Limits...
Evaluation of MCF10A as a Reliable Model for Normal Human Mammary Epithelial Cells.

Qu Y, Han B, Yu Y, Yao W, Bose S, Karlan BY, Giuliano AE, Cui X - PLoS ONE (2015)

Bottom Line: However, there is limited knowledge about whether MCF10A cells reliably represent normal human mammary cells.When grown in suspension culture, MCF10A cells showed low mammosphere-forming ability.Our results raise a question as to whether the commonly used MCF10A cell line is a suitable model for human mammary cell studies.

View Article: PubMed Central - PubMed

Affiliation: Department of Surgery, Samuel Oschin Comprehensive Cancer Institute, Cedars-Sinai Medical Center, Los Angeles, California, United States of America.

ABSTRACT
Breast cancer is the most common cancer in women and a leading cause of cancer-related deaths for women worldwide. Various cell models have been developed to study breast cancer tumorigenesis, metastasis, and drug sensitivity. The MCF10A human mammary epithelial cell line is a widely used in vitro model for studying normal breast cell function and transformation. However, there is limited knowledge about whether MCF10A cells reliably represent normal human mammary cells. MCF10A cells were grown in monolayer, suspension (mammosphere culture), three-dimensional (3D) "on-top" Matrigel, 3D "cell-embedded" Matrigel, or mixed Matrigel/collagen I gel. Suspension culture was performed with the MammoCult medium and low-attachment culture plates. Cells grown in 3D culture were fixed and subjected to either immunofluorescence staining or embedding and sectioning followed by immunohistochemistry and immunofluorescence staining. Cells or slides were stained for protein markers commonly used to identify mammary progenitor and epithelial cells. MCF10A cells expressed markers representing luminal, basal, and progenitor phenotypes in two-dimensional (2D) culture. When grown in suspension culture, MCF10A cells showed low mammosphere-forming ability. Cells in mammospheres and 3D culture expressed both luminal and basal markers. Surprisingly, the acinar structure formed by MCF10A cells in 3D culture was positive for both basal markers and the milk proteins β-casein and α-lactalbumin. MCF10A cells exhibit a unique differentiated phenotype in 3D culture which may not exist or be rare in normal human breast tissue. Our results raise a question as to whether the commonly used MCF10A cell line is a suitable model for human mammary cell studies.

No MeSH data available.


Related in: MedlinePlus

MCF10A cells in “on-top” of Matrigel.MCF10A cells were cultured in “on-top” of Matrigel for 14 days. 3D cultures were fixed, embedded and cut into sections. Immunohistochemistry staining of basal markers (B), luminal markers (C), and breast tissue-specific markers (D) were performed. Bars: 50μm. Original magnification: ×600.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4493126&req=5

pone.0131285.g005: MCF10A cells in “on-top” of Matrigel.MCF10A cells were cultured in “on-top” of Matrigel for 14 days. 3D cultures were fixed, embedded and cut into sections. Immunohistochemistry staining of basal markers (B), luminal markers (C), and breast tissue-specific markers (D) were performed. Bars: 50μm. Original magnification: ×600.

Mentions: We then tested the expression of markers using continuous thin sections from fixed acini. Similar to mammospheres, MCF10A spheres in “on-top” Matrigel did not express stem-like markers such as CD49f (data not shown) but expressed the basal markers CK14 and SMA (Fig 5A). However, P63 was not detected (Fig 5A). These cells also expressed luminal CK18 and CK8 (Fig 5B) as well as breast specific markers CSN2 and LALBA (Fig 5C), suggesting that luminal-, basal- and breast tissue-specific- markers were co-expressed in the cells. Of note, immunofluorescence staining of mammosphere thin sections showed that MCF10A cells did not express the stem/progenitor markers CD49f, CD44, and ALDH1A3 at day 7 and day 10 of Matrigel culture (S3 Fig), suggesting that Matrigel may induce the loss of stem/progenitor marker expression in MCF10A cells. To further test whether these spheres were positive for both luminal- and basal-markers, we performed immunofluorescence co-staining. As shown in S4A Fig, both CK18 and CK14 were positive in the acinar structure and thus co-expressed in the same cells. In contrast, the majority of luminal cells were CK18+ and basal cells were CK14+ in the normal human mammary gland tissue (S4B Fig). More importantly, cells in the duct did not co-express CK14 and CSN2 (S4C Fig). MCF10A cells embedded in Matrigel showed similar results as the “on-top” culture (data not shown). We also found that co-culture of mammary fibroblast cells did not alter MCF10A cell differentiation profiles or luminal, basal, breast tissue marker expression pattern in Matrigel 3D culture (data not shown). These data suggest that MCF10A cells are able to form a polarized spheroid structure but may not represent normal human breast epithelial cells or may represent a rare mammary epithelial cell type.


