Clinical significance of the Epithelial-Mesenchymal Transition in the malignant transformation of canine mammary gland tumors.

上皮间质转化在犬乳腺肿瘤恶变中的临床意义。

• 批准号: 21780285
• 负责人: NAKAGAWA Takayuki
• 金额: $ 2.83万
• 依托单位: The University of Tokyo
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Young Scientists (B)
• 财政年份: 2009
• 资助国家: 日本
• 起止时间: 2009 至 2010
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-21780285/
• 关键词: 外科; 上皮間葉化; 犬; 乳腺腫瘍; TGF-β; Smad; 上皮間葉化現象; 獣医学; 癌; 細胞・組織

To clarify the significance of epithelial-mesenchymal transition(EMT) in the malignant transformation of canine mammary gland tumors(cMGTs), serial experiments were intended. EMT related factors were examined on tissue specimens collected by the surgical resection from clinical cases of spontaneous cMGTs. Immunohistochemical analysis revealed that the protein expression pattern indicating EMT was observed only in the adenocarcinoma cases, but not in the adenoma cases. For further in vitro studies, we obtained two clonal cell lines derived from the same cMGT patient, which show the different malignancy in the culture condition and the transplanted mouse model. Highly malignant clonal cell line, CHMp5b, was found to have the mesencymal characteristics with high expressions of vimentin and N-cadherin, whereas low malignant cell line, CHMp13a, showed the epithelial characteristics with high expressions of E-cadherin and β-catenin. In the trans-well assay, CHMp5b showed significantly higher motility and invasive potential than CHMp13a. These results suggested the relationship between the EMT status and the malignancy of cMGT cells. By the stimulus of TGF-β on these cell lines, vimentin expression was induced and this expression was paralleled with the phosphorylation of Smad2. In the experiment using xCELLigence system, changes of expressions of EMT related proteins were correlated with motility and invasive potential of these cells dynamically. From the results of these experiments, EMT might relate to the malignant transformation of cMGT dynamically and Smad2 cascades might be a crucial factor in converting the EMT status. We continue the further study on the significance of EMT in canine MGT malignancy with the controlled experiment by RNAi method, and confirm these findings obtained from these studies in the clinical cMGT cases.

为探讨上皮间质转化（EMT）在犬乳腺肿瘤（cMGTs）恶性转化中的意义，本研究进行了一系列实验。对临床自发性cMGT病例手术切除的组织标本进行EMT相关因素检测。免疫组化分析显示，EMT的蛋白表达模式仅在腺癌中观察到，而在腺瘤中未观察到。为了进一步的体外研究，我们获得了来自同一cMGT患者的两个克隆细胞系，它们在培养条件和移植小鼠模型中显示出不同的恶性度。高度恶性克隆细胞系CHMp 5 b具有间质细胞特征，高表达vimentin和N-cadherin，而低度恶性克隆细胞系CHMp 13 a具有上皮细胞特征，高表达E-cadherin和β-catenin。在trans-well测定中，CHMp 5 b显示出比CHMp 13 a显著更高的运动性和侵袭潜力。这些结果表明，EMT状态与cMGT细胞的恶性程度有关。在TGF-β刺激下，Vimentin表达增加，并与Smad 2磷酸化有关。在xCELLigence系统的实验中，EMT相关蛋白表达的变化与这些细胞的运动和侵袭能力动态相关。从这些实验的结果来看，EMT可能与cMGT的恶性转化动态相关，Smad 2级联可能是EMT状态转换的关键因素。本研究通过RNAi方法的对照实验，进一步探讨了EMT在犬MGT恶性肿瘤中的意义，并在临床病例中证实了这些研究结果。

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