Circular RNA Atp9b, a competing endogenous RNA, regulates the progression of osteoarthritis by targeting miR-138-5p.

Osteoarthritis (OA) is the most common joint disease and is mainly characterized by degradation of the articular cartilage. Recently, circular RNAs (circRNAs), novel noncoding RNAs with different biological functions and pathological implications, have been reported to be closely associated with various diseases. Growing evidence indicates that circRNAs act as competing endogenous RNAs (ceRNAs) that bind with microRNAs (miRNAs) and regulate their downstream functions. Here, we identified a new circRNA, circRNA_Atp9b, and further investigated its function in OA using a well-established mouse chondrocyte model. We demonstrated that circRNA_Atp9b expression was significantly up-regulated in mouse chondrocytes after stimulation with interleukin-1 beta (IL-1β), and that knockdown of circRNA_Atp9b promoted the expression of type II collagen while inhibiting the generation of MMP13, COX-2 and IL-6. Moreover, there was a negative correlation between the expression levels of circRNA_Atp9b and microRNA (miR)-138-5p, indicating that miR-138-5p also played a role in IL-1β-induced chondrocytes. Bioinformatics analysis predicted circRNA_Atp9b directly target miR-138-5p, which was validated by dual-luciferase assay. Further functional experiments revealed that down-regulation of miR-138-5p partly reversed the effects of circRNA_Atp9b on extracellular matrix (ECM) catabolism and inflammation. Taken together, these results suggest that circRNA_Atp9b regulates OA progression by modulating ECM catabolism and inflammation in chondrocytes via sponging miR-138-5p. Our findings provide novel insight into the regulatory mechanism of circRNA_Atp9b in OA and may contribute to establishing potential therapeutic strategies.

Zhou ZB ，Du D ，Huang GX ，Chen A ，Zhu L ... -  《-》

Circular RNA expression profile of articular chondrocytes in an IL-1β-induced mouse model of osteoarthritis.

Osteoarthritis (OA) is a widely prevalent degenerative joint disease characterized by articular cartilage degradation and joint inflammation. The pathogenesis of OA remains unclear, leading to a lack of effective treatment. Previous studies have reported that circular RNAs (circRNAs) are involved in the development of various diseases. However, the function of circRNAs and their roles in OA is largely unknown. Therefore, we aimed to investigate changes in circRNA expression and predict their functions in OA by using bioinformatics analysis. An OA model was established in mouse articular chondrocytes (MACs) treated by interleukin-1β (IL-1β), and then the circRNA profile was screened by Next Generation Sequencing. By comparing circRNA expression in IL-1β- treated MACs and normal controls, differentially expressed circRNAs were identified during OA pathogenesis, and differential expression levels of selected circRNAs were validated by qRT-PCR. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were employed to predict the functions of these circRNAs. Because circRNAs can act as "miRNA sponges", we also constructed a circRNA-miRNA network to predict their functions. A total of 255 circRNAs were found to be differentially expressed in IL-1β-treated MACs (p≤0.05; fold-change≥2) from the expression of the normal controls. Among them, 119 circRNAs were significantly up-regulated, and the other 136 were down-regulated. Seven circRNAs were randomly selected to verify the reliability of these profiles by quantitative qRT-PCR. After obtaining the parental genes of differentially expressed circRNA, the top 30 enrichment GO entries and KEGG pathways were annotated. Then, two significantly differentially expressed circRNAs (mmu-circRNA-30365 and mmu-circRNA-36866) were identified and selected for further analysis, meanwhile a circRNA-miRNA regulation network was created and the top five most likely functional-related target miRNAs of the circRNAs were collected. Although the exact mechanisms and biological functions of these circRNAs in the development of OA need further exploration, our findings do suggest that the differentially expressed circRNAs were involved in the pathogenesis of OA. Thus, our study brings us closer to understanding the pathogenic mechanisms and finding new molecular targets for the clinical treatment of osteoarthritis.

Zhou Z ，Du D ，Chen A ，Zhu L ... -  《-》

CircRNA hsa_circ_0005105 upregulates NAMPT expression and promotes chondrocyte extracellular matrix degradation by sponging miR-26a.

Osteoarthritis (OA) is a chronic disease pathologically characterized by articular cartilage degeneration and damage. Currently, studies have found that circular RNA (circRNA) is involved in intracellular RNA regulating network and is closely related to the occurrence and development of diseases, therefore it may become a new biological marker and therapeutic target. After stimulating chondrocytes with interleukin-1 beta (IL-1β), hsa_circ_0005105 expression was significantly upregulated, while miR-26a expression was significantly inhibited. Hsa_circ_0005105 did not influence miR-26a expression but inhibited its transcriptional activity so as to upregulate the expression of its target NAMPT. Studies further indicated that hsa_circ_0005105 can inhibit the expression of type II collagen and aggrecan, promote the expression of MMP-13 and ADAMTS-4, and the generation of PGE2, IL-6, and IL-8, but the linear sequence of hsa_circ_0005105 cannot. MiR-26a has the opposite effect, and hsa_circ_0005105 can antagonize the function of miR-26a. When NAMPT expression was downregulated, the above function of hsa_circ_0005105 was significantly weakened. Therefore, hsa_circ_0005105 can promote extracellular matrix (ECM) degradation by regulating the expression of miR-26a target NAMPT. These findings will provide new targets for treatment and prevention of OA and other orthopedic diseases.

