Role of epimorphin in bile duct formation of rat liver epithelial stem-like cells: involvement of small G protein RhoA and C/EBPβ

J Cell Physiol. 2011 Nov;226(11):2807-16. doi: 10.1002/jcp.22625.

Abstract

Epimorphin/syntaxin 2 is a high conserved and very abundant protein involved in epithelial morphogenesis in various organs. We have shown recently that epimorphin (EPM), a protein exclusively expressed on the surface of hepatic stellate cells and myofibroblasts of the liver, induces bile duct formation of hepatic stem-like cells (WB-F344 cells) in a putative biophysical way. Therefore, the aim of this study was to present some of the molecular mechanisms by which EPM mediates bile duct formation. We established a biliary differentiation model by co-culture of EPM-overexpressed mesenchymal cells (PT67(EPM)) with WB-F344 cells. Here, we showed that EPM could promote WB-F344 cells differentiation into bile duct-like structures. Biliary differentiation markers were also elevated by EPM including Yp, Cx43, aquaporin-1, CK19, and gamma glutamyl transpeptidase (GGT). Moreover, the signaling pathway of EPM was analyzed by focal adhesion kinase (FAK), extracellular regulated kinase 1/2 (ERK1/2), and RhoA Western blot. Also, a dominant negative (DN) RhoA-WB-F344 cell line (WB(RhoA-DN)) was constructed. We found that the levels of phosphorylation (p) of FAK and ERK1/2 were up-regulated by EPM. Most importantly, we also showed that RhoA is necessary for EPM-induced activation of FAK and ERK1/2 and bile duct formation. In addition, a dual luciferase-reporter assay and CHIP assay was performed to reveal that EPM regulates GGT IV and GGT V expression differentially, possibly mediated by C/EBPβ. Taken together, these data demonstrated that EPM regulates bile duct formation of WB-F344 cells through effects on RhoA and C/EBPβ, implicating a dual aspect of this morphoregulator in bile duct epithelial morphogenesis.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Aquaporin 1 / metabolism
  • Bile Ducts / cytology
  • Bile Ducts / growth & development*
  • Bile Ducts / metabolism
  • CCAAT-Enhancer-Binding Protein-beta / metabolism*
  • Cell Differentiation / drug effects
  • Cell Line
  • Coculture Techniques
  • Connexin 43 / metabolism
  • Glutathione S-Transferase pi / metabolism
  • Liver / cytology
  • Liver / growth & development*
  • Liver / metabolism
  • Membrane Glycoproteins / metabolism*
  • Mesoderm / cytology
  • Mesoderm / metabolism
  • Protein Kinases / metabolism
  • Rats
  • Signal Transduction
  • Stem Cells / cytology
  • Stem Cells / metabolism*
  • Up-Regulation
  • gamma-Glutamyltransferase / metabolism
  • rhoA GTP-Binding Protein / metabolism*

Substances

  • Aqp1 protein, rat
  • CCAAT-Enhancer-Binding Protein-beta
  • Cebpb protein, rat
  • Connexin 43
  • Membrane Glycoproteins
  • Stx2 protein, rat
  • Aquaporin 1
  • gamma-Glutamyltransferase
  • Glutathione S-Transferase pi
  • Protein Kinases
  • rhoA GTP-Binding Protein