环状 RNA ECE1 通过海绵吸附 miR-588 调控 RAB3D 促进骨肉瘤进展。

CircECE1 promotes osteosarcoma progression through regulating RAB3D by sponging miR-588.

机构信息

Department of Bone and Soft Tissue Oncology, Shanxi Province Cancer Hospital, Shanxi Hospital Affiliated to Cancer Hospital, Chinese Academy of Medical Sciences, Cancer Hospital Affiliated to Shanxi Medical University, No.3, Zhigong New Street, Xinghualing District, Taiyuan, 030013, China.

出版信息

J Orthop Surg Res. 2023 Aug 9;18(1):587. doi: 10.1186/s13018-023-04045-4.

DOI:10.1186/s13018-023-04045-4

PMID:37559140

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC10410784/

Abstract

BACKGROUND

Circular RNAs (circRNAs) have been confirmed to be involved in cancer pathogenesis. However, the underlying mechanism of circRNA endothelin converting enzyme 1 (circECE1) in osteosarcoma (OS) development is still not understood.

METHODS

The expression levels of circECE1, microRNA-588 (miR-588) and RAB3D, member RAS oncogene family (RAB3D) were gauged by quantitative real-time polymerase chain reaction (qRT-PCR) and western blot. OS cell proliferation was assessed by cell counting kit-8 (CCK-8) assay and 5-ethynyl-2'-deoxyuridine (EdU) assay. OS cell apoptosis rate and metastasis were identified by flow cytometry and transwell assay. Dual-luciferase reporter analysis and RNA immunoprecipitation (RIP) assay were performed to confirm the interactions among circECE1, miR-588 and RAB3D. Xenograft tumor models were established to explore circECE1 function in vivo. Immunohistochemistry (IHC) assay was applied to analyze RAB3D level after circECE1 knockdown.

RESULTS

In OS, circECE1 expression was higher than that in normal chondroma tissues. High levels of circECE1 were positively linked to OS cell viability, proliferation, migration and invasion, and negatively linked to OS cell apoptosis rate. It was found that circECE1 was a miR-588 sponge, and miR-588 inhibitor abrogated the influence of si-circECE1 on OS cells. MiR-588 targeted RAB3D to further regulate the pathological process of OS. Moreover, silencing circECE1 blocked OS tumor growth in vivo.

CONCLUSION

We elucidated the function of a novel circECE1/miR-588/RAB3D axis in OS progression.

摘要

背景

环状 RNA（circRNAs）已被证实参与癌症的发病机制。然而，circRNA 内皮素转换酶 1（circECE1）在骨肉瘤（OS）发展中的潜在机制尚不清楚。

方法

通过实时定量聚合酶链反应（qRT-PCR）和 Western blot 测定 circECE1、microRNA-588（miR-588）和 RAS 癌基因家族成员 RAB3D（RAB3D）的表达水平。通过细胞计数试剂盒-8（CCK-8）测定和 5-乙炔基-2'-脱氧尿苷（EdU）测定评估 OS 细胞增殖。通过流式细胞术和 Transwell 测定鉴定 OS 细胞凋亡率和转移。通过双荧光素酶报告分析和 RNA 免疫沉淀（RIP）分析证实 circECE1、miR-588 和 RAB3D 之间的相互作用。建立异种移植肿瘤模型以研究 circECE1 在体内的功能。免疫组织化学（IHC）测定用于分析 circECE1 敲低后 RAB3D 的水平。

结果

在 OS 中，circECE1 的表达高于正常软骨瘤组织。高水平的 circECE1 与 OS 细胞活力、增殖、迁移和侵袭呈正相关，与 OS 细胞凋亡率呈负相关。发现 circECE1 是 miR-588 的海绵体，miR-588 抑制剂消除了 si-circECE1 对 OS 细胞的影响。miR-588 靶向 RAB3D 以进一步调节 OS 的病理过程。此外，沉默 circECE1 阻断了 OS 肿瘤在体内的生长。