miR-9-5p靶向Ambra1促进舌癌细胞增殖

miRNAs在肿瘤中异常表达，且与肿瘤的发生发展密切相关。目前发现，miR-9-5p在肿瘤中可能发挥原癌或抑癌效应，功能尚未完全阐述清楚。本文拟探讨miR-9-5p在舌癌中的作用。前期研究中收集10例舌癌组织及配对的癌旁组织，实时荧光定量PCR技术检测后发现，miR-9-5p在舌癌组织中的表达量显著高于癌旁组织，且其在舌癌细胞中的表达量也明显高于正常舌上皮细胞。此外，在舌癌细胞Tca8113中过表达miR-9-5p显著增加细胞的增殖能力。生物信息学预测及双荧光素酶报告基因实验证实，miR-9-5p可直接结合在自噬/苄氯素1调节因子1（activating molecule in beclin1-regulated autophagy, Ambra1）的 3′-UTR区域，靶向抑制Ambra1表达。Western印迹结果证实过表达miR-9-5p降低Ambra1的表达，反之亦然。Ambra1在舌癌细胞中的表达量显著低于正常舌上皮细胞。BrdU实验证实在舌癌细胞SCC-25中过表达Ambra1可显著抑制其增殖能力；相反，使用siRNA技术沉默Ambra1能够显著促进Tca8113细胞的增殖。在干预miR-9-5p的细胞中同时干预Ambra1的表达，结果发现Ambra1可显著逆转miR-9-5p对舌癌细胞增殖的促进作用。总之，miR-9-5p在舌癌中可能发挥原癌基因样作用，通过直接靶向抑制Ambra1表达进而促进舌癌细胞发生增殖。

miR-9-5p Promotes the Proliferation of Tongue Cancer Cells by Targeted Suppression of Ambra1

miRNAs are aberrantly expressed in tumors, and are closely related with tumorigenesis and progression. The roles of miR-9-5p in tumors are not very clear, as evidence shows that miR-9-5p functions as either an oncogene or a suppressor in tumors. We aimed to explore its function in the tongue cancer. First, qRT-PCR assays were used to detect miR-9-5p expression levels in ten cases of tongue cancer and normal tissues. The result showed that miR-9-5p was highly expressed in tongue cancer tissues, compared with the normal tissues. Similarly, miR-9-5p was much higher in tongue cancer cells than wild type tongue epithelial cells. Furthermore, the proliferation of Tca8113 cells was remarkably enhanced by overexpression of miR-9-5p. Bioinformatics analysis combined with dual luciferase assays demonstrated that miR-9-5p directly bound to the 3′-UTR of activating molecule in beclin1-regulated autophagy (Ambra1) and inhibited its expression. Western blotting assays suggested that overexpression of miR-9-5p down-regulated Ambra1, while knocking down of miR-9-5p did the opposite. Ambra1 was lowly expressed in the tongue cancer cell line SCC-25, compared with wild-type tongue epithelial cells. BrdU assays indicated that overexpressing Ambra1 markedly inhibited proliferation of SCC-25 cells. On the contrary, knocking down Ambra1 by siRNAs promoted proliferation of SCC-25 cells. Finally, we simultaneously modulated miR-9-5p and Ambra1 expression in tongue cancer cells, and found that Ambra1 overexpression could significantly reverse the proliferation of tongue cancer cells with miR-9-5p. Taken together, these data suggested that miR-9-5p might play an oncogenic role in tongue cancers, and silencing miR-9-5p promoted tongue cancer cell proliferation via directly targeting Ambra1.