Background

Colorectal cancer (CRC) is one of the most common malignancies worldwide and a major threat to human life and health. Circular RNA (circRNA)-microRNA (miRNA)-mRNA mechanism is considered to occur in various cancers. However, the mechanism of circ_101692 in CRC is still unclear.

Methods

Quantitative real-time PCR was used to detect the expression of circ_101692, miR-449b-5p and Golgi phosphoprotein 3 (GOLPH3) in CRC tissues and cell lines. Cell proliferation was determined using cell counting kit-8 (CCK8), colony formation assay and 5-ethynyl-2′-deoxyuridine (EdU). Flow cytometry and transwell were performed to measure apoptosis and cell invasion, respectively. Besides, glucose uptake and lactate production were tested using commercial kits to determine the glycolytic of CRC. Furthermore, dual-luciferase reporter assay and RNA immunoprecipitation assay were employed to verify the relationship between miR-449b-5p and circ_101692 or GOLPH3. And the protein levels were monitored using western blot assay. The xenotransplantation model was established to study the role of circ_101692 in vivo.

Results

Circ_101692 and GOLPH3 were highly expressed, while miR-449b-5p expression was down-regulated in CRC tissues and cell lines compared to normal tissues and cell lines. Circ_101692 negatively targeted miR-449b-5p, and GOLPH3 was the downstream gene of miR-449b-5p. Silencing circ_101692 suppressed CRC cell proliferation, invasion and glycolysis, as well as promoted apoptosis, while these influences could be reverted by miR-449b-5p inhibitor. Similarly, overexpression of GOLPH3 abolished the influence of miR-449b-5p mimic on CRC cell behavior. In addition, silencing of circ_101692 restricted CRC tumor growth in vivo.

Conclusion

Our results showed that circ_101692 promoted CRC progression by modulating the miR-449b-5p/GOLPH3 axis, implying that circ_101692 might be a possible target for CRC treatment.

Graphical abstract

中文翻译：

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Circ_101692通过调控miR-449b-5p/GOLPH3轴促进结直肠癌增殖、侵袭和糖酵解

背景

结直肠癌（CRC）是全球最常见的恶性肿瘤之一，对人类生命和健康构成重大威胁。环状RNA（circRNA）-微小RNA（miRNA）-mRNA机制被认为发生在多种癌症中。然而，circ_101692在CRC中的作用机制仍不清楚。

方法

采用实时定量PCR检测CRC组织和细胞系中circ_101692、miR-449b-5p和高尔基体磷蛋白3（GOLPH3）的表达。使用细胞计数试剂盒-8 (CCK8)、集落形成测定和 5-乙炔基-2'-脱氧尿苷 (EdU) 测定细胞增殖。流式细胞仪和transwell分别测量细胞凋亡和细胞侵袭。此外，使用商业试剂盒测试葡萄糖摄取和乳酸产生，以确定CRC的糖酵解。此外，采用双荧光素酶报告基因测定和RNA免疫沉淀测定来验证miR-449b-5p与circ_101692或GOLPH3之间的关系。并使用蛋白质印迹测定法监测蛋白质水平。建立异种移植模型来研究circ_101692在体内的作用。

结果

与正常组织和细胞系相比，CRC组织和细胞系中Circ_101692和GOLPH3高表达，而miR-449b-5p表达下调。Circ_101692负向靶向miR-449b-5p，GOLPH3是miR-449b-5p的下游基因。沉默circ_101692可抑制CRC细胞增殖、侵袭和糖酵解，并促进细胞凋亡，而这些影响可被miR-449b-5p抑制剂逆转。同样，GOLPH3 的过度表达消除了 miR-449b-5p 模拟物对 CRC 细胞行为的影响。此外，circ_101692的沉默限制了体内CRC肿瘤的生长。

结论

我们的结果表明，circ_101692 通过调节 miR-449b-5p/GOLPH3 轴促进 CRC 进展，这意味着 circ_101692 可能是 CRC 治疗的可能靶点。

图形概要