Wang Huiqiang, Chen Bin, Duan Bo, Zheng Jiaxin, Wu Xinyi
Department of Urology, First Affliated Hospital of Xiamen University, Xiamen, Fujian 361003, P.R. China.
Department of Breast Surgery, First Affliated Hospital of Xiamen University, Xiamen, Fujian 361003, P.R. China.
Mol Med Rep. 2016 Oct;14(4):3343-9. doi: 10.3892/mmr.2016.5589. Epub 2016 Aug 4.
The present study aimed to determine the importance of microRNA‑205 (miR‑205) in the proliferation, apoptosis, invasion and metastasis of renal cell carcinoma (RCC) cells and the underlying molecular mechanisms. Reverse transcription‑polymerase chain reaction was used to quantify the expression levels of miR‑205 in RCC tissue, normal tissue adjacent to carcinoma, RCC cells and normal renal cells. It was determined that the expression levels of miR‑205 in RCC tissue and cells were reduced compared with those in normal tissue and renal cells. miR‑205 mimics and the negative control were prepared and transfected into RCC cells. Cell viability and apoptosis were investigated using methyl thiazolyl tetrazolium assay and Annexin V‑fluorescein isothiocyanate/propidium iodide staining, respectively. Cell migration and invasion were evaluated with Transwell assays. The protein expression levels of E2F transcription factor 1 (E2F1), B‑cell lymphoma‑2 (Bcl‑2), E‑cadherin, vimentin, phosphatase and tensin homolog (PTEN) and phosphorylated AKT serine/threonine kinase 1 (p‑AKT) were determined with western blot analysis. It was revealed that miR‑205 promoted the apoptosis of RCC cells and suppressed their proliferation, metastasis and invasion compared with the negative control. The expression levels of E2F1, Bcl‑2, vimentin and p‑AKT were downregulated compared with the negative control. The expression levels of E‑cadherin and PTEN were upregulated in the cells transfected with miR‑205 mimics compared with the negative control group. Therefore, it was concluded that miR‑205 suppressed cell proliferation, invasion, and metastasis in RCC cells via regulation of the PTEN/AKT signaling pathway. The present study may contribute to future miRNA‑based RCC therapy.
本研究旨在确定微小RNA-205(miR-205)在肾细胞癌(RCC)细胞增殖、凋亡、侵袭和转移中的重要性及其潜在分子机制。采用逆转录-聚合酶链反应定量RCC组织、癌旁正常组织、RCC细胞和正常肾细胞中miR-205的表达水平。结果显示,与正常组织和肾细胞相比,RCC组织和细胞中miR-205的表达水平降低。制备miR-205模拟物和阴性对照并转染至RCC细胞。分别采用甲基噻唑基四氮唑蓝法和膜联蛋白V-异硫氰酸荧光素/碘化丙啶染色法研究细胞活力和凋亡情况。通过Transwell实验评估细胞迁移和侵袭能力。采用蛋白质印迹分析测定E2F转录因子1(E2F1)、B细胞淋巴瘤-2(Bcl-2)、E-钙黏蛋白、波形蛋白、磷酸酶和张力蛋白同源物(PTEN)以及磷酸化AKT丝氨酸/苏氨酸激酶1(p-AKT)的蛋白表达水平。结果表明,与阴性对照相比,miR-205可促进RCC细胞凋亡,并抑制其增殖、转移和侵袭。与阴性对照相比,E2F1、Bcl-2、波形蛋白和p-AKT的表达水平下调。与阴性对照组相比,转染miR-205模拟物的细胞中E-钙黏蛋白和PTEN的表达水平上调。因此,得出结论:miR-205通过调控PTEN/AKT信号通路抑制RCC细胞的增殖、侵袭和转移。本研究可能为未来基于miRNA的RCC治疗做出贡献。