Department of Orthopedics, Huizhou Central People's Hospital, Huizhou, Guangdong Province, China.
Department of Trauma Surgery, Huizhou Central People's Hospital, Huizhou, Guangdong Province, China.
Innate Immun. 2022 Feb;28(2):79-90. doi: 10.1177/17534259221077078.
The aim of this study was to explore the role of hsa_circRNA_0000205 (circ_0000205) in chondrocyte injury in osteoarthritis (OA) and the underlying mechanism. Expression of circ_0000205, microRNA (miR)-766-3p and a disintegrin and metalloproteinase with thrombospondin motif (ADAMTS)-5 was detected by quantitative real time (qRT)-polymerase chain reaction (PCR) and Western blot assays. Cell proliferation, apoptosis, and extracellular matrix (ECM) synthesis were examined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and 5-ethynyl-2-deoxyuridine assays, flow cytometry, and qRT-PCR and Western blot assays. The target relationship between miR-766-3p and circ_0000205 or ADAMTS5 was confirmed by luciferase reporter assay and RNA immunoprecipitation. IL-1β treatment could attenuate cell viability of primary chondrocytes and proliferating cell nuclear antigen (PCNA) and collagen II type alpha-1 (COL2A1) levels, and elevate apoptosis rate and cleaved caspase-3, ADAMTS5 and matrix metalloproteinase-13 (MMP13) levels, suggesting that IL-1β induced chondrocyte apoptosis and ECM degradation. Expression of circ_0000205 was up-regulated in OA tissues and IL-1β-induced primary chondrocytes, accompanied with miR-766-3p down-regulation and ADAMTS5 up-regulation. Knockdown of circ_0000205 could mitigate IL-1β-induced above effects and improve cell proliferation. Moreover, both depleting miR-766-3p and promoting ADAMTS5 could partially counteract circ_0000205 knockdown roles in IL-1β-cultured primary chondrocytes. Notably, circ_0000205 was verified as a sponge for miR-766-3p via targeting, and ADAMTS5 was a direct target for miR-766-3p. Silencing circ_0000205 could protect chondrocytes from IL-1β-induced proliferation reduction, apoptosis, and ECM degradation by targeting miR-766-3p/ADAMTS5 axis.
本研究旨在探讨 hsa_circRNA_0000205(circ_0000205)在骨关节炎(OA)软骨细胞损伤中的作用及其机制。通过实时定量聚合酶链反应(qRT-PCR)和 Western blot 检测 circ_0000205、微小 RNA(miR)-766-3p 和解整合素金属蛋白酶与凝血酶样 5(ADAMTS)-5 的表达。通过 3-(4,5-二甲基噻唑-2-基)-2,5-二苯基四氮唑溴盐和 5-乙炔基-2-脱氧尿苷测定、流式细胞术、qRT-PCR 和 Western blot 检测细胞增殖、凋亡和细胞外基质(ECM)合成。通过荧光素酶报告基因检测和 RNA 免疫沉淀证实了 miR-766-3p 与 circ_0000205 或 ADAMTS5 的靶关系。IL-1β 处理可降低原代软骨细胞增殖细胞核抗原(PCNA)和胶原 II 型α-1(COL2A1)水平,增加细胞凋亡率和 cleaved caspase-3、ADAMTS5 和基质金属蛋白酶 13(MMP13)水平,表明 IL-1β 诱导软骨细胞凋亡和 ECM 降解。OA 组织和 IL-1β 诱导的原代软骨细胞中 circ_0000205 的表达上调,同时伴有 miR-766-3p 下调和 ADAMTS5 上调。circ_0000205 敲低可减轻 IL-1β 诱导的上述作用并改善细胞增殖。此外,miR-766-3p 耗竭和 ADAMTS5 促进均可部分拮抗 IL-1β 培养的原代软骨细胞中 circ_0000205 敲低的作用。值得注意的是,circ_0000205 通过靶向被验证为 miR-766-3p 的海绵,而 ADAMTS5 是 miR-766-3p 的直接靶标。沉默 circ_0000205 可通过靶向 miR-766-3p/ADAMTS5 轴保护软骨细胞免受 IL-1β 诱导的增殖减少、凋亡和 ECM 降解。