Lei Jianjun, Fan Lin, Wei Guangbing, Chen Xin, Duan Wanxing, Xu Qinhong, Sheng Wei, Wang Kang, Li Xuqi
Department of Hepatobiliary Surgery, First Affiliated Hospital Medical College of Xi'an Jiaotong University, Xi'an, Shaanxi, 710061, China.
Tumour Biol. 2015 Apr;36(4):3119-26. doi: 10.1007/s13277-014-2948-z. Epub 2014 Dec 14.
Hypoxia can induce HIF-1α expression and promote the epithelial-mesenchymal transition (EMT) and invasion of cancer cells. However, their mechanisms remain unclear. The objective of this study was to evaluate the role of Gli-1, an effector of the Hedgehog pathway, in the hypoxia-induced EMT and invasion of breast cancer cells. Human breast cancer MDA-MB-231 cells were transfected with HIF-1α or Gli-1-specific small interfering RNA (siRNA) and cultured under a normoxic or hypoxic condition. The relative levels of HIF-1α, Gli-1, E-cadherin, and vimentin in the cells were characterized by quantitative RT-PCR and Western blot assays, and the invasion of MDA-MB-231 cells was determined. Data was analyzed by Student T test, one-way ANOVA, and post hoc LSD test or Mann-Whitney U when applicable. We observed that hypoxia significantly upregulated the relative levels of vimentin expression, but downregulated E-cadherin expression and promoted the invasion of MDA-MB-231 cells, associated with upregulated HIF-1α translation and Gil-1 expression. Knockdown of HIF-1α mitigated hypoxia-modulated Gil-1, vimentin and E-cadherin expression, and invasion of MDA-MB-231 cells. Knockdown of Gil-1 did not significantly change hypoxia-upregulated HIF-1α translation but completely eliminated hypoxia-modulated vimentin and E-cadherin expression and invasion of MDA-MB-231 cells. These data indicate that Gil-1 is crucial for hypoxia-induced EMT and invasion of breast cancer cells and may be a therapeutic target for intervention of breast cancer metastasis.
缺氧可诱导缺氧诱导因子-1α（HIF-1α）表达，促进癌细胞的上皮-间质转化（EMT）及侵袭。然而，其机制仍不清楚。本研究的目的是评估Hedgehog信号通路的效应分子Gli-1在缺氧诱导的乳腺癌细胞EMT及侵袭中的作用。将人乳腺癌MDA-MB-231细胞用HIF-1α或Gli-1特异性小干扰RNA（siRNA）转染，并在常氧或缺氧条件下培养。通过定量逆转录-聚合酶链反应（RT-PCR）和蛋白质免疫印迹法检测细胞中HIF-1α、Gli-1、E-钙黏蛋白和波形蛋白的相对水平，并测定MDA-MB-231细胞的侵袭能力。数据采用学生t检验、单因素方差分析及事后最小显著差法检验（LSD检验），或在适用时采用曼-惠特尼U检验进行分析。我们观察到，缺氧显著上调波形蛋白表达的相对水平，但下调E-钙黏蛋白表达，并促进MDA-MB-231细胞的侵袭，这与HIF-1α翻译及Gli-1表达上调有关。敲低HIF-1α可减轻缺氧调节的Gli-1、波形蛋白和E-钙黏蛋白表达以及MDA-MB-231细胞的侵袭。敲低Gli-1并未显著改变缺氧上调的HIF-1α翻译，但完全消除了缺氧调节的波形蛋白和E-钙黏蛋白表达以及MDA-MB-231细胞的侵袭。这些数据表明，Gli-1对缺氧诱导的乳腺癌细胞EMT及侵袭至关重要，可能是干预乳腺癌转移的治疗靶点。