Evaluation of MCF10A as a Reliable Model for Normal Human Mammary Epithelial Cells.

Qu Y, Han B, Yu Y, Yao W, Bose S, Karlan BY, Giuliano AE, Cui X - PLoS ONE (2015)

MCF10A cells in “on-top” of Matrigel.MCF10A cells were cultured in “on-top” of Matrigel for 14 days. 3D cultures were fixed, embedded and cut into sections. Immunohistochemistry staining of basal markers (B), luminal markers (C), and breast tissue-specific markers (D) were performed. Bars: 50μm. Original magnification: ×600.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4493126&req=5

pone.0131285.g005: MCF10A cells in “on-top” of Matrigel.MCF10A cells were cultured in “on-top” of Matrigel for 14 days. 3D cultures were fixed, embedded and cut into sections. Immunohistochemistry staining of basal markers (B), luminal markers (C), and breast tissue-specific markers (D) were performed. Bars: 50μm. Original magnification: ×600.
Mentions: We then tested the expression of markers using continuous thin sections from fixed acini. Similar to mammospheres, MCF10A spheres in “on-top” Matrigel did not express stem-like markers such as CD49f (data not shown) but expressed the basal markers CK14 and SMA (Fig 5A). However, P63 was not detected (Fig 5A). These cells also expressed luminal CK18 and CK8 (Fig 5B) as well as breast specific markers CSN2 and LALBA (Fig 5C), suggesting that luminal-, basal- and breast tissue-specific- markers were co-expressed in the cells. Of note, immunofluorescence staining of mammosphere thin sections showed that MCF10A cells did not express the stem/progenitor markers CD49f, CD44, and ALDH1A3 at day 7 and day 10 of Matrigel culture (S3 Fig), suggesting that Matrigel may induce the loss of stem/progenitor marker expression in MCF10A cells. To further test whether these spheres were positive for both luminal- and basal-markers, we performed immunofluorescence co-staining. As shown in S4A Fig, both CK18 and CK14 were positive in the acinar structure and thus co-expressed in the same cells. In contrast, the majority of luminal cells were CK18+ and basal cells were CK14+ in the normal human mammary gland tissue (S4B Fig). More importantly, cells in the duct did not co-express CK14 and CSN2 (S4C Fig). MCF10A cells embedded in Matrigel showed similar results as the “on-top” culture (data not shown). We also found that co-culture of mammary fibroblast cells did not alter MCF10A cell differentiation profiles or luminal, basal, breast tissue marker expression pattern in Matrigel 3D culture (data not shown). These data suggest that MCF10A cells are able to form a polarized spheroid structure but may not represent normal human breast epithelial cells or may represent a rare mammary epithelial cell type.

Bottom Line: However, there is limited knowledge about whether MCF10A cells reliably represent normal human mammary cells.When grown in suspension culture, MCF10A cells showed low mammosphere-forming ability.Our results raise a question as to whether the commonly used MCF10A cell line is a suitable model for human mammary cell studies.

View Article: PubMed Central - PubMed

Affiliation: Department of Surgery, Samuel Oschin Comprehensive Cancer Institute, Cedars-Sinai Medical Center, Los Angeles, California, United States of America.

ABSTRACT
Breast cancer is the most common cancer in women and a leading cause of cancer-related deaths for women worldwide. Various cell models have been developed to study breast cancer tumorigenesis, metastasis, and drug sensitivity. The MCF10A human mammary epithelial cell line is a widely used in vitro model for studying normal breast cell function and transformation. However, there is limited knowledge about whether MCF10A cells reliably represent normal human mammary cells. MCF10A cells were grown in monolayer, suspension (mammosphere culture), three-dimensional (3D) "on-top" Matrigel, 3D "cell-embedded" Matrigel, or mixed Matrigel/collagen I gel. Suspension culture was performed with the MammoCult medium and low-attachment culture plates. Cells grown in 3D culture were fixed and subjected to either immunofluorescence staining or embedding and sectioning followed by immunohistochemistry and immunofluorescence staining. Cells or slides were stained for protein markers commonly used to identify mammary progenitor and epithelial cells. MCF10A cells expressed markers representing luminal, basal, and progenitor phenotypes in two-dimensional (2D) culture. When grown in suspension culture, MCF10A cells showed low mammosphere-forming ability. Cells in mammospheres and 3D culture expressed both luminal and basal markers. Surprisingly, the acinar structure formed by MCF10A cells in 3D culture was positive for both basal markers and the milk proteins β-casein and α-lactalbumin. MCF10A cells exhibit a unique differentiated phenotype in 3D culture which may not exist or be rare in normal human breast tissue. Our results raise a question as to whether the commonly used MCF10A cell line is a suitable model for human mammary cell studies.

No MeSH data available.


Related in: MedlinePlus