Purpose

Multiple circular RNAs (circRNAs) have been reported to be dysregulated in hepatocellular carcinoma (HCC). However, their functions and modes of action are still largely unclear. Identifying key circRNAs and revealing their potential functions and molecular mechanisms is considered important for improving the diagnosis and treatment of HCC.

Methods

Dysregulated circRNAs in HCC were identified through integration of three human HCC circRNAs microarray datasets (GSE94508, GSE97332 and GSE 78520), followed by qRT-PCR validation in primary HCC tissues and cell lines. circRNA characteristics were verified through Sanger sequencing, RNase R treatment, northern blotting and intracellular localization analyses. In addition, circRNA functions in HCC development were assessed using CCK8, colony formation, EDU incorporation, flow cytometry, transwell and scratch wound healing assays in vitro and tumor xenograft assays in vivo. Next, underlying molecular mechanisms in HCC were assessed using dual-luciferase reporter, RNA pull-down, RNA immunoprecipitation and western blotting assays.

Results

We found that a novel circular RNA, circ-102,166, was down-regulated in HCC and that its expression level was significantly associated with multiple clinicopathologic characteristics, as well as the clinical prognosis of HCC patients. In vitro and in vivo experiments revealed that circ-102,166 overexpression significantly inhibited the proliferation, invasion, migration and tumorigenicity of HCC cells. Furthermore, we found that circ-102,166 can bind to miR-182 and miR-184 to regulate the expression of several of their downstream targets (FOXO3a, MTSS1, SOX7, p-RB and c-MYC).

Conclusion

Our data revealed a tumor-suppressing role of circ-102,166 in HCC. Down-regulation of circ-102,166 enhanced the proliferation and invasion of HCC cells by releasing the oncomiRs miR-182 and miR-184.

中文翻译：

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环状 RNA circ-102,166 作为 miR-182 和 miR-184 的海绵抑制肝细胞癌的增殖和侵袭

目的

据报道，多个环状 RNA（circRNA）在肝细胞癌（HCC）中失调。然而，它们的功能和作用方式在很大程度上仍不清楚。识别关键的 circRNA 并揭示其潜在功能和分子机制被认为对改善 HCC 的诊断和治疗具有重要意义。

方法

通过整合三个人类 HCC circRNA 微阵列数据集（GSE94508、GSE97332 和 GSE 78520），然后在原代 HCC 组织和细胞系中进行 qRT-PCR 验证，确定了 HCC 中失调的 circRNA。通过 Sanger 测序、RNase R 处理、northern 印迹和细胞内定位分析验证了 circRNA 特征。此外，使用 CCK8、集落形成、EDU 掺入、流式细胞术、Transwell 和划痕伤口愈合试验以及体内肿瘤异种移植试验评估了 circRNA 在 HCC 发展中的功能。接下来，使用双荧光素酶报告基因、RNA 下拉、RNA 免疫沉淀和蛋白质印迹分析评估了 HCC 中的潜在分子机制。

结果

我们发现一种新型环状 RNA circ-102,166 在 HCC 中下调，其表达水平与多种临床病理特征以及 HCC 患者的临床预后显着相关。体内外实验表明circ-102,166过表达显着抑制HCC细胞的增殖、侵袭、迁移和致瘤性。此外，我们发现 circ-102,166 可以与 miR-182 和 miR-184 结合以调节其几个下游靶标（FOXO3a、MTSS1、SOX7、p-RB 和 c-MYC）的表达。

结论

我们的数据揭示了 circ-102,166 在 HCC 中的肿瘤抑制作用。circ-102,166 的下调通过释放 oncomiRs miR-182 和 miR-184 增强了 HCC 细胞的增殖和侵袭。