LncRNA MEG3 Protects Chondrocytes From IL-1β-Induced Inflammation via Regulating miR-9-5p/KLF4 Axis

Yijiang Huang; Daosen Chen; Zijian Yan; Jingdi Zhan; Xinghe Xue; Xiaoyun Pan; Huachen Yu

doi:10.3389/fphys.2021.617654

LncRNA MEG3 Protects Chondrocytes From IL-1β-Induced Inflammation via Regulating miR-9-5p/KLF4 Axis

Front Physiol. 2021 Mar 11:12:617654. doi: 10.3389/fphys.2021.617654. eCollection 2021.

Authors

Yijiang Huang^{1

2}, Daosen Chen^{1

2}, Zijian Yan^{1

2}, Jingdi Zhan^{1

2}, Xinghe Xue^{1

2}, Xiaoyun Pan^{1

2}, Huachen Yu^{1

2}

Affiliations

¹ Department of Orthopaedics, The Second Affiliated Hospital and Yuying Children's Hospital of Wenzhou Medical University, Wenzhou, China.
² Key Laboratory of Orthopaedics of Zhejiang Province, Wenzhou, China.

Abstract

Background: Osteoarthritis (OA) is a chronic degenerative disease of the joints characterized by articular cartilage damage, subchondral bone remodeling, osteophyte formation, and inflammatory changes. This work aims to investigate the protective role of long non-coding RNA (lncRNA) maternally expressed 3 (MEG3) against the apoptosis of chondrocytes.

Methods: Chondrocyte cell lines, CHON-001, and ATDC5 were treated with different doses of interleukin-1β (IL-1β) to mimic the inflammatory response during OA pathogenesis. Quantitative real-time polymerase chain reaction was performed to measure MEG3, miR-9-5p, and Krüppel-like factor 4 (KLF4) mRNA expression levels. MEG3 and KLF4 overexpression plasmids, MEG3 shRNA, miR-9-5p mimics, and miR-9-5p inhibitors were transfected into the cells. Cell counting kit-8, wound healing assay, and flow cytometry were conducted to determine cell viability, migration, and apoptotic rate. Dual-luciferase reporter assay was adopted to verify the targeting relationships among MEG3, miR-9-5p, and KLF4. Western blot was used to detect KLF4 protein expression. Enzyme-linked immunosorbent assay was employed to measure the levels of inflammatory factors.

Results: MEG3 expression in chondrocytes was down-regulated by the stimulation of IL-1β, and MEG3 negatively regulated miR-9-5p expression but positively regulated KLF4 expression. MEG3 overexpression strengthened the viability and migration of CHON-001 and ATDC5 cells but restrained the apoptosis and inflammatory response, while MEG3 knockdown had opposite effects. miR-9-5p inhibition or KLF4 overexpression could counteract the effects of MEG3 knockdown on chondrocytes. Besides that, MEG3 was proved to be a molecular sponge for miR-9-5p, and KLF4 was verified as the target of miR-9-5p.

Conclusion: MEG3 can promote chondrocyte proliferation and migration and inhibit apoptosis and inflammation by sponging miR-9-5p to induce KLF4 expression, which provides a promising therapy target for OA treatment.

Keywords: KLF4; MEG3; OA; apoptosis; inflammation; miR-9-5p; proliferation.