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Increasingly transformed MCF-10A cells have a progressively tumor-like phenotype in three-dimensional basement membrane culture.

Imbalzano KM, Tatarkova I, Imbalzano AN, Nickerson JA - Cancer Cell Int. (2009)

Bottom Line: The pre-malignant MCF-10AT cells and malignant MCF-10CA1a lines were sequentially derived from the MCF-10A parental cell line first by expression of a constitutively active T24 H-Ras generating the MCF-10AT cell line.Features of tissue morphogenesis were assessed, including proliferation, basement membrane formation, polarization of alpha-6 beta-4 integrin to the basement membrane, formation of cell:cell junctions, and apoptosis for luminal clearance.The matched series of normal MCF-10A, pre-malignant MCF-10AT, and malignant MCF-10CA1a cells offers a unique opportunity to study the mechanisms of malignant progression both in a three-dimensional microenvironment and in the same cell background.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Cell Biology, University of Massachusetts Medical School, Worcester, MA 01655, USA. karen.imbalzano@umassmed.edu

ABSTRACT

Background: MCF-10A cells are near diploid and normal human mammary epithelial cells. In three-dimensional reconstituted basement membrane culture, they undergo a well-defined program of proliferation, differentiation, and growth arrest, forming acinar structures that recapitulate many aspects of mammary architecture in vivo. The pre-malignant MCF-10AT cells and malignant MCF-10CA1a lines were sequentially derived from the MCF-10A parental cell line first by expression of a constitutively active T24 H-Ras generating the MCF-10AT cell line. This was followed by repeated selection for increasingly aggressive tumor formation from cells recovered from xenograft tumors in immuno-compromised mice, generating the MCF-10CA1a cell line. When inoculated subcutaneously into the flanks of immuno-compromised mice, MCF-10AT cells occasionally form tumors, whereas MCF-10CA1a cells invariably form tumors with a shorter latency than MCF-10AT derived tumors.

Results: MCF-10AT cells grown in three-dimensional basement membrane culture form complex multi-acinar structures that produce a basement membrane but undergo delayed cell cycle arrest and have incomplete luminal development. MCF-10CA1a cells grown in three-dimensional basement membrane culture form large, hyper-proliferative masses, that retain few characteristics of MCF10A acini and more closely resemble tumors.

Conclusion: Here we report on the growth and differentiation properties of these three matched cell lines in three-dimensional basement membrane culture. Features of tissue morphogenesis were assessed, including proliferation, basement membrane formation, polarization of alpha-6 beta-4 integrin to the basement membrane, formation of cell:cell junctions, and apoptosis for luminal clearance. The matched series of normal MCF-10A, pre-malignant MCF-10AT, and malignant MCF-10CA1a cells offers a unique opportunity to study the mechanisms of malignant progression both in a three-dimensional microenvironment and in the same cell background.

No MeSH data available.


Related in: MedlinePlus

Morphology observed for MCF-10A, MCF-10AT, and MCF-10CA1a cells grown in rBM for 20 days. Phase contrast images of normal MCF-10A cells were cultured in overlay rBM for 2, 4, 8, 12, 16, and 20 days (top, A to F). Transformed MCF-10AT cells (middle, G to L) and malignant MCF-10CA1a cells (bottom, M to R) were cultured in three-dimensions at the same time points. Scale bar, 100 μm. Normal MCF-10A cells increase in size to day 8 and form polarized spherical acini. Transformed MCF-10AT cells increase in size for 20 days, retain polarity, and form multi-acinar structures. Malignant MCF-10CA1a cells continue to proliferate throughout the 20 days and form large, complex masses.
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Figure 2: Morphology observed for MCF-10A, MCF-10AT, and MCF-10CA1a cells grown in rBM for 20 days. Phase contrast images of normal MCF-10A cells were cultured in overlay rBM for 2, 4, 8, 12, 16, and 20 days (top, A to F). Transformed MCF-10AT cells (middle, G to L) and malignant MCF-10CA1a cells (bottom, M to R) were cultured in three-dimensions at the same time points. Scale bar, 100 μm. Normal MCF-10A cells increase in size to day 8 and form polarized spherical acini. Transformed MCF-10AT cells increase in size for 20 days, retain polarity, and form multi-acinar structures. Malignant MCF-10CA1a cells continue to proliferate throughout the 20 days and form large, complex masses.

Mentions: Parental MCF-10A, pre-malignant MCF-10AT, and malignant MCF-10CA1a cell lines were cultured in rBM. In this procedure, cells were plated on a layer of rBM with an overlay created by adding rBM to the culture medium [9]. After two days, cultures of all three cell lines formed similarly-sized spherical masses of cells (Figure 2A, G, M). After 4 days, the masses of MCF-10AT and MCF-10CA1a cells were larger than the masses of MCF-10A cells (Figure 2B, H, N), an indicator of increased proliferation. After 8 days in three-dimensional culture, the MCF-10AT structures had an acinar appearance but these acini were larger than those for the parental MCF-10A cells and were more irregular in shape (Figure 2C and 2I).


