Factors affecting morphogenesis of normal gallbladder epithelial cells and those regulationg differentiation in gallbladder cancer cells.

兔胆囊上皮细胞（RGEC）悬浮在胶原凝胶中形成具有形态极性的球形囊肿。EGF、HGF、表硫啡和FCM通过加速透明极化囊泡的增殖和聚集来促进囊肿成熟。相比之下，TGF-β1在单层和胶原凝胶培养中均显著抑制DNA合成，并促进胶原凝胶中分支结构的形成。此外，在EGF存在下，TGF-β1诱导了形态发生的剧烈变化，形成分支网络，仅在分支接触的部位显示细胞-细胞接触。形成rgec的多细胞囊肿不表达波形蛋白，但表达大量的细胞角质蛋白和恢复的连接复合物。而TGF-β1处理的细胞强表达vimentin并伴有分支结构，细胞角蛋白表达和连接复合物降低。因此，TGF-β1诱导间质样细胞形态，并伴有细胞骨架痣…更多的眼球变化，上皮极化和连接复合物的丧失。这些结果表明，RGEC的形态发生程序可能是由这些肽的相互作用和它们存在的时间决定的。为了阐明细胞粘附分子在胆囊癌形态学中的作用，我们在体外对四种人胆囊癌细胞系（GB-d1、kg - c、GBK-1和G-415）进行了检测。它们在我们的标准凝胶培养（SC）中表现出明显不同的形态。GB-dl和KMG-C分别形成囊状和球状结构，似乎分别代表良好和中度分化的癌症。GBK-1和G-415分别显示分支和“伪腺”结构，两者似乎都表明原始的去分化癌症。在浮凝胶培养（FC）中，只有GB-d1显示出高度增加的囊肿形成倾向。GBK-1和G-415细胞中无E-cadherin和α-catenin表达。此外，FC中E-cadherin在GB-d1中的表达量是SC的1.82倍，而kg -c中E-cadherin的表达量没有变化。SC和FC在GB-d1和KMG-C中α-catenin的表达均无差异。GB-d1的免疫染色显示这些蛋白定位在细胞膜上。相比之下，在SC和FC的KMG-C的球形结构中检测到这些蛋白的异质定位。电镜检查显示，仅在FC培养的GB-d1细胞中出现了连接复合物的重建。一种抗人E-cadherin抗体完全抑制了GB-d1囊性结构的形成。在胆囊癌细胞中，e -钙粘蛋白的表达及其膜定位是良好分化型形态发生所必需的。少

Rabbit gallbladder epithelial cells (RGEC) suspended in collagen gels form spherical cysts with morphologic polarity. EGF, HGF, epimorphin, and FCM promoted cyst maturation by accelerating the proliferation and aggregation of clear, polarized vesicles. In contrast, TGF-β1 markedly inhibited DNA synthesisin both monolayer and collagen gel cultures an promoted formation of branching structures in collagen gels. Furthermore, in the presence of EGF, TGF-β1 induced a drastic change in morphogenesis, with the formation of branching networks that showed cell-cell contact only at sites where branches touched. RGEC-forming multicellular cysts did not express vimentin but expressed significant amounts of cytoskeratin and regained junctional complexes. In contrast, TGF-β1 treated cells strongly expressed vimentin along with branching structures and showed decreases in cytokeratin expression and junctional complexes. Thus TGF-β1 induces a mesenchyme-like cell shape accompanied by cytoskeletal mole … More cular changes, with loss of both epithelial polarization and junctional complexes. These results suggest that the morphogenetic program of RGEC is likely tobe determined by the interaction of these peptides and the timing of their presence.To clarify the role of cell adhesion molecules in the morphology of gallbladder cancers, four human gallbladder cancer cell lines (GB-d1, KMG-C, GBK-1 and G-415) were eximined in vitro. They showed noticeably different morphologics in our standard gel cultures (SC). GB-dl and KMG-C formed cystic and spheroid structures, respectively, which seemed to represent well- and moderately-differentiated cancers, respectively. GBK-1 and G-415 showed branching and "pseudo-glandular" structures, respectively, both of which seemed to indicate original dedifferentiated cancers. In floating gel culture (FC), only GB-d1 showed a highly increased tendency toward cyst formation. Expression of E-cadherin and α-catenin was detected in GB-d1 and KMG-C, but not in GBK-1 and G-415 cells. Furthermore, E-cadherin expression in GB-d1 was 1.82 times greater in FC than in SC, while E-cadherin expression levels of KMG-C did not change. Neither GB-d1 nor KMG-C showed any difference in α-catenin expression between SC and FC.Immunostaining of GB-d1 revealed that these proteins were localized to the cell membrane. In contrast, heterogeneous localization of these proteins was detected in the spheroid structures of KMG-C, in both SC and FC.Electromicroscopic examination revealed that reestablishment of junctional complex was occurred only in GB-d1 cells cultured in FC.The formation of cystic structures in GB-d1 was completely inhibited by an antibody against human E-cadherin. Both expression of E-cadherin and its membranous localization are required for well-differentiated-type morphogenesis in gallbladder cancer cells. Less