, CD24 and other breast epithelial differentiation markers decreased [8]. This indicates that the gene expression pattern of MCF10A cells changes when interacting with cancer cells, thereby affecting their function.
2. Stem cell/progenitor cell markers:
MCF10A cells exhibit different expression levels of stem/progenitor cell markers under different culture conditions. For example, in 3D Matrigel culture, MCF10A cells lost stem cell markers CD49f, CD44, and ALDH1A3, but gained epithelial markers CK8, CK18, and CK7 [2]. This indicates that MCF10A cells may lose their stem cell characteristics under specific conditions and acquire more mature epithelial cell features.
3. Signal pathway and function:
After RLO treatment, the expression levels of p53, Caspase-3, DMP1, and p21 were significantly increased in MCF10A cells, while the expression level of HER2 decreased [6]. These changes indicate that MCF10A cells undergo adjustments in their signaling pathways and functions when stimulated by specific chemicals, thereby affecting cell proliferation and apoptosis.
· After co culturing with breast cancer cells, MCF10A cells showed significant imbalance in a variety of cancer related pathways, including metabolic pathway, cell adhesion, growth factor signal transduction and TP53 pathway [8]. These changes may reflect the functional reprogramming experienced by MCF10A cells in the cancer environment.
4. Morphology and structure:
MCF10A cells can reproduce acinar and ductal like structures in co culture or triple culture, exhibiting a more mature phenotype and function than single-cell culture [9]. However, despite being considered a "normal" mammary epithelial cell line, MCF10A cells exhibit genetic abnormalities, carrying genetic and epigenetic abnormalities associated with long-term in vitro culture [9].
5. Composition of nuclear matrix proteins:
· MCF10A cell line and its derived cells showed a phenotype containing nuclear matrix proteins found in normal and breast cancer tissues [12]. This indicates that MCF10A cells may represent an intermediate step in the transformation of breast cancer, and the changes of nuclear matrix proteins may serve as biomarkers in the transformation of breast cancer.
Although MCF10A cells are often used as a model for normal breast epithelial cells, significant differences still exist in gene expression, stem cell characteristics, signaling pathways, morphology, and nuclear matrix protein composition.
How to evaluate the effectiveness of MCF 10A cells as a research model for breast cancer?
To evaluate the effectiveness of MCF10A cells as a research model for breast cancer, we can analyze it from many aspects, including its biological characteristics, molecular marker expression, cell behavior, and simulation ability at different stages of breast cancer progression.
The MCF10A cell line is derived from benign proliferative breast tissue, with a stable near diploid karyotype and moderate genetic modifications such as loss of the p16 gene [4]. These cells exhibit important features of normal breast tissue in three-dimensional reconstructed basement membrane (rBM) culture, such as proliferation, cell cycle arrest, apical basal polarization, and ultimately apoptosis, forming the nipple space [4]. These characteristics make MCF10A cells an ideal model for studying the malignant progression of breast cancer.
MCF10A cells can express breast cancer related genes and produce molecular markers related to breast cancer [19]. For example, proton induced biomolecular changes in MCF-10A breast cells were evaluated using Fourier transform infrared spectroscopy (µ - FT-IR), showing the different contributions of DNA, proteins, lipids, and carbohydrate cellular components [19]. In addition, the genome changes of MCF10A cell line series in three-dimensional spheroid culture were analyzed by RNA sequencing technology, which verified the effectiveness of this system as an accurate molecular model of breast cancer progression [15].
MCF10A cells exhibit high heterogeneity and variability in three-dimensional culture systems, but they show high heterogeneity and variability in in vitro culture [19]. Through a series of cell biology and molecular biology analysis methods, such as SRB staining, BrdU labeling, FITC Annexin V Apoptosis Detection Kit I, etc., the protein expression, DNA synthesis, apoptosis, cell invasion, colony formation, tumor formation, and cell differentiation of MCF10A cells can be evaluated in detail [18].
The MCF10A cell line series provides a unique opportunity to study malignant progression in a molecular defined manner within a common cellular context. Compared with MCF10A cells, MCF-10AT cells form a multi papillary structure in three-dimensional rBM, while MCF-10CA1a cells form large highly proliferative cell clusters in three-dimensional rBM, with changes in tissue structure and no papillary structures [4]. This indicates that pre malignant cells have lost normal proliferation control, but retained normal cell polarity, intercellular and cell basement membrane adhesion. Completely malignant cells have lost the ability to undergo normal nipple morphology.
