title: The role of 3D microenvironmental organization in MCF-7 epithelial-mesenchymal transition after 7 culture days.
abstract: We present a multi-technique study on in vitro epithelial-mesenchymal transition (EMT) in human MCF-7 cells cultured on electrospun scaffolds of poly(l-lactic acid) (PLA), with random and aligned fiber orientations. Our aim is to investigate the morphological and genetic characteristics induced by extracellular matrix in tumor cells cultured in different 3D environments, and at different time points. Cell vitality was assessed with AlamarBlue at days 1, 3, 5 and 7. Scanning electron microscopy was performed at culture days 3 and 7. Immunohistochemistry (for E-cadherin, β-catenin, cytokeratins, nucleophosmin, tubulin, Ki-67 and vimentin), immunofluorescence (for F-actin) western blot (for E-cadherin, β-catenin and vimentin) and transmission electron microscopy were carried out at day 7. An EMT gene array followed by PCR analysis confirmed the regulation of selected genes. At day 7, scanning electron microscopy on aligned-PLA revealed spindle-shaped cells gathered in buds and ribbon-like structures, with a higher nucleolar/nuclear ratio and a loss in E-cadherin and β-catenin at immunohistochemistry and western blot. An up-regulation of SMAD2, TGF-β2, TFPI2 and SOX10 was found in aligned-PLA compared to random-PLA cultured cells. The topography of the extracellular matrix has a role in tumor EMT, and a more aggressive phenotype characterizes MCF-7 cells cultured on aligned-PLA scaffold.
authors.affiliation: Pathology Unit, Department of Haematology, Oncology and Clinical Pathology, S. Orsola-Malpighi Hospital, Bologna University, Italy.
authors.forename: Laura
authors.initials: L
authors.lastname: Foroni
authors.forename: Francesco
authors.initials: F
authors.lastname: Vasuri
authors.forename: Sabrina
authors.initials: S
authors.lastname: Valente
authors.forename: Chiara
authors.initials: C
authors.lastname: Gualandi
authors.forename: Maria Letizia
authors.initials: ML
authors.lastname: Focarete
authors.forename: Giacomo
authors.initials: G
authors.lastname: Caprara
authors.forename: Mariastella
authors.initials: M
authors.lastname: Scandola
authors.forename: Antonia
authors.initials: A
authors.lastname: D'Errico-Grigioni
authors.forename: Gianandrea
authors.initials: G
authors.lastname: Pasquinelli
mesh_terms: D000199:Actins / Q000235:genetics / Q000378:metabolism; D015703:Antigens, CD; D015153:Blotting, Western; D015820:Cadherins / Q000235:genetics / Q000378:metabolism; D002448:Cell Adhesion; D048430:Cell Shape; D058750:Epithelial-Mesenchymal Transition; D005109:Extracellular Matrix / Q000378:metabolism*; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D006023:Glycoproteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D019394:Ki-67 Antigen / Q000235:genetics / Q000378:metabolism; D019344:Lactic Acid / Q000378:metabolism; D061986:MCF-7 Cells; D008855:Microscopy, Electron, Scanning; D046529:Microscopy, Electron, Transmission; D020411:Oligonucleotide Array Sequence Analysis; D010078:Oxazines; D010641:Phenotype; D051899:Smad2 Protein / Q000235:genetics / Q000378:metabolism; D013997:Time Factors; D054457:Tissue Scaffolds; D053781:Transforming Growth Factor beta2 / Q000235:genetics / Q000378:metabolism; D059016:Tumor Microenvironment; D014746:Vimentin / Q000235:genetics / Q000378:metabolism; D014966:Xanthenes; D051176:beta Catenin / Q000235:genetics / Q000378:metabolism
chemical_list: D000199:Actins; D015703:Antigens, CD; C513384:CDH1 protein, human; C495270:CTNNB1 protein, human; D015820:Cadherins; D006023:Glycoproteins; D019394:Ki-67 Antigen; D010078:Oxazines; C497782:SMAD2 protein, human; D051899:Smad2 Protein; D053781:Transforming Growth Factor beta2; D014746:Vimentin; D014966:Xanthenes; D051176:beta Catenin; C021240:tissue-factor-pathway inhibitor 2; C005843:resazurin; D019344:Lactic Acid