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

影响正常胆囊上皮细胞形态发生的因素和调节胆囊癌细胞分化的因素。

• 批准号: 09671322
• 负责人: MIYAZAKI Kohji
• 金额: $ 2.11万
• 依托单位: Saga Medical School
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1997
• 资助国家: 日本
• 起止时间: 1997 至 1999
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-09671322/
• 关键词: Epithelial Cells; Gallbladder cancer; Morphogenesis; E-cadherin; α-catenin; Collagen gel culture; Proliferation; Differentiation; 胆嚢癌; 形態調節; 分化度; extracellular matrix; 胆嚢癌株細胞; 増殖因子; 形態形成; 接着因子

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

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、epimorphin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。相反，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诱导形态发生的急剧变化，形成分支网络，仅在分支接触的部位显示细胞-细胞接触。RGEC形成的多细胞囊肿不表达波形蛋白，但表达了大量的cytoskeratin和恢复连接复合物。相反，TGF-β1处理的细胞强烈表达波形蛋白沿着与分支结构，并显示细胞角蛋白表达和连接复合物减少。因此，TGF-β1诱导间充质样细胞的形状伴随着细胞骨架摩尔 ...更多信息 角膜改变，上皮极化和连接复合体丧失。为探讨细胞粘附分子在胆囊癌形态发生中的作用，本研究对四种人胆囊癌细胞株（GB-d1、KMG-C、GBK-1和G-415）进行了体外实验。它们在我们的标准凝胶培养物（SC）中显示出明显不同的形态学。GB-dl和KMG-C分别形成囊状和球状结构，其似乎分别代表良好分化和中度分化的癌症。GBK-1和G-415分别显示分支和“假腺”结构，这两者似乎都表明原始的去分化癌。在漂浮凝胶培养（FC），只有GB-d1表现出高度增加的趋势，对囊肿形成。GB-d1和KMG-C细胞表达E-cadherin和α-catenin，而GBK-1和G-415细胞不表达。此外，E-钙粘蛋白表达在GB-d1是1.82倍以上，在FC比SC，而E-钙粘蛋白表达水平KMG-C没有改变。GB-d1和KMG-C的α-catenin在SC和FC中的表达无明显差异。相反，这些蛋白质的异质性定位在KMG-C的球体结构中，在SC和FC。电镜检查显示，连接复合物的重建只发生在GB-d1细胞培养在FC中。GB-d1中的囊性结构的形成完全抑制的抗体对人E-cadherin。E-钙粘蛋白的表达及其膜定位是胆囊癌细胞高分化型形态发生所必需的。少

项目成果

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T.Hamamoto 等人：“在原发性胃癌附近的不完整型肠化生中改变了微卫星。”

向井伸介 他: "サイトカインによる胆嚢癌増殖形態の検討" 消化器癌の発生と進展. 9. 375-380 (1997)

Shinsuke Mukai等人：“通过细胞因子检查胆囊癌的生长模式”胃肠癌的发生和进展（1997年）。

M.Katano et.al.: "Prognostic value of platelet-derived factor(PDGF-A)in gastric carcinoma." Ann.Surg.227. 365-371 (1998)

M.Katano 等人：“血小板衍生因子 (PDGF-A) 在胃癌中的预后价值。”

森倫人: "胆嚢癌におけるE型カドヘリン,αカテニンの発現性と予後."日消外会誌. 33. 584-589 (2000)

Michihito Mori：“E 型钙粘蛋白和 α-连环蛋白在胆囊癌中的表达和预后。”Nichishu Gaikai 杂志 33. 584-589 (2000)