Aili Dilihumaer, Wu Tong, Gu Yuan, Chen Ziyuan, Wang Wanchun
Department of Orthopaedics, The Second Xiangya Hospital of Central South University, Changsha, Hunan 410011, P.R. China.
Exp Ther Med. 2021 May;21(5):455. doi: 10.3892/etm.2021.9886. Epub 2021 Mar 1.
Numerous studies have reported the critical roles of long non-coding RNAs (lncRNAs) in the regulation of osteoarthritis (OA) development. The present study aimed to assess the function and regulatory mechanism of a lncRNA, KCNQ1 opposite strand/antisense transcript 1 (KCNQ1OT1), in OA . C28/I2 cells were treated with lipopolysaccharide (LPS) to generate an OA model. The relative expression levels of KCNQ1OT1, microRNA (miR)-211-5p and transcription factor 4 (TCF4) were determined via reverse transcription-quantitative polymerase chain reaction. The associations between KCNQ1OT1, miR-211-5p and TCF4 were confirmed using a dual-luciferase reporter assay. Furthermore, cell viability was assessed using the MTT assay. Inflammatory cytokine levels were measured using ELISA. The protein expression levels of matrix metalloproteinase-3/13, collagen II/X and TCF4 were detected by western blotting. KCNQ1OT1 and TCF4 were highly expressed in the cartilage tissues of patients with OA and C28/I2 cells treated with LPS (OA cells), whereas miR-211-5p was downregulated concomitantly in OA tissues and cells. Knockdown of KCNQ1OT1 stimulated cell viability, and suppressed the inflammation and degradation of the extracellular matrix (ECM) in OA cells. In addition, overexpression of miR-211-5p stimulated cell viability, and inhibited inflammation and degradation of the ECM in OA cells. Notably, miR-211-5p was revealed to be the target of, and was negatively regulated by, KCNQ1OT1. TCF4 was targeted and negatively modulated by miR-211-5p. Transfection of cells with the miR-211-5p inhibitor or pcDNA-TCF4 reversed the suppressive effects of short hairpin RNA (sh)-KCNQ1OT1 on inflammation and ECM degradation, as well as the promotive effect of sh-KCNQ1OT1 on viability in OA . Therefore, KCNQ1OT1 may regulate the miR-211-5p/TCF4 axis to ameliorate OA .
众多研究报道了长链非编码RNA（lncRNA）在骨关节炎（OA）发展调控中的关键作用。本研究旨在评估一种lncRNA，即钾离子通道蛋白Q1反义转录本1（KCNQ1OT1）在OA中的功能及调控机制。用脂多糖（LPS）处理C28/I2细胞以构建OA模型。通过逆转录-定量聚合酶链反应测定KCNQ1OT1、微小RNA（miR）-211-5p和转录因子4（TCF4）的相对表达水平。使用双荧光素酶报告基因检测法确认KCNQ1OT1、miR-211-5p和TCF4之间的关联。此外，使用MTT法评估细胞活力。采用酶联免疫吸附测定法测量炎性细胞因子水平。通过蛋白质印迹法检测基质金属蛋白酶-3/13、胶原蛋白II/X和TCF4的蛋白表达水平。KCNQ1OT1和TCF4在OA患者软骨组织以及经LPS处理的C28/I2细胞（OA细胞）中高表达，而miR-211-5p在OA组织和细胞中同时下调。敲低KCNQ1OT1可促进细胞活力，并抑制OA细胞中细胞外基质（ECM）的炎症和降解。此外，miR-211-5p过表达可促进细胞活力，并抑制OA细胞中ECM的炎症和降解。值得注意的是，miR-211-5p被证实是KCNQ1OT1的靶标且受其负调控。TCF4是miR-211-5p的靶标并受其负向调节。用miR-211-5p抑制剂或pcDNA-TCF4转染细胞可逆转短发夹RNA（sh）-KCNQ1OT1对炎症和ECM降解以及sh-KCNQ1OT1对OA细胞活力的促进作用。因此，KCNQ1OT1可能通过调节miR-211-5p/TCF4轴来改善OA。
