微小核糖核酸-145藉由抑制ARF6調控上泌尿道上皮癌的侵襲能力
徐偉齊1,7、李怡琛2、李威明3,7,9、張玲麗1, 4、黃阿梅1,5,6、林慧惠7、
吳文正1,3,7、李經家3,7、連培因8、柯宏龍1,3,7
高雄醫學大學 醫學院醫學研究所1；高雄醫學大學 醫學院醫學研究所 解剖學科2，泌尿學科3，微生物與免疫學科4，生物化學科5；高雄醫學大學 臨床醫學研究所6；高雄醫學大學附設醫院泌尿科7，病理科8；衛生福利部屏東醫院泌尿科9
MicroRNA-145 regulates invasion in upper tract urothelial carcinoma by targeting ARF6
Wei-Chi Hsu 1,7, Yi-Chen Lee 2, Wei-Ming Li 3,7,9, Lin-Li Chang1, 4, A-Mei Huang 1,5,6, Hui-Hui Lin 7, Wen-Jeng Wu 1,3,7, Ching-Chia Li 3,7, Peir-In Liang 8, Hung-Lung Ke 1,3,7
Graduate Institute of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan1 ; Department of Anatomy2, Urology3, Microbiology4, and Biochemistry5 School of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan ; Graduate Institute of Clinical Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan6 ; Department of Urology7 and Pathology8, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan; Department of Urology, Ministry of Health and Welfare Pingtung Hospital, Pingtung, Taiwan9.
Purpose: Upper tract urothelial carcinoma (UTUC) is a rare tumor with extraordinarily different feature between Eastern and Western countries. Approximately 60% of UTUCs are invasive at diagnosis compared with 15-25% of bladder cancer. However, the molecular mechanisms of invasion in UTUC are not well understood. This study intended to explain that the role of ADP-ribosylation factor 6 (ARF6) in UTUC invasion. We also aimed to demonstrate the clinical significance of the microRNA (miR)-145 /ARF6 pathway, especially the predictive invasive function of ARF6 in UTUC.
Materials and methods: ARF6 expression was evaluated by immunohistochemistry and miR-145 expression was analyzed by real-time qPCR in patients with UTUC, and the correlation between their expression and clinicopathological variables were also examined. We demonstrated that the regulatory relationship between ARF6 and miR-145 by western blot and dual luciferase reporter assay. The role of the miR-145/ARF6 pathway in motility and invasion was analyzed in vitro.
Results: Our data showed that ARF6 expression was higher and miR-145 expression was lower in UTUC than normal urothelium. Up-regulation of ARF6 in UTUC was negatively correlated with miR-145 expression. In vitro study, miR-145 inhibited ARF6 expression by directly targeting its 3'-UTR. The results indicated that cell motility and invasion were all suppressed in UTUC cells transfected with miR-145 mimics or ARF6 siRNA. Moreover, ARF6 overexpression restored the inhibitory motility and invasion effects of miR-145. Notably, over-expression of miR-145 reduced MMP2, MMP9, MMP7 and N-cadherin protein levels and E-cadherin was increased in UTUC cells, in addition to these levels were reversed by ARF6 overexpression. We also found the expression of MMP2, MMP9 and N-cadherin were decreased, and cell invasion were repressed by MMP7 knockdown in UTUC cell lines.
Conclusion: These findings suggest that miR-145 may inhibit cell motility and invasion in UTUC cell lines by direct targeting ARF6/MMP7 through regulating epithelial-to-mesenchymal transition.