核糖核酸酶抑制因子与整合素连接激酶对 膀胱癌上皮- 间质转化与转移的影响

Crosstalk between RI and ILK regulating EMT and metastasis of bladder cancer

摘要:

目的 研究核糖核酸酶抑制因子（RI）与整合素连接激酶（ILK）相互作用对膀胱癌上皮- 间质转 化（EMT）与转移的影响。方法 构建过表达RI 或ILK 的膀胱癌EJ 细胞系，免疫荧光检测RI 与ILK 在EJ 细胞 中的表达及共定位，倒置相差显微镜下观察细胞形态，Western blot 检测细胞中RI、ILK 及EMT 标志物的表达， 复制膀胱癌裸鼠移植瘤模型，免疫组织化学分析瘤组织中RI、ILK 及EMT 标志物的表达，HE 染色观察裸鼠肺 转移。结果 EJ 细胞中RI 与ILK 存在共定位的现象，RI 与ILK 存在相互作用。Western blot 结果显示，EJ-RI 组E- 钙黏蛋白（E-cadherin）表达高于EJ 组（P <0.05），EJ-ILK 组基质金属蛋白酶2（MMP2）、基质金属蛋白 酶9（MMP9）、N- 钙黏蛋白（N-cadherin）、波形蛋白（Vimentin）、扭转蛋白（Twist）、核转录因子（Snail） 与重组人S100 钙结合蛋白A4（S100A4）表达高于EJ 组（P <0.05）。过表达RI 抑制膀胱癌体内外EMT 及移 植瘤自发性肺转移，而过表达ILK 促进EMT 发生及膀胱癌转移。结论 RI 与ILK 相互作用调节膀胱癌EMT 与 转移。

Abstract:

Objective To investigate the regulative effect of crosstalk between ribonuclease inhibitor (RI) and intergrin-linked kinase (ILK) on epithelial-mesenchymal transition (EMT) and metastasis of bladder cancer. Methods Bladder cancer cell lines with overexpression of RI or ILK were constructed. Western blot was performed to analyze the expressions of RI, ILK and EMT markers. Immunofluorescence assay was performed to identify colocalization of RI and ILK. Cell morphology was observed under phase contrast microscope. The model of bladder cancer xenografts in nude mice was established. HE staining of lung tissues was conducted to evaluate pulmonary metastasis of bladder cancer. Results The co-localization and interaction of RI and ILK were observed and verified in EJ cells. Western blot showed that the expression of E-cadherin was significantly increased in EJ-RI cells compared with that in EJ cells. Expression levels of MMP-2, MMP-9, N-cadherin, Vimentin, Twist, Snail and S100A4 in EJ-ILK cells were dramatically upregulated when compared with EJ cells. Overexpression of RI suppressed EMT and metastasis both in vitro and in vivo. Overexpression of ILK promoted EMT and metastasis both in vitro and in vivo. Conclusion Crosstalk between RI and ILK regulates EMT and metastasis of bladder cancer.