title: Reduced adhesive ligand density in engineered extracellular matrices induces an epithelial-mesenchymal-like transition.
abstract: A synergistic effect of biochemical and mechanical cues emanating from the extracellular matrix (ECM) on inducing malignant transformation of epithelial cells has been observed recently. However, the effect of quantitative changes in biochemical stimuli on cell phenotype, without changes in ECM component and rigidity, remains unknown. To determine this effect, we grew Madin-Darby canine kidney epithelial cells (MDCK) on gold surfaces immobilized with varying densities of cyclic arginine-glycine-aspartate (cRGD) peptide and analyzed cell morphology, cell migration, cytoskeletal organization, and protein expression. Cells grown on a surface presenting a higher density of cRGD displayed an epithelial morphology and grew in clusters, while those grown on a diluted cRGD surface transformed into an elongated, fibroblast-like form with extensive scattering. Time-lapse imaging of cell clusters grown on the concentrated cRGD surface revealed collective migration with intact cell-cell contacts accompanied by the development of cortical actin. In contrast, cells migrated individually and formed stress fibers on the substrate with sparse cRGD. These data point towards transdifferentiation of epithelial cells to mesenchymal-like cells when plated on a diluted cRGD surface. Supporting this hypothesis, immunofluorescence microscopy and western blot analysis revealed increased membrane localization and total expression of N-cadherin and vimentin in cells undergoing mesenchymal-like transition. Taken together, these results suggest a possible role of decreased biochemical stimuli from the ECM in regulating epithelial phenotype switching.
  
  
  
  Epithelial-mesenchymal transition (EMT) is a process where adherent epithelial cells convert into individual migratory mesenchymal phenotype. It plays an important role both in physiological and pathological processes. Recent studies demonstrate that the program is not only governed by soluble factors and gene expressions, but also modulated by biochemical and mechanical cues in ECMs. In this study, we developed chemically defined surfaces presenting controlled ECM ligand densities and studied their impact on the EMT progression. Morphological and biochemical analyses of epithelial cells cultured on the surfaces indicate the cells undergo an EMT-like transition on the diluted cRGD surface while retaining epithelial characteristics on the cRGD-rich substrate, suggesting an important role of the ECM ligand density in epithelial phenotype switching.
authors.affiliation: World Premier International (WPI) Research Center Initiative, International Center for Materials Nanoarchitectonics (MANA), National Institute for Materials Science (NIMS), 1-1 Namiki, Tsukuba 305-0044, Japan.
authors.forename: Saw
authors.initials: S
authors.lastname: Marlar
authors.affiliation: World Premier International (WPI) Research Center Initiative, International Center for Materials Nanoarchitectonics (MANA), National Institute for Materials Science (NIMS), 1-1 Namiki, Tsukuba 305-0044, Japan.
authors.forename: Shimaa A
authors.initials: SA
authors.lastname: Abdellatef
authors.affiliation: World Premier International (WPI) Research Center Initiative, International Center for Materials Nanoarchitectonics (MANA), National Institute for Materials Science (NIMS), 1-1 Namiki, Tsukuba 305-0044, Japan. Electronic address: NAKANISHI.Jun@nims.go.jp.
authors.forename: Jun
authors.initials: J
authors.lastname: Nakanishi
mesh_terms: D000818:Animals; D002465:Cell Movement; D003599:Cytoskeleton / Q000378:metabolism*; D004285:Dogs; D058750:Epithelial-Mesenchymal Transition; D005109:Extracellular Matrix / Q000737:chemistry*; D006046:Gold; D008024:Ligands; D061985:Madin Darby Canine Kidney Cells; D010456:Peptides, Cyclic / Q000737:chemistry*; D014176:Protein Biosynthesis; D013499:Surface Properties; D023822:Tissue Engineering
chemical_list: D008024:Ligands; D010456:Peptides, Cyclic; C507169:cyclic arginine-glycine-aspartic acid peptide; D006046:Gold
keywords: Epithelial-mesenchymal transition; Extracellular matrix; MDCK; Mechanobiology; Microenvironment; cRGD