Cervical cancer is a critically malignant tumor with the second mortality of females worldwide. MicroRNAs (miRNAs) are short but regulatory non-coding RNAs playing a pivotal role in many biological processes including tumorigenesis. However, the exact role of miR-140-3p in cervical cancer remains to be elucidated. Here we identified that miR-140-3p was significantly reduced in cervical cancer tissues by comprehensive analysis of TCGA data, hinting that higher expression level of miR-140-3p predicted a good clinical prognosis. Quantitative real-time PCR (RT-qPCR) assay was performed to confirm the negative correlation between miR-140-3p expression level and human cervical cancer tissues as well as various cervical cancer cell lines. To clarify the certain role of miR-140-3p, forced expression by microRNA mimics was applied in Caski and C33A cells, showing that miR-140-3p overexpression significantly impeded the proliferation of cervical cancer cells by cell count kit (CCK-8) assay. Western blot analysis of cell cycle-related proteins Cyclin A, Cyclin B1 and Cyclin D1 have further confirmed the cell cycle arrest was induced by the ectopic expression of miR-140-3p. Annexin-V based FACS analysis also found the simultaneous appearance of early apoptotic cell population in miR-140-3p overexpression cells. The protein level of BCL-2 was attenuated in accompany with elevated Bax and Cleaved caspase-3 protein, indicating miR-140-3p overexpression induced early apoptosis. Mechanistically, we demonstrated that miR-140-3p could target the 3'UTR of RRM2 which has been proved to be highly involved in the onset of cancer. Furthermore, upregulation of miR-140-3p and RRM2 failed to inhibit the proliferation of human cervical cancer cells, revealing that RRM2 served as the target downstream gene of miR-140-3p abolishing its ability as a tumor suppressor. Overall, we figured out the new role of miR-140-3p in cervical cancer and concluded that miR-140-3p was a candidate of cancer control in preclinical.

中文翻译：

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miR-140-3p通过靶向RRM2诱导细胞周期停滞和早期凋亡来阻止人宫颈癌细胞的增殖。

宫颈癌是一种严重的恶性肿瘤，全世界女性的死亡率均居第二位。微小RNA（miRNA）短，但调控性非编码RNA在包括肿瘤发生在内的许多生物学过程中起着关键作用。但是，miR-140-3p在宫颈癌中的确切作用仍有待阐明。在这里，我们通过对TCGA数据的全面分析，发现宫颈癌组织中的miR-140-3p显着降低，这表明miR-140-3p的较高表达水平预示了良好的临床预后。进行定量实时PCR（RT-qPCR）分析以确认miR-140-3p表达水平与人宫颈癌组织以及各种宫颈癌细胞系之间的负相关性。为了阐明miR-140-3p的特定作用，将microRNA模拟物强制表达应用于Caski和C33A细胞，通过细胞计数试剂盒（CCK-8）分析显示，miR-140-3p的过表达显着阻碍了宫颈癌细胞的增殖。细胞周期相关蛋白Cyclin A，Cyclin B1和Cyclin D1的蛋白质印迹分析进一步证实了miR-140-3p的异位表达诱导了细胞周期阻滞。基于膜联蛋白-V的FACS分析还发现miR-140-3p过表达细胞中早期凋亡细胞群同时出现。BCL-2的蛋白水平随Bax和Cleaved caspase-3蛋白的升高而减弱，表明miR-140-3p过表达诱导了早期凋亡。从机理上讲，我们证明了miR-140-3p可以靶向RRM2的3'UTR，这已被证明与癌症的发生密切相关。此外，miR-140-3p和RRM2的上调未能抑制人宫颈癌细胞的增殖，表明RRM2作为miR-140-3p的目标下游基因，从而取消了其作为肿瘤抑制因子的能力。总体而言，我们发现了miR-140-3p在子宫颈癌中的新作用，并得出结论：miR-140-3p是临床前癌症控制的候选者。