环状 RNA ATRNL1 通过靶向 miR-153-3p 和 KLF5 对骨关节炎起保护作用。

CircATRNL1 protects against osteoarthritis by targeting miR-153-3p and KLF5.

机构信息

Department of Orthopaedics, the 1st Affiliated Hospital of Harbin Medical University, Harbin 150001, China.

出版信息

Int Immunopharmacol. 2021 Jul;96:107704. doi: 10.1016/j.intimp.2021.107704. Epub 2021 May 7.

DOI:10.1016/j.intimp.2021.107704

PMID:33971492

Abstract

BACKGROUND

Osteoarthritis (OA) is characterized by chondrocyte injury. Circular RNAs (circRNAs) are involved in the pathogenesis of various diseases, including OA. The purpose of this study was to determine the potential role of circATRNL1 in OA pathology in vitro.

METHODS

Human chondrocytes were isolated and treated with interleukin-1 beta (IL-1β) to mimic OA in vitro. High-throughput RNA sequencing was performed to identify differentially expressed circRNAs, miRNAs and mRNAs between IL and 1β-treated chondrocytes and normal chondrocytes. The expression of circATRNL1, miR-153-3p and KLF5 was measured using quantitative real-time polymerase chain reaction (qRT-PCR). For functional analyses, cell apoptosis was assessed using a flow cytometry assay. Extracellular matrix (ECM) degradation was monitored by measuring the levels of ECM-associated proteins by Western blot. The potential target miRNAs of circATRNL1 were screened by bioinformatics analysis and verified by dual-luciferase reporter assay.

RESULTS

The expression of circATRNL1 was decreased in IL-1β-treated chondrocytes. CircATRNL1 overexpression ameliorated cell apoptosis and ECM degradation, which were promoted by IL-1β treatment. Mechanistic analysis revealed that circATRNL1 directly targeted miR-153-3p and that miR-153-3p could reverse the inhibitory effects of circATRNL1 overexpression on inflammatory responses, cell apoptosis and ECM degradation. KLF5 is a target of miR-153-3p.

CONCLUSION

Taken together, the results in this study suggested that circATRNL1 might ameliorate the development and progression of OA through regulating miR-153-3p/KLF5 axis. Our study increased the understanding of circRNAs as therapeutic targets in the treatment of OA.

摘要

背景

骨关节炎（OA）的特征是软骨细胞损伤。环状 RNA（circRNAs）参与多种疾病的发病机制，包括 OA。本研究旨在确定 circATRNL1 在 OA 病理中的潜在作用。

方法

分离人软骨细胞并用白细胞介素-1β（IL-1β）处理以模拟 OA 体外。高通量 RNA 测序用于鉴定 IL-1β处理的软骨细胞与正常软骨细胞之间差异表达的 circRNAs、miRNAs 和 mRNAs。使用实时定量聚合酶链反应（qRT-PCR）测量 circATRNL1、miR-153-3p 和 KLF5 的表达。进行功能分析时，通过流式细胞术评估细胞凋亡。通过 Western blot 监测细胞外基质（ECM）降解，以测量 ECM 相关蛋白的水平。通过生物信息学分析筛选 circATRNL1 的潜在靶 miRNA，并通过双荧光素酶报告基因检测进行验证。

结果

IL-1β处理的软骨细胞中 circATRNL1 的表达降低。circATRNL1 过表达可改善 IL-1β处理促进的细胞凋亡和 ECM 降解。机制分析表明，circATRNL1 直接靶向 miR-153-3p，miR-153-3p 可逆转 circATRNL1 过表达对炎症反应、细胞凋亡和 ECM 降解的抑制作用。KLF5 是 miR-153-3p 的靶基因。