Research shows that MCF10A cell line shows the increase of cell proliferation and migration ability, as well as the transformation of epithelial to mesenchymal markers in the breast cancer progression model [14]. In addition, after knocking down the Mettl3 gene using CRISPR/Cas9 technology, the mRNA level of one candidate gene significantly increased in the MCF10A cell line, while the migration ability of another candidate gene significantly decreased [14]. These results further confirmed the validity of MCF10A cell line as a research model for breast cancer.
What are the applications of MCF 10A cells in simulating breast development and disease processes?
The application of MCF10A cells in simulating breast development and disease processes is very extensive, mainly reflected in the following aspects:
The MCF10A cell line is widely used to study the developmental process of breast tissue. By constructing 3D models, including duct and acinar structures, which gradually form during the cultivation process, the developmental process of breast tissue was simulated [20]. This model can express breast gland markers and display the developmental process of breast glands [20].
MCF10A cells and their sub lines (such as MCF10AnoT, MCF10AT, MCF10DCIS and MCF10CA1) are used to study different stages of breast cancer. For example, MCF10A cells represent normal breast epithelial cells, while MCF10AT cells mimic the early stages of breast proliferative diseases through HRAS gene transformation [22,23]. In addition, MCF10CA1h and MCF10CA1a cells represent different stages of breast cancer progression, including malignant tumors and distant metastasis [24].
MCF10A cell line is also used to study the signal pathway of breast cancer cells. For example, through phosphorylated peptide enrichment and LC-MS analysis, researchers can study the dynamic changes of key signal pathways such as MAPK-AKT-mTOR in breast cancer [26].
MCF10A cells contain a subpopulation expressing stem/progenitor cell markers and can be characterized for their self-renewal ability through blastocyst culture methods [2]. Although its blastocyst formation rate is low, the characteristics of breast stem cells can be further studied by sorting specific subpopulations of markers [2].
Due to the high similarity between MCF10A cells and normal human breast epithelial cells, they are widely used in toxicity studies. For example, studying the effects of bisphenol A and its analogues on MCF10A cells [25].
MCF10 series of cell lines also played an important role in the study of heterogeneity in spontaneous ectopic breast cancer and breast ductal carcinoma. These models provide important tools for early diagnosis, treatment and prognosis of breast cancer [27].
What are the different views and research results in the academic community regarding the controversy over MCF 10A cells?
There are different views and research results in the academic community regarding the controversy over MCF 10A cells. Here are several main research directions and findings:
1. Tumor formation ability:
Tumor forming ability of MCF10A cells: Studies have shown that MCF10A cells treated with random mutations (such as MCF10A β cells) can form large and continuously growing tumors in nude mice [21]. However, another study indicated that even in immunocompromised mice, MCF10A cells treated with Cd ^ 2+failed to form tumors [7]. This indicates that MCF10A cells may exhibit different tumor forming abilities under different conditions.
2. Gene expression and genomic changes:
Gene expression pattern: Studies have found that MCF10A cells have a basal like gene expression pattern and are ER and PR negative [7]. In addition, MCF10A cells showed clear intercellular connections and basement membrane deposition in three-dimensional culture, but their proliferation ability was limited [4].
Genomic changes: Copy number analysis of Illumina Human Omni 2.5M SNP array data revealed significant differences in gene amplification and loss between MCF10A cells and MCF10CA1a cells [28]. For example, in the MCF10A cell line, there was an increase of 2374 protein coding genes and a loss of 461 genes, while in the MCF10CA1a cell line, there was an increase of 1433 genes and a loss of 33 genes [28].
3. Drug sensitivity and chemotherapy response:
Chemotherapy sensitivity: Studies have shown that MCF10A and MCF10CA1a cell lines have the same sensitivity to Docetaxel/paclitaxel chemotherapy, regardless of the presence of EGFR mutations [28]. This indicates that MCF10A cells have a certain sensitivity to certain chemotherapy drugs.
4. Cell cycle regulation:
Cell cycle regulation: The study investigated the behavior of MCF10A cells under different resting conditions through phase contrast and fluorescence exposure image analysis, as well as measurement of cell cycle related proteins. The results showed that when the level of p21 protein was high, the intermediate time of the cell cycle (IMT) was longer, indicating that p21 plays an important role in regulating the cell cycle [33].
5. Effects of contrast agents on ultrasound imaging:
The effect of ultrasound microbubbles on MCF10A cells: Studies have shown that the improved ultrasound microbubble CNC SonoVue has little effect on the proliferation of MCF10A cells, indicating that its introduction does not increase cytotoxicity [29,32].
The controversy surrounding MCF 10A cells mainly focuses on their tumor forming ability, gene expression and genomic changes, drug sensitivity, cell cycle regulation, and the effects of ultrasound imaging contrast agents.
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