Wu Y ，Zhang Y ，Zhang Y ，Wang JJ ... -  《-》

Circ_DHRS3 positively regulates GREM1 expression by competitively targeting miR-183-5p to modulate IL-1β-administered chondrocyte proliferation, apoptosis and ECM degradation.

Osteoarthritis (OA) is a chronic inflammatory disease caused by degenerative changes of articular cartilage, involving in the expression changes of special circular RNAs (circRNAs). This study aimed to explore the role of circ_DHRS3 in OA cell models and provide a potential mechanism. OA cell models were constructed using human chondrocytes with Interleukin-1 beta (IL-1β) treatment. The expression of circ_DHRS3, microRNA (miR)-183-5p and Gremlin 1 (GREM1) mRNA was detected using real-time quantitative polymerase chain reaction (RT-qPCR). Cell proliferation was identified using 3-[4, 5-dimethylthiazol-2-yl]-2, 5 diphenyl tetrazolium bromide (MTT) assay. Cell apoptosis was investigated using flow cytometry assay. The protein levels of proliferation- and apoptosis-related proteins were quantified by western blot. The levels of extracellular matrix (ECM)-associated proteins were quantified by western blot to assess ECM degradation. The relationship between miR-183-5p and circ_DHRS3 or GREM1 was predicted and then verified by dual-luciferase reporter assay. Circ_DHRS3 expression was elevated in OA cartilage tissues and IL-1β-treated chondrocytes. Circ_DHRS3 was resistant to RNase R and Actinomycin D. Circ_DHRS3 knockdown promoted chondrocyte proliferation inhibited by IL-1β, and alleviated IL-1β-induced apoptosis and ECM degradation, which were reversed by the inhibition of miR-183-5p, a target of circ_DHRS3. MiR-183-5p restoration also enhanced IL-1β-blocked cell proliferation, and relieved IL-1β-induced cell apoptosis and ECM degradation, while GREM1 (a target of miR-183-5p) overexpression abolished the effects of miR-183-5p restoration. Moreover, circ_DHRS3 regulated GREM1 expression by targeting miR-183-5p. Circ_DHRS3 mediated IL-1β-administered chondrocyte proliferation, apoptosis and ECM degradation by positively regulating GREM1 expression via competitively targeting miR-183-5p.

Jiang R ，Gao H ，Cong F ，Zhang W ，Song T ，Yu Z ... -  《-》

CircRNA circ_SEC24A upregulates DNMT3A expression by sponging miR-26b-5p to aggravate osteoarthritis progression.

Osteoarthritis (OA) is a chronic degenerative disease characterized by degeneration and injury of articular cartilage. Circular RNA_SEC24A (circ_SEC24A; circBase ID: hsa_circ_0005105) is upregulated and promotes multiple tumor processes. However, its role in OA progression remained mostly unknown. Quantitative real-time PCR (qRT-PCR) was used to detect the RNA expression of circ_SEC24A, miR-26b-5p and DNA methyltransferase 3 alpha (DNMT3A). Cell proliferation was verified by 3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2-H-tetrazolium bromide (MTT) and 5-ethynyl-2'-deoxyuridine (EdU) assays. Flow cytometry was used to detect apoptosis. Western blot was used to detect protein expression of DNMT3A, proliferating cell nuclear antigen (PCNA), extracellular matrix (ECM) proteins (Collagen II and Aggrecan), and ECM degrading enzymes (matrix metalloproteinase-13 [MMP13] and metallopeptidase with thrombospondin type 1 motif 5 [ADAMTS5]). The target relationship between miR-26b-5p and circ_SEC24A or DNMT3A was predicted by Statbase3.0 or TargetScan and confirmed by dual-luciferase reporter assay, RNA pull-down assay and RNA immunoprecipitation. Circ_SEC24A was upregulated in osteoarthritic cartilage tissues and IL-1β-induced chondrocytes, accompanying with miR-26b-5p downregulation and DNMT3A upregulation. Circ_SEC24A expression was resistant to RNase R digestion and mainly expressed in the cytoplasm. Interfering circ_SEC24A abolished IL-1β-induced effects on proliferation inhibition, apoptosis, and ECM degradation in chondrocytes, but overexpressing circ_SEC24A had the opposite effects. Inhibiting miR-26b-5p counteracted but upregulating miR-26a-5p mimicked the functions of circ_SEC24A silencing. Reinforcing DNMT3A reversed miR-26b-5p overexpression's role in IL-1β-induced chondrocytes. Mechanically, circ_SEC24A and DNMT3A were competitive endogenous RNAs (ceRNAs) for miR-26b-5p. Circ_SEC24A was a promoting factor for IL-1β-induced OA progression via circ_SEC24A/miR-26b-5p/DNMT3A ceRNA axis.

Zhang Z ，Yang B ，Zhou S ，Wu J ... -  《-》