Department of Gastrointestinal Surgery, Taizhou Hospital of Zhejiang Province, Wenzhou Medical University, Linhai City, Zhejiang Province, China.
Anticancer Drugs. 2021 Oct 1;32(9):919-929. doi: 10.1097/CAD.0000000000001084.
Circular RNAs (circRNA) are a key regulator of cancer progression, including colorectal cancer (CRC). Nevertheless, the role of circRASSF2 in CRC remains unclear. Quantitative real-time PCR was used to measure the expression of circRASSF2 and miR-195-5p. Cell counting kit 8 assay, colony formation assay, flow cytometry and transwell assay were used to determine the proliferation, apoptosis, migration and invasion of cells, respectively. The levels of proliferation, metastasis and Wnt/β-catenin signaling pathway-related proteins, as well as Frizzled 4 (FZD4) protein, were determined using western blot analysis. Furthermore, a dual-luciferase reporter assay, RNA immunoprecipitation assay and RNA pull-down assay were used to illumine the mechanism of circRASSF2. Animal experiments were used to determine the role of circRASSF2 in the tumor growth of CRC in vivo. Our study reported that circRASSF2 was upregulated in CRC tissues and cells, and its high expression was related to the poor prognosis of CRC patients. CircRASSF2 knockdown could inhibit proliferation, migration, invasion, and enhance apoptosis in CRC cells, and its overexpression had the opposite effect. Besides, our data revealed that circRASSF2 could sponge miR-195-5p, and miR-195-5p could target FZD4. The rescue experiments indicated that both miR-195-5p inhibitor and FZD4 overexpression could reverse the negative regulation of circRASSF2 silencing on CRC progression. Moreover, circRASSF2 could positively regulate the activity of Wnt/β-catenin signaling pathway by the miR-195-5p/FZD4 axis. In addition, circRASSF2 knockdown restrained the tumor growth of CRC in vivo. Our findings suggested that circRASSF2 might function as a tumor promoter to accelerate the progression of CRC via regulating the miR-195-5p/FZD4/Wnt/β-catenin pathway.
环状 RNA（circRNA）是癌症进展的关键调节剂，包括结直肠癌（CRC）。然而，circRASSF2 在 CRC 中的作用尚不清楚。采用实时定量 PCR 检测 circRASSF2 和 miR-195-5p 的表达。细胞计数试剂盒 8 检测、集落形成实验、流式细胞术和 Transwell 实验分别用于检测细胞的增殖、凋亡、迁移和侵袭，Western blot 分析检测细胞增殖、转移和 Wnt/β-catenin 信号通路相关蛋白以及 Frizzled 4（FZD4）蛋白的水平。此外，还采用双荧光素酶报告基因实验、RNA 免疫沉淀实验和 RNA 下拉实验来阐明 circRASSF2 的作用机制。动物实验用于确定 circRASSF2 在 CRC 体内肿瘤生长中的作用。本研究报道 circRASSF2 在 CRC 组织和细胞中上调，其高表达与 CRC 患者的不良预后相关。circRASSF2 敲低可抑制 CRC 细胞的增殖、迁移、侵袭，促进细胞凋亡，而过表达则具有相反的作用。此外，我们的数据显示 circRASSF2 可以海绵 miR-195-5p，miR-195-5p 可以靶向 FZD4。挽救实验表明，miR-195-5p 抑制剂和 FZD4 过表达均可逆转 circRASSF2 沉默对 CRC 进展的负调控作用。此外，circRASSF2 可通过 miR-195-5p/FZD4 轴正向调节 Wnt/β-catenin 信号通路的活性。此外，circRASSF2 敲低可抑制 CRC 在体内的肿瘤生长。我们的研究结果表明，circRASSF2 可能通过调节 miR-195-5p/FZD4/Wnt/β-catenin 通路作为肿瘤促进剂加速 CRC 的进展。
