A 3D in vitro model to explore the inter-conversion between epithelial and mesenchymal states during EMT and its reversion

Sci Rep. 2016 Jun 3:6:27072. doi: 10.1038/srep27072.

Abstract

Epithelial-to-mesenchymal transitions (EMT) are strongly implicated in cancer dissemination. Intermediate states, arising from inter-conversion between epithelial (E) and mesenchymal (M) states, are characterized by phenotypic heterogeneity combining E and M features and increased plasticity. Hybrid EMT states are highly relevant in metastatic contexts, but have been largely neglected, partially due to the lack of physiologically-relevant 3D platforms to study them. Here we propose a new in vitro model, combining mammary E cells with a bioengineered 3D matrix, to explore phenotypic and functional properties of cells in transition between E and M states. Optimized alginate-based 3D matrices provided adequate 3D microenvironments, where normal epithelial morphogenesis was recapitulated, with formation of acini-like structures, similar to those found in native mammary tissue. TGFβ1-driven EMT in 3D could be successfully promoted, generating M-like cells. TGFβ1 removal resulted in phenotypic switching to an intermediate state (RE cells), a hybrid cell population expressing both E and M markers at gene/protein levels. RE cells exhibited increased proliferative/clonogenic activity, as compared to M cells, being able to form large colonies containing cells with front-back polarity, suggesting a more aggressive phenotype. Our 3D model provides a powerful tool to investigate the role of the microenvironment on metastable EMT stages.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Actins / genetics
  • Actins / metabolism
  • Alginates / chemistry
  • Animals
  • Biomarkers / metabolism
  • Cadherins / genetics
  • Cadherins / metabolism
  • Cell Culture Techniques
  • Cell Line
  • Epithelial Cells / cytology
  • Epithelial Cells / drug effects*
  • Epithelial Cells / metabolism
  • Epithelial-Mesenchymal Transition / drug effects*
  • Epithelial-Mesenchymal Transition / genetics
  • Female
  • Gene Expression
  • Glucuronic Acid / chemistry
  • Hexuronic Acids / chemistry
  • Ki-67 Antigen / genetics
  • Ki-67 Antigen / metabolism
  • Mammary Glands, Animal / cytology
  • Mammary Glands, Animal / drug effects
  • Mammary Glands, Animal / metabolism
  • Mice
  • Occludin / genetics
  • Occludin / metabolism
  • Phenotype
  • Spheroids, Cellular / cytology*
  • Spheroids, Cellular / metabolism
  • Transforming Growth Factor beta1 / pharmacology*

Substances

  • Actins
  • Alginates
  • Biomarkers
  • Cadherins
  • Cdh1 protein, mouse
  • Hexuronic Acids
  • Ki-67 Antigen
  • Mki67 protein, mouse
  • Occludin
  • Ocln protein, mouse
  • Transforming Growth Factor beta1
  • Glucuronic Acid