Xu Fei, Hu Qun-Fang, Li Jia, Shi Chang-Jiang, Luo Jin-Wei, Tian Wei-Chao, Pan Li-Wei
Department of Orthopedics, Affiliated Hospital of Chengde Medical College, Chengde 067000, Hebei Province, People's Republic of China.
Department of Orthopedics, Affiliated Hospital of Chengde Medical College, Chengde 067000, Hebei Province, People's Republic of China.
Cytokine. 2022 Apr;152:155805. doi: 10.1016/j.cyto.2022.155805. Epub 2022 Feb 21.
To clarify the expression and underlying network of long non-coding RNA (lncRNA) MCM3AP-AS1 in osteoarthritis (OA).
Human articular cartilage samples, OA model rats and IL-1β-treated C28/I2 cells were used in this study. The expression changes of genes and proteins were assessed by real-time quantitative PCR (qRT-PCR) and western blot. Cell viability, apoptosis, autophagy and extracellular matrix (ECM) degradation were assessed by Cell Counting Kit-8 (CCK-8), immunohistochemistry (IHC), flow cytometry, immunofluorescence and western blot assays, respectively. Molecule interactions were validated by dual luciferase and Chromatin immunoprecipitation (ChIP) assays. H&E staining was used to detect the pathological changes of cartilage.
MCM3AP-AS1 was upregulated in OA patients and IL-1β-induced chondrocytes. Knockdown of MCM3AP-AS1 enhanced autophagy, while alleviated ECM degradation and cartilage injury. Mechanistically, overexpression of SOX4 boosted the transcription of MCM3AP-AS1. Moreover, MCM3AP-AS1 functioned as a molecular sponge or epigenetic regulator of miR-149-5p to facilitate Notch1 expression. Functional rescue experiments showed that either inhibition of miR-149-5p nor ectopic expression of Notch1 dramatically weakened the biological impacts of MCM3AP-AS1 silencing.
These finding demonstrated that SOX4-activated MCM3AP-AS1 aggravated OA progression by modulating autophagy and ECM degradation via targeting miR-149-5p/Notch1 axis. These data supported that inhibition of MCM3AP-AS1 might be a potential treatment strategy of OA.
阐明长链非编码RNA（lncRNA）MCM3AP-AS1在骨关节炎（OA）中的表达及潜在调控网络。
本研究使用人关节软骨样本、OA模型大鼠及白细胞介素-1β（IL-1β）处理的C28/I2细胞。通过实时定量聚合酶链反应（qRT-PCR）和蛋白质免疫印迹法评估基因和蛋白质的表达变化。分别采用细胞计数试剂盒-8（CCK-8）、免疫组织化学（IHC）、流式细胞术、免疫荧光和蛋白质免疫印迹法检测细胞活力、凋亡、自噬及细胞外基质（ECM）降解情况。通过双荧光素酶和染色质免疫沉淀（ChIP）试验验证分子间相互作用。采用苏木精-伊红（H&E）染色检测软骨的病理变化。
MCM3AP-AS1在OA患者及IL-1β诱导的软骨细胞中表达上调。敲低MCM3AP-AS1可增强自噬，同时减轻ECM降解及软骨损伤。机制上，SRY-box转录因子4（SOX4）过表达可促进MCM3AP-AS1的转录。此外，MCM3AP-AS1作为微小RNA-149-5p（miR-149-5p）的分子海绵或表观遗传调节因子，促进Notch受体1（Notch1）表达。功能挽救实验表明，抑制miR-149-5p或异位表达Notch1均未显著减弱MCM3AP-AS1沉默的生物学效应。
这些发现表明，SOX4激活的MCM3AP-AS1通过靶向miR-149-5p/Notch1轴调节自噬和ECM降解，从而加重OA进展。这些数据支持抑制MCM3AP-AS1可能是OA的一种潜在治疗策略。
