Exogenous expression of E-cadherin in gallbladder carcinoma cell line G-415 restores its cellular polarity and differentiation.
E-cadherin (E-cad) is critical for the development and maintenance of polarity and differentiation of epithelial cells. Our previous study revealed gallbladder (GB) cancer cell lines expressing E-cadherin could form cystic or spheroid-like structures in a 3-D matrix. This differentiated phenotype was disrupted by addition of anti-E-cad antibody. By extrapolation, we postulated that E-cad might directly regulate epithelial morphogenesis in GB carcinoma. In the present study, we introduced an expression plasmid harboring mouse E-cad cDNA (pBATEM2) into an undifferentiated GB cancer cell line (G-415) which did not express E-cad. We systemically compared differences including cell phenotype, morphogenesis, and ultrastructure between the parental G-415 cells (parental-cells) and E-cad transfected cells (E-cad-cells) under various conditions such as monolayer culture, 3-D gel culture as well as inoculation of nude mice. In contrast to parental-cells, E-cad-cells presented a typical epithelial shape with tight intercellular connections in monolayer culture. They developed round multicellular cystic structures in 3-D gel. The cysts secreted mucous substances via the apical surface into the cystic lumen, in which the substances were positively stained by alcian blue (pH 2.5). Ultrastructural sections of gel/xenograft tumor revealed re-establishment of adherens junctions and restoration of cell polarization. Immunohistochemical and immunofluorescent staining showed exogenous E-cad primarily accumulated at the extended cell-cell contact sites. Moreover, the membranous translocation of alpha-, and beta-catenin from cytoplasm was also found in 3-D matrix and xenograft tumor sections. Histopathologic examination revealed papillary adenocarcinoma features in the central region of the xenograft tumors. Our results demonstrate that exogenous expression of E-cad in undifferentiated GB cancer G-415 cells reverses differentiated characteristics in spatial growth conditions. Simultaneous translocation of alpha-, beta-catenin to membrane following functional E-cad expression may be a critical event for this process.
Jiao
W
W
Department of Surgery, Saga Medical School, Saga 849-8501, Japan.
Miyazaki
K
K
Kitajima
Y
Y
D000231:Adenocarcinoma, Papillary / Q000378:metabolism* / Q000473:pathology; D000818:Animals; D015153:Blotting, Western; D015820:Cadherins / Q000235:genetics / Q000378:metabolism*; D002454:Cell Differentiation / Q000502:physiology*; D016764:Cell Polarity / Q000502:physiology*; D017931:DNA Primers / Q000737:chemistry; D019084:Fluorescent Antibody Technique, Indirect; D005706:Gallbladder Neoplasms / Q000378:metabolism* / Q000473:pathology; D049452:Green Fluorescent Proteins; D006801:Humans; D007124:Immunoenzyme Techniques; D008164:Luminescent Proteins / Q000378:metabolism; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008854:Microscopy, Electron; D010957:Plasmids; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection; D014407:Tumor Cells, Cultured / Q000378:metabolism / Q000648:ultrastructure
D015820:Cadherins; D017931:DNA Primers; D008164:Luminescent Proteins; D049452:Green Fluorescent Proteins