Epithelial-to-mesenchymal transition in pancreatic ductal adenocarcinoma: Characterization in a 3D-cell culture model

Nicoletta Gagliano, Giuseppe Celesti, Lorenza Tacchini, Stefano Pluchino, Chiarella Sforza, Marco Rasile, Vincenza Valerio, Luigi Laghi, Vincenzo Conte, Patrizia Procacci

Research output: Contribution to journalArticle

Abstract

AIM: To analyze the effect of three-dimensional (3D)-arrangement on the expression of epithelial-tomesenchymal transition markers in pancreatic adenocarcinoma (PDAC) cells. METHODS: HPAF-II, HPAC, and PL45 PDAC cells were cultured in either 2D-monolayers or 3D-spheroids. Ultrastructure was analyzed by transmission electron microscopy. The expression of E-cadherin, β-catenin, N-cadherin, collagen type I (COL-I), vimentin, a-smooth muscle actin (αSMA), and podoplanin was assayed by confocal microscopy in cells cultured on 12-mm diameter round coverslips and in 3D-spheroids. Gene expression for E-cadherin, Snail, Slug, Twist, Zeb1, and Zeb2 was quantified by real-time PCR. E-cadherin protein level and its electrophoretic pattern were studied by Western blot in cell lysates obtained from cells grown in 2D-monolayers and 3D-spheroids. RESULTS: The E-cadherin/β-catenin complex was expressed in a similar way in plasma membrane cell boundaries in both 2D-monolayers and 3D-spheroids. E-cadherin increased in lysates obtained from 3D-spheroids, while cleavage fragments were more evident in 2D-monolayers. N-cadherin expression was observed in very few PDAC cells grown in 2D-monolayers, but was more evident in 3D-spheroids. Some cells expressing COL-I were observed in 3D-spheroids. Podoplanin, expressed in collectively migrating cells, and αSMA were similarly expressed in both experimental conditions. The concomitant maintenance of the E-cadherin/β-catenin complex at cell boundaries supports the hypothesis of a collective migration for these cells, which is consistent with podoplanin expression. CONCLUSION: We show that a 3D-cell culture model could provide deeper insight into understanding the biology of PDAC and allow for the detection of marked differences in the phenotype of PDAC cells grown in 3D-spheroids.

Original languageEnglish
Pages (from-to)4466-4483
Number of pages18
JournalWorld Journal of Gastroenterology
Volume22
Issue number18
DOIs
Publication statusPublished - May 14 2016

Keywords

  • 3D-spheroids
  • E-cadherin
  • Epithelial-to-mesenchymal transition
  • Pancreatic ductal adenocarcinoma
  • Podoplanin

ASJC Scopus subject areas

  • Gastroenterology

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  • Cite this

    Gagliano, N., Celesti, G., Tacchini, L., Pluchino, S., Sforza, C., Rasile, M., Valerio, V., Laghi, L., Conte, V., & Procacci, P. (2016). Epithelial-to-mesenchymal transition in pancreatic ductal adenocarcinoma: Characterization in a 3D-cell culture model. World Journal of Gastroenterology, 22(18), 4466-4483. https://doi.org/10.3748/wjg.v22.i18.4466