Increasingly transformed MCF-10A cells have a progressively tumor-like phenotype in three-dimensional basement membrane culture.

Imbalzano KM, Tatarkova I, Imbalzano AN, Nickerson JA - Cancer Cell Int. (2009)

Morphology observed for MCF-10A, MCF-10AT, and MCF-10CA1a cells grown in rBM for 20 days. Phase contrast images of normal MCF-10A cells were cultured in overlay rBM for 2, 4, 8, 12, 16, and 20 days (top, A to F). Transformed MCF-10AT cells (middle, G to L) and malignant MCF-10CA1a cells (bottom, M to R) were cultured in three-dimensions at the same time points. Scale bar, 100 μm. Normal MCF-10A cells increase in size to day 8 and form polarized spherical acini. Transformed MCF-10AT cells increase in size for 20 days, retain polarity, and form multi-acinar structures. Malignant MCF-10CA1a cells continue to proliferate throughout the 20 days and form large, complex masses.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2666639&req=5

Figure 2: Morphology observed for MCF-10A, MCF-10AT, and MCF-10CA1a cells grown in rBM for 20 days. Phase contrast images of normal MCF-10A cells were cultured in overlay rBM for 2, 4, 8, 12, 16, and 20 days (top, A to F). Transformed MCF-10AT cells (middle, G to L) and malignant MCF-10CA1a cells (bottom, M to R) were cultured in three-dimensions at the same time points. Scale bar, 100 μm. Normal MCF-10A cells increase in size to day 8 and form polarized spherical acini. Transformed MCF-10AT cells increase in size for 20 days, retain polarity, and form multi-acinar structures. Malignant MCF-10CA1a cells continue to proliferate throughout the 20 days and form large, complex masses.
Mentions: Parental MCF-10A, pre-malignant MCF-10AT, and malignant MCF-10CA1a cell lines were cultured in rBM. In this procedure, cells were plated on a layer of rBM with an overlay created by adding rBM to the culture medium [9]. After two days, cultures of all three cell lines formed similarly-sized spherical masses of cells (Figure 2A, G, M). After 4 days, the masses of MCF-10AT and MCF-10CA1a cells were larger than the masses of MCF-10A cells (Figure 2B, H, N), an indicator of increased proliferation. After 8 days in three-dimensional culture, the MCF-10AT structures had an acinar appearance but these acini were larger than those for the parental MCF-10A cells and were more irregular in shape (Figure 2C and 2I).

Bottom Line: The pre-malignant MCF-10AT cells and malignant MCF-10CA1a lines were sequentially derived from the MCF-10A parental cell line first by expression of a constitutively active T24 H-Ras generating the MCF-10AT cell line.Features of tissue morphogenesis were assessed, including proliferation, basement membrane formation, polarization of alpha-6 beta-4 integrin to the basement membrane, formation of cell:cell junctions, and apoptosis for luminal clearance.The matched series of normal MCF-10A, pre-malignant MCF-10AT, and malignant MCF-10CA1a cells offers a unique opportunity to study the mechanisms of malignant progression both in a three-dimensional microenvironment and in the same cell background.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Cell Biology, University of Massachusetts Medical School, Worcester, MA 01655, USA. karen.imbalzano@umassmed.edu

ABSTRACT

Background: MCF-10A cells are near diploid and normal human mammary epithelial cells. In three-dimensional reconstituted basement membrane culture, they undergo a well-defined program of proliferation, differentiation, and growth arrest, forming acinar structures that recapitulate many aspects of mammary architecture in vivo. The pre-malignant MCF-10AT cells and malignant MCF-10CA1a lines were sequentially derived from the MCF-10A parental cell line first by expression of a constitutively active T24 H-Ras generating the MCF-10AT cell line. This was followed by repeated selection for increasingly aggressive tumor formation from cells recovered from xenograft tumors in immuno-compromised mice, generating the MCF-10CA1a cell line. When inoculated subcutaneously into the flanks of immuno-compromised mice, MCF-10AT cells occasionally form tumors, whereas MCF-10CA1a cells invariably form tumors with a shorter latency than MCF-10AT derived tumors.

Results: MCF-10AT cells grown in three-dimensional basement membrane culture form complex multi-acinar structures that produce a basement membrane but undergo delayed cell cycle arrest and have incomplete luminal development. MCF-10CA1a cells grown in three-dimensional basement membrane culture form large, hyper-proliferative masses, that retain few characteristics of MCF10A acini and more closely resemble tumors.

Conclusion: Here we report on the growth and differentiation properties of these three matched cell lines in three-dimensional basement membrane culture. Features of tissue morphogenesis were assessed, including proliferation, basement membrane formation, polarization of alpha-6 beta-4 integrin to the basement membrane, formation of cell:cell junctions, and apoptosis for luminal clearance. The matched series of normal MCF-10A, pre-malignant MCF-10AT, and malignant MCF-10CA1a cells offers a unique opportunity to study the mechanisms of malignant progression both in a three-dimensional microenvironment and in the same cell background.

No MeSH data available.


Related in: MedlinePlus