Can widely used cell type markers predict the suitability of immortalized or primary mammary epithelial cell models?

BACKGROUND: Mammary cell cultures are convenient tools for in vitro studies of mammary gland biology. However, the heterogeneity of mammary cell types, e.g., glandular milk secretory epithelial or myoepithelial cells, often complicates the interpretation of cell-based data. The present study was und...

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Autores principales:	Ontsouka,Edgar Corneille, Bertschi,Janique Sabina, Huang,Xiao, Lüthi,Michael, Müller,Stefan, Albrecht,Christiane
Lenguaje:	English
Publicado:	Sociedad de Biología de Chile 2016
Materias:	Cell characterization Flow cytometry Gene expression Lactation marker Mammalian Mammary epithelial cell
Acceso en línea:	http://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0716-97602016000100001
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spelling	oai:scielo:S0716-976020160001000012016-12-01Can widely used cell type markers predict the suitability of immortalized or primary mammary epithelial cell models?Ontsouka,Edgar CorneilleBertschi,Janique SabinaHuang,XiaoLüthi,MichaelMüller,StefanAlbrecht,Christiane Cell characterization Flow cytometry Gene expression Lactation marker Mammalian Mammary epithelial cell BACKGROUND: Mammary cell cultures are convenient tools for in vitro studies of mammary gland biology. However, the heterogeneity of mammary cell types, e.g., glandular milk secretory epithelial or myoepithelial cells, often complicates the interpretation of cell-based data. The present study was undertaken to determine the relevance of bovine primary mammary epithelial cells isolated from American Holstein (bMEC US) or Swiss Holstein-Friesian (bMEC CH) cows, and of primary bovine mammary alveolar epithelial cells stably transfected with simian virus-40 (SV-40) large T-antigen (MAC-T) for in vitro analyses. This was evaluated by testing their expression pattern of cytokeratin (CK) 7, 18, 19, vimentin, and α-smooth muscle actin (α-SMA. RESULTS: The expression of the listed markers was assessed using real-time quantitative PCR, flow cytometry and immunofluorescence microscopy. Characteristic markers of the mesenchymal (vimentin), myoepithelial (α-SMA) and glandular secretory cells (CKs) showed differential expression among the studied cell cultures, partly depending on the analytical method used. The relative mRNA expression of vimentin, CK7 and CK19, respectively, was lower (P < 0.05) in immortalized than in primary mammary cell cultures. The stain index (based on flow cytometry) of CK7 and CK19 protein was lower (P < 0.05) in MAC-T than in bMECs, while the expression of α-SMA and CK18 showed an inverse pattern. Immunofluorescence microscopy analysis mostly confirmed the mRNA data, while partly disagreed with flow cytometry data (e.g., vimentin level in MAC-T). The differential expression of CK7 and CK19 allowed discriminating between immortal and primary mammary cultures. CONCLUSIONS: The expression of the selected widely used cell type markers in primary and immortalized MEC cells did not allow a clear preference between these two cell models for in vitro analyses studying aspects of milk composition. All tested cell models exhibited to a variable degree epithelial and mesenchymal features. Thus, based on their characterization with widely used cell markers, none of these cultures represent an unequivocal alveolar mammary epithelial cell model. For choosing the appropriate in vitro model additional properties such as the expression profile of specific proteins of interest (e.g., transporter proteins) should equally be taken into account.info:eu-repo/semantics/openAccessSociedad de Biología de ChileBiological Research v.49 20162016-01-01text/htmlhttp://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0716-97602016000100001en10.1186/s40659-015-0063-2
institution	Scielo Chile
collection	Scielo Chile
language	English
topic	Cell characterization Flow cytometry Gene expression Lactation marker Mammalian Mammary epithelial cell
spellingShingle	Cell characterization Flow cytometry Gene expression Lactation marker Mammalian Mammary epithelial cell Ontsouka,Edgar Corneille Bertschi,Janique Sabina Huang,Xiao Lüthi,Michael Müller,Stefan Albrecht,Christiane Can widely used cell type markers predict the suitability of immortalized or primary mammary epithelial cell models?
description	BACKGROUND: Mammary cell cultures are convenient tools for in vitro studies of mammary gland biology. However, the heterogeneity of mammary cell types, e.g., glandular milk secretory epithelial or myoepithelial cells, often complicates the interpretation of cell-based data. The present study was undertaken to determine the relevance of bovine primary mammary epithelial cells isolated from American Holstein (bMEC US) or Swiss Holstein-Friesian (bMEC CH) cows, and of primary bovine mammary alveolar epithelial cells stably transfected with simian virus-40 (SV-40) large T-antigen (MAC-T) for in vitro analyses. This was evaluated by testing their expression pattern of cytokeratin (CK) 7, 18, 19, vimentin, and α-smooth muscle actin (α-SMA. RESULTS: The expression of the listed markers was assessed using real-time quantitative PCR, flow cytometry and immunofluorescence microscopy. Characteristic markers of the mesenchymal (vimentin), myoepithelial (α-SMA) and glandular secretory cells (CKs) showed differential expression among the studied cell cultures, partly depending on the analytical method used. The relative mRNA expression of vimentin, CK7 and CK19, respectively, was lower (P < 0.05) in immortalized than in primary mammary cell cultures. The stain index (based on flow cytometry) of CK7 and CK19 protein was lower (P < 0.05) in MAC-T than in bMECs, while the expression of α-SMA and CK18 showed an inverse pattern. Immunofluorescence microscopy analysis mostly confirmed the mRNA data, while partly disagreed with flow cytometry data (e.g., vimentin level in MAC-T). The differential expression of CK7 and CK19 allowed discriminating between immortal and primary mammary cultures. CONCLUSIONS: The expression of the selected widely used cell type markers in primary and immortalized MEC cells did not allow a clear preference between these two cell models for in vitro analyses studying aspects of milk composition. All tested cell models exhibited to a variable degree epithelial and mesenchymal features. Thus, based on their characterization with widely used cell markers, none of these cultures represent an unequivocal alveolar mammary epithelial cell model. For choosing the appropriate in vitro model additional properties such as the expression profile of specific proteins of interest (e.g., transporter proteins) should equally be taken into account.
author	Ontsouka,Edgar Corneille Bertschi,Janique Sabina Huang,Xiao Lüthi,Michael Müller,Stefan Albrecht,Christiane
author_facet	Ontsouka,Edgar Corneille Bertschi,Janique Sabina Huang,Xiao Lüthi,Michael Müller,Stefan Albrecht,Christiane
author_sort	Ontsouka,Edgar Corneille
title	Can widely used cell type markers predict the suitability of immortalized or primary mammary epithelial cell models?
title_short	Can widely used cell type markers predict the suitability of immortalized or primary mammary epithelial cell models?
title_full	Can widely used cell type markers predict the suitability of immortalized or primary mammary epithelial cell models?
title_fullStr	Can widely used cell type markers predict the suitability of immortalized or primary mammary epithelial cell models?
title_full_unstemmed	Can widely used cell type markers predict the suitability of immortalized or primary mammary epithelial cell models?
title_sort	can widely used cell type markers predict the suitability of immortalized or primary mammary epithelial cell models?
publisher	Sociedad de Biología de Chile
publishDate	2016
url	http://www.scielo.cl/scielo.php?script=sci_arttext&pid=S0716-97602016000100001
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Can widely used cell type markers predict the suitability of immortalized or primary mammary epithelial cell models?

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