title: Can widely used cell type markers predict the suitability of immortalized or primary mammary epithelial cell models?
abstract: 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 (bMECUS) or Swiss Holstein-Friesian (bMECCH) 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.
authors.affiliation: Faculty of Medicine, Institute of Biochemistry and Molecular Medicine, University of Bern, Buehlstrasse 28, 3012, Bern, Switzerland. corneille.ontsouka@ibmm.unibe.ch.|Swiss National Center of Competence in Research, NCCR TransCure, University of Bern, Bern, Switzerland. corneille.ontsouka@ibmm.unibe.ch.
authors.forename: Edgar Corneille
authors.initials: EC
authors.lastname: Ontsouka
authors.affiliation: Faculty of Medicine, Institute of Biochemistry and Molecular Medicine, University of Bern, Buehlstrasse 28, 3012, Bern, Switzerland. janique.21@hotmail.com.
authors.forename: Janique Sabina
authors.initials: JS
authors.lastname: Bertschi
authors.affiliation: Faculty of Medicine, Institute of Biochemistry and Molecular Medicine, University of Bern, Buehlstrasse 28, 3012, Bern, Switzerland. xiao.huang@ibmm.unibe.ch.
authors.forename: Xiao
authors.initials: X
authors.lastname: Huang
authors.affiliation: Faculty of Medicine, Institute of Biochemistry and Molecular Medicine, University of Bern, Buehlstrasse 28, 3012, Bern, Switzerland. michael.luethi@ibmm.unibe.ch.|Swiss National Center of Competence in Research, NCCR TransCure, University of Bern, Bern, Switzerland. michael.luethi@ibmm.unibe.ch.
authors.forename: Michael
authors.initials: M
authors.lastname: Lüthi
authors.affiliation: Department of Clinical Research, Faculty of Medicine, University of Bern, 3010, Bern, Switzerland. stefan.mueller@dkf.unibe.ch.
authors.forename: Stefan
authors.initials: S
authors.lastname: Müller
authors.affiliation: Faculty of Medicine, Institute of Biochemistry and Molecular Medicine, University of Bern, Buehlstrasse 28, 3012, Bern, Switzerland. christiane.albrecht@ibmm.unibe.ch.|Swiss National Center of Competence in Research, NCCR TransCure, University of Bern, Bern, Switzerland. christiane.albrecht@ibmm.unibe.ch.
authors.forename: Christiane
authors.initials: C
authors.lastname: Albrecht
mesh_terms: D000199:Actins / Q000032:analysis*; D000704:Analysis of Variance; D000818:Animals; D000957:Antigens, Viral, Tumor; D002417:Cattle; D002460:Cell Line; D002478:Cells, Cultured; D004847:Epithelial Cells / Q000737:chemistry / Q000166:cytology*; D005260:Female; D005434:Flow Cytometry / Q000379:methods; D007633:Keratins / Q000032:analysis*; D008321:Mammary Glands, Animal / Q000737:chemistry / Q000166:cytology*; D008856:Microscopy, Fluorescence / Q000379:methods; D061251:Primary Cell Culture; D060888:Real-Time Polymerase Chain Reaction; D013539:Simian virus 40; D014746:Vimentin / Q000032:analysis*
chemical_list: D000199:Actins; D000957:Antigens, Viral, Tumor; D014746:Vimentin; D007633:Keratins