Background

Hypoxia is a key characteristic of osteosarcoma (OS). Increasing data suggested that circular RNA (circRNAs) were involve in the progression of cancers and the regulation of hypoxia, including OS. This study aims to examine the biological mechanism of circRNA cytochrome P450 family 51 subfamily A member 1 (circCYP51A1) in OS under hypoxia.

Methods

The expression levels of circCYP51A1, microRNA-490-3p (miR-490-3p) and kruppel-like factor 12 (KLF12) were detected by quantitative real-time polymerase chain reaction (qRT-PCR) in OS tissues and cells. Cell proliferation, migration and invasion were determined by colony formation assay and transwell assay. Lactate production and glucose consumption in OS cells were measured by using lactate assay kit and glucose assay kit, respectively. Western blot assay and immunohistochemistry assay were used to test protein levels. The associated relationship between miR-490-3p and circCYP51A1 or KLF12 was predicted using Starbase or DIANA online database and verified by dual-luciferase reporter assay. The xenograft model was used to explore the role of circCYP51A1 in vivo.

Results

CircCYP51A1 and KLF12 expression were dramatically increased, whereas miR-490-3p was decreased in OS cells under hypoxia condition. Deficiency of circCYP51A1 hindered hypoxia-induced cell proliferation, migration, invasion and glycolysis in OS cells. CircCYP51A1 enhanced KLF12 expression by sponging miR-490-3p. MiR-490-3p inhibitor weakened the inhibition effect of circCYP51A1 knockdown on the progression of OS under hypoxia. Besides, overexpression of miR-490-3p inhibited cell progression of OS under hypoxia condition, while the effects were attenuated by KLF12 overexpression. Importantly, knockdown of circCYP51A1 inhibited tumor growth in vivo.

Conclusion

CircCYP51A1 mediated cell proliferation, migration, invasion and glycolysis by regulating miR-490-3p/KLF12 axis in OS cells under hypoxia condition.

中文翻译：

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circCYP51A1的沉默通过调节缺氧条件下骨肉瘤中的miR-490-3p/KLF12轴来抑制细胞进程和糖酵解

背景

缺氧是骨肉瘤（OS）的一个关键特征。越来越多的数据表明，环状 RNA (circRNA) 参与了癌症的进展和缺氧的调节，包括 OS。本研究旨在探讨circRNA细胞色素P450家族51亚家族A成员1（circCYP51A1）在缺氧条件下在OS中的生物学机制。

方法

通过定量实时聚合酶链反应 (qRT-PCR) 检测 OS 组织和细胞中 circCYP51A1、microRNA-490-3p (miR-490-3p) 和 kruppel 样因子 12 (KLF12) 的表达水平。通过集落形成试验和transwell试验确定细胞增殖、迁移和侵袭。分别使用乳酸测定试剂盒和葡萄糖测定试剂盒测量OS细胞中的乳酸产生和葡萄糖消耗。蛋白质印迹测定和免疫组织化学测定用于测试蛋白质水平。使用 Starbase 或 DIANA 在线数据库预测 miR-490-3p 与 circCYP51A1 或 KLF12 之间的相关关系，并通过双荧光素酶报告基因分析验证。异种移植模型用于探索circCYP51A1在体内的作用。

结果

缺氧条件下 OS 细胞中 CircCYP51A1 和 KLF12 表达显着增加，而 miR-490-3p 降低。circCYP51A1 的缺乏会阻碍 OS 细胞中缺氧诱导的细胞增殖、迁移、侵袭和糖酵解。CircCYP51A1 通过海绵化 miR-490-3p 增强 KLF12 的表达。MiR-490-3p 抑制剂减弱了 circCYP51A1 敲低对缺氧条件下 OS 进展的抑制作用。此外，miR-490-3p的过表达抑制了缺氧条件下OS的细胞进展，而KLF12过表达则减弱了这种作用。重要的是，circCYP51A1 的敲低抑制了体内肿瘤的生长。

结论

CircCYP51A1 通过调节缺氧条件下 OS 细胞中的 miR-490-3p/KLF12 轴介导细胞增殖、迁移、侵袭和糖酵解。