Department of Urology, Tongde Hospital of Zhejiang Province, Hangzhou, Zhejiang, People's Republic of China.
Eur Rev Med Pharmacol Sci. 2017 Oct;21(20):4557-4565.
To dissect the functioning mode of miR-645 on renal clear cell carcinoma cell metastasis and growth, and provide therapeutic targets for renal clear cell carcinoma.
Quantitative Real-time PCR (qRT-PCR) assay was employed to detect miR-645 expression level. Wound healing assay and transwell assay were performed to investigate metastasis capacity of renal clear cell carcinoma cells. Cell Counting Kit 8 (CCK8) assay was incorporated to assess cell proliferation capacity. Flow cytometry was used to identify cell apoptosis and cell cycle distribution. Protein levels were assessed by Western blotting assay. The target gene was predicted and verified by bioinformatics analysis and luciferase assay.
MiR-645 was upregulated in renal clear cell carcinoma tissues when compared with para-carcinoma tissues (n=32). Downregulated miR-645 could attenuate cell migration and invasion capacities, as well as inhibited cell proliferation capacity, promoted cell apoptosis and cell cycle arrest at G0/G1 phase. GK5 was chosen as the target gene of miR-645 by bioinformatics analysis and luciferase reporter assay. Moreover, silence of GK5 could rescue tumor suppression role of downregulated miR-645 on renal clear cell carcinoma metastasis.
Knockdown of miR-645 exerted tumor-suppressive effects on renal clear cell carcinoma metastasis and growth via targeting GK5 in vitro, which provided an innovative and candidate target for diagnose and treatment of renal clear cell carcinoma.
剖析 miR-645 对肾透明细胞癌细胞转移和生长的作用方式，为肾透明细胞癌提供治疗靶点。
采用实时定量聚合酶链反应（qRT-PCR）检测 miR-645 的表达水平。通过划痕愈合实验和 Transwell 实验研究肾透明细胞癌细胞的转移能力。通过细胞计数试剂盒 8（CCK8）检测细胞增殖能力。采用流式细胞术检测细胞凋亡和细胞周期分布。通过 Western blot 检测蛋白水平。通过生物信息学分析和荧光素酶报告实验预测和验证靶基因。
与癌旁组织相比（n=32），肾透明细胞癌组织中 miR-645 呈上调表达。下调 miR-645 可减弱细胞迁移和侵袭能力，抑制细胞增殖能力，促进细胞凋亡和细胞周期阻滞在 G0/G1 期。生物信息学分析和荧光素酶报告实验选择 GK5 作为 miR-645 的靶基因。此外，沉默 GK5 可挽救下调 miR-645 对肾透明细胞癌转移的抑制作用。
在体外，下调 miR-645 通过靶向 GK5 对肾透明细胞癌转移和生长发挥肿瘤抑制作用，为肾透明细胞癌的诊断和治疗提供了一种创新的候选靶点。
