Department of Orthopedics, Taizhou Clinical Medical School of Nanjing Medical University (Taizhou People's Hospital), Taizhou, People's Republic of China.
Department of Orthopedics, Jinling Hospital, Jinling Clinical Medical College of Nanjing Medical University, Nanjing, People's Republic of China.
Bioengineered. 2021 Dec;12(1):7020-7032. doi: 10.1080/21655979.2021.1971483.
Previous research has identified N-myc downstream-regulated gene 2 (NDRG2) as one of the differentially expressed genes common to rat models of osteoarthritis (OA) and human OA. The purpose of this study was to investigate the role of NDRG2 in OA. In this study, an in vitro OA model was constructed by challenging ATDC5 chondrocytes with 10 ng/ml IL-1β. After transfection of pcDNA3.1(+)/NDRG2, qPCR and western blot were performed to assay NDRG2 expression. The analyses of cell viability, apoptosis and inflammatory molecule expression were employed respectively by CCK-8, TUNEL and ELISA. The protein expression related to apoptosis, inflammation or extracellular matrix (ECM) degradation was detected by western blot. The binding of Krüppel-like factor 5 (KLF5) to NDRG2 promoter was verified by means of dual-luciferase reporter assay. After overexpression of both NDRG2 and KLF5 in IL-1β-stimulated ATDC5 chondrocytes, corresponding assays were performed to examine cell viability, apoptosis, inflammatory response and ECM degradation. In ATDC5 chondrocytes challenged with IL-1β, NDRG2 expression was much lower than that in the control group, whereas it's overexpression helped restored cell viability and reduce cell apoptosis, inflammatory response and ECM degradation. It was also observed that KLF5 expression was decreased in IL-1β-stimulated ATDC5 chondrocytes, and that KLF5 bound to the NDRG2 promoter. Importantly, overexpressing KLF5 could reverse the protective effect of NDRG2 overexpression on IL-1β-stimulated ATDC5. Overall, NDRG2 could be transcriptionally regulated by transcription factor KLF5 and may play a protective role against chondrocyte the inflammatory response and ECM degradation in OA.
先前的研究已经确定 N- myc 下游调节基因 2（NDRG2）是骨关节炎（OA）大鼠模型和人类 OA 共有的差异表达基因之一。本研究旨在探讨 NDRG2 在 OA 中的作用。在这项研究中，通过用 10ng/ml IL-1β 刺激 ATDC5 软骨细胞构建体外 OA 模型。转染 pcDNA3.1（+）/NDRG2 后，通过 qPCR 和 Western blot 检测 NDRG2 表达。通过 CCK-8、TUNEL 和 ELISA 分别分析细胞活力、凋亡和炎症分子表达。通过 Western blot 检测与凋亡、炎症或细胞外基质（ECM）降解相关的蛋白表达。通过双荧光素酶报告基因检测验证 Krüppel 样因子 5（KLF5）与 NDRG2 启动子的结合。在 IL-1β 刺激的 ATDC5 软骨细胞中过表达 NDRG2 和 KLF5 后，进行相应的检测以检查细胞活力、凋亡、炎症反应和 ECM 降解。在受 IL-1β 刺激的 ATDC5 软骨细胞中，NDRG2 表达明显低于对照组，而过表达有助于恢复细胞活力并减少细胞凋亡、炎症反应和 ECM 降解。还观察到 KLF5 在 IL-1β 刺激的 ATDC5 软骨细胞中表达降低，并且 KLF5 与 NDRG2 启动子结合。重要的是，过表达 KLF5 可以逆转 NDRG2 过表达对 IL-1β 刺激的 ATDC5 的保护作用。总的来说，NDRG2 可能受到转录因子 KLF5 的转录调控，并且在 OA 中可能对软骨细胞的炎症反应和 ECM 降解发挥保护作用。
