title: Recapitulating epithelial tumor microenvironment in vitro using three dimensional tri-culture of human epithelial, endothelial, and mesenchymal cells.
abstract: Three-dimensional (3-D) cultures of cancer cells can potentially bridge the gap between 2-D drug screening and in vivo xenografts. The objective of this study was to characterize the cellular and extracellular matrix characteristics of spheroids composed of human lung epithelial cells (epi), pulmonary vascular endothelial (endo) cells, and human marrow-derived mesenchymal stems cells (MSCs).
  
  
  
  Spheroids composed of epi/endo/MSCs, termed herein as synthetic tumor microenvironment mimics (STEMs), were prepared by the hanging drop method. Cellular composition and distribution in the STEMs was characterized using fluorescence microscopy. Induction of reactive oxygen species and upregulation of efflux transporters was quantified using fluorometry and PCR, respectively, and phenotypic markers were qualitatively assessed using immunohistochemistry.
  
  
  
  STEMs exhibited three unique characteristics not captured in other spheroid cultures namely, the presence of a spheroid core devoid of epithelial cells and primarily composed of MSCs, a small viable population of endothelial cells hypothesized to be closely associated with MSCs within the hypoxic core, and discrete regions with high expression for vimentin and cytokeratin-18, whose co-expression is co-related with enhanced metastasis. Although cells within STEMs show elevated levels of reactive oxygen species and mRNA for ABC-B1, an efflux transporter associated with drug resistance, they exhibited only modest resistance to paclitaxel and gemcitabine in comparison to 2-D tri-cultures.
  
  
  
  The epi/endo/MSC spheroid model described herein offers a promising platform for understanding tumor biology and drug testing in vitro.
authors.affiliation: Institute for Macromolecular Chemistry, University of Freiburg, Hermann-Staudinger-Haus Stefan-Meier-Straße 31, 79104, Freiburg, Germany.
authors.forename: Surya P
authors.initials: SP
authors.lastname: Lamichhane
authors.affiliation: Institute for Macromolecular Chemistry, University of Freiburg, Hermann-Staudinger-Haus Stefan-Meier-Straße 31, 79104, Freiburg, Germany.|Helmholtz Virtual Institute on Multifunctional Biomaterials for Medicine, Kantstr. 55, 14513, Teltow, Germany.
authors.forename: Neha
authors.initials: N
authors.lastname: Arya
authors.affiliation: Institute for Macromolecular Chemistry, University of Freiburg, Hermann-Staudinger-Haus Stefan-Meier-Straße 31, 79104, Freiburg, Germany.
authors.forename: Esther
authors.initials: E
authors.lastname: Kohler
authors.affiliation: Institute for Macromolecular Chemistry, University of Freiburg, Hermann-Staudinger-Haus Stefan-Meier-Straße 31, 79104, Freiburg, Germany.
authors.forename: Shengnan
authors.initials: S
authors.lastname: Xiang
authors.affiliation: Institute for Macromolecular Chemistry, University of Freiburg, Hermann-Staudinger-Haus Stefan-Meier-Straße 31, 79104, Freiburg, Germany.|BIOSS-Centre for Biological Signalling Studies, University of Freiburg, Schänzlestraße 18, 79104, Freiburg, Germany.
authors.forename: Jon
authors.initials: J
authors.lastname: Christensen
authors.affiliation: Institute for Macromolecular Chemistry, University of Freiburg, Hermann-Staudinger-Haus Stefan-Meier-Straße 31, 79104, Freiburg, Germany. prasad.shastri@gmail.com.|BIOSS-Centre for Biological Signalling Studies, University of Freiburg, Schänzlestraße 18, 79104, Freiburg, Germany. prasad.shastri@gmail.com.|Helmholtz Virtual Institute on Multifunctional Biomaterials for Medicine, Kantstr. 55, 14513, Teltow, Germany. prasad.shastri@gmail.com.
authors.forename: V Prasad
authors.initials: VP
authors.lastname: Shastri
mesh_terms: D045744:Cell Line, Tumor; D018920:Coculture Techniques; D003841:Deoxycytidine / Q000031:analogs & derivatives* / Q000494:pharmacology; D019008:Drug Resistance, Neoplasm; D042783:Endothelial Cells / Q000166:cytology* / Q000378:metabolism; D004847:Epithelial Cells / Q000166:cytology* / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D053538:Keratin-18 / Q000235:genetics / Q000378:metabolism; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000378:metabolism; D017239:Paclitaxel / Q000494:pharmacology*; D017382:Reactive Oxygen Species / Q000378:metabolism; D018874:Spheroids, Cellular / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D059016:Tumor Microenvironment; D014746:Vimentin / Q000235:genetics / Q000378:metabolism; D000093542:Gemcitabine
chemical_list: D053538:Keratin-18; D017382:Reactive Oxygen Species; D014746:Vimentin; D003841:Deoxycytidine; D017239:Paclitaxel; D000093542:Gemcitabine
keywords: Drug resistance; Drug screening; Mesenchymal stem cells; Multicellular spheroids; Oxidative stress