title: Expression of E-cadherin repressors SNAIL, ZEB1 and ZEB2 by tumour and stromal cells influences tumour-budding phenotype and suggests heterogeneity of stromal cells in pancreatic cancer.
abstract: BACKGROUND
  There is evidence that tumour-stroma interactions have a major role in the neoplastic progression of pancreatic ductal adenocarcinoma (PDAC). Tumour budding is thought to reflect the process of epithelial-mesenchymal transition (EMT); however, the relationship between tumour buds and EMT remains unclear. Here we characterize the tumour-budding- and stromal cells in PDAC at protein and mRNA levels concerning factors involved in EMT.
  
  
  METHODS
  mRNA in situ hybridisation and immunostaining for E-cadherin, β-catenin, SNAIL1, ZEB1, ZEB2, N-cadherin and TWIST1 were assessed in the main tumour, tumour buds and tumour stroma on multipunch tissue microarrays from 120 well-characterised PDACs and associated with the clinicopathological features, including peritumoural (PTB) and intratumoural (ITB) budding.
  
  
  RESULTS
  Tumour-budding cells showed increased levels of ZEB1 (P<0.0001) and ZEB2 (P=0.0119) and reduced E-cadherin and β-catenin (P<0.0001, each) compared with the main tumour. Loss of membranous β-catenin in the main tumour (P=0.0009) and tumour buds (P=0.0053), without nuclear translocation, as well as increased SNAIL1 in tumour and stromal cells (P=0.0002, each) correlated with high PTB. ZEB1 overexpression in the main tumour-budding and stromal cells was associated with high ITB (P=0.0084; 0.0250 and 0.0029, respectively) and high PTB (P=0.0005; 0.0392 and 0.0007, respectively). ZEB2 overexpression in stromal cells correlated with higher pT stage (P=0.03), lymphatic invasion (P=0.0172) and lymph node metastasis (P=0.0152).
  
  
  CONCLUSIONS
  In the tumour microenvironment of phenotypically aggressive PDAC, tumour-budding cells express EMT hallmarks at protein and mRNA levels underlining their EMT-type character and are surrounded by stromal cells expressing high levels of the E-cadherin repressors ZEB1, ZEB2 and SNAIL1, this being strongly associated with the tumour-budding phenotype. Moreover, our findings suggest the existence of subtypes of stromal cells in PDAC with phenotypical and functional heterogeneity.
authors.affiliation: Translational Research Unit, Institute of Pathology, University of Bern, Murtenstrasse 31, Bern CH-3010, Switzerland.
authors.forename: J A
authors.initials: JA
authors.lastname: Galván
authors.affiliation: Translational Research Unit, Institute of Pathology, University of Bern, Murtenstrasse 31, Bern CH-3010, Switzerland.
authors.forename: I
authors.initials: I
authors.lastname: Zlobec
authors.affiliation: 1] Translational Research Unit, Institute of Pathology, University of Bern, Murtenstrasse 31, Bern CH-3010, Switzerland [2] Division of Clinical Pathology, Institute of Pathology, University of Bern, Murtenstrasse 31, Bern CH-3010, Switzerland.
authors.forename: M
authors.initials: M
authors.lastname: Wartenberg
authors.affiliation: 1] Translational Research Unit, Institute of Pathology, University of Bern, Murtenstrasse 31, Bern CH-3010, Switzerland [2] Division of Clinical Pathology, Institute of Pathology, University of Bern, Murtenstrasse 31, Bern CH-3010, Switzerland.
authors.forename: A
authors.initials: A
authors.lastname: Lugli
authors.affiliation: Department of Visceral Surgery, Insel University Hospital, Freiburgstrasse 4, CH-3010 Bern, Switzerland.
authors.forename: B
authors.initials: B
authors.lastname: Gloor
authors.affiliation: 1] Translational Research Unit, Institute of Pathology, University of Bern, Murtenstrasse 31, Bern CH-3010, Switzerland [2] Division of Clinical Pathology, Institute of Pathology, University of Bern, Murtenstrasse 31, Bern CH-3010, Switzerland.
authors.forename: A
authors.initials: A
authors.lastname: Perren
authors.affiliation: 1] Translational Research Unit, Institute of Pathology, University of Bern, Murtenstrasse 31, Bern CH-3010, Switzerland [2] Division of Clinical Pathology, Institute of Pathology, University of Bern, Murtenstrasse 31, Bern CH-3010, Switzerland.
authors.forename: E
authors.initials: E
authors.lastname: Karamitopoulou
mesh_terms: D015820:Cadherins / Q000378:metabolism; D021441:Carcinoma, Pancreatic Ductal / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D058750:Epithelial-Mesenchymal Transition; D018398:Homeodomain Proteins / Q000096:biosynthesis* / Q000235:genetics; D006801:Humans; D010190:Pancreatic Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D010641:Phenotype; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*; D012097:Repressor Proteins / Q000096:biosynthesis* / Q000235:genetics; D000071250:Snail Family Transcription Factors; D017154:Stromal Cells / Q000473:pathology*; D014157:Transcription Factors / Q000096:biosynthesis* / Q000235:genetics; D059016:Tumor Microenvironment; D000074165:Zinc Finger E-box Binding Homeobox 2; D000071799:Zinc Finger E-box-Binding Homeobox 1
chemical_list: D015820:Cadherins; D018398:Homeodomain Proteins; D012333:RNA, Messenger; D012097:Repressor Proteins; C000604407:SNAI1 protein, human; D000071250:Snail Family Transcription Factors; D014157:Transcription Factors; C486307:ZEB1 protein, human; C492363:ZEB2 protein, human; D000074165:Zinc Finger E-box Binding Homeobox 2; D000071799:Zinc Finger E-box-Binding Homeobox 1