Deng Gang, Xu Yizhou, Li Zhengnan, Zeng Guangxuan
Department of Sports Medicine, Ganzhou People's Hospital, No.16 Meiguan Avenue, Zhanggong District, Ganzhou City, Jiangxi Province, China.
Department of Spinal Surgery, Orthopedic Medical Center, Zhujiang Hospital, Southern Medical University, Guangzhou, China.
J Orthop Surg Res. 2024 Dec 5;19(1):826. doi: 10.1186/s13018-024-05262-1.
Osteoarthritis (OA) poses a significant risk to the mobility of patients. Carbonic anhydrase 12 (CA12) can boost apoptosis and inflammation in several cancers, but its role in OA is unknown.
Differentially expressed genes in OA were analyzed using the GEO database (GSE169077). RT-qPCR and western blot estimated relative mRNA and protein levels of CA12. Cell viability and apoptosis were estimated by cell counting and flow cytometry assays. Oxidative stress (OxS) was determined by detecting with ROS and MDA levels, as well as CAT and SOD activities. Cytokine levels of IL-6 and TNF-α were detected by ELISA. Parameters associated with apoptosis and extracellular matrix (ECM) were detected by western blot. The m6A modification profile was determined by methylated RNA immunoprecipitation assays.
Relative CA12 and wilms' tumor 1-associating protein (WTAP) mRNA and protein levels were overexpressed in OA articular cartilages and IL-1β-challenged chondrocytes CHON-001. CA12 silencing impaired IL-1β-induced cell apoptosis, inflammation, OxS, and ECM degradation in chondrocytes. Yet, CA12 overexpression exerted an opposing function. WTAP reinforced the stability of CA12 mRNA depending on the m6A modification. Furthermore, WTAP knockdown weakened cell apoptosis, inflammation, OxS, and ECM degradation in chondrocytes induced by IL-1β, but these changes were impaired after CA12 overexpression. In addition, WTAP knockdown mitigates cartilage degeneration in DMM-induced mouse models.
IL-1β-induced WTAP enhances CA12 mRNA stability depending on m6A modification, thus promoting chondrocyte apoptosis, inflammatory response, OxS, and ECM degradation, providing evidence to support the possibility of WTAP and CA12 as potential targets for OA treatment.
骨关节炎（OA）对患者的活动能力构成重大风险。碳酸酐酶12（CA12）可促进多种癌症中的细胞凋亡和炎症反应，但其在OA中的作用尚不清楚。
使用GEO数据库（GSE169077）分析OA中差异表达的基因。RT-qPCR和蛋白质免疫印迹法估计CA12的相对mRNA和蛋白质水平。通过细胞计数和流式细胞术检测细胞活力和凋亡。通过检测活性氧（ROS）和丙二醛（MDA）水平以及过氧化氢酶（CAT）和超氧化物歧化酶（SOD）活性来确定氧化应激（OxS）。通过酶联免疫吸附测定（ELISA）检测白细胞介素-6（IL-6）和肿瘤坏死因子-α（TNF-α）的细胞因子水平。通过蛋白质免疫印迹法检测与细胞凋亡和细胞外基质（ECM）相关的参数。通过甲基化RNA免疫沉淀测定法确定m6A修饰谱。
在OA关节软骨和白细胞介素-1β刺激的软骨细胞CHON-001中，CA12和肾母细胞瘤1相关蛋白（WTAP）的相对mRNA和蛋白质水平过表达。CA12沉默可损害白细胞介素-1β诱导的软骨细胞凋亡、炎症、氧化应激和细胞外基质降解。然而，CA12过表达则发挥相反的作用。WTAP通过m6A修饰增强了CA12 mRNA的稳定性。此外，WTAP敲低减弱了白细胞介素-1β诱导的软骨细胞凋亡、炎症、氧化应激和细胞外基质降解，但在CA12过表达后这些变化受到损害。此外，WTAP敲低减轻了在半月板切除（DMM）诱导的小鼠模型中的软骨退变。
白细胞介素-1β诱导的WTAP通过m6A修饰增强了CA12 mRNA的稳定性，从而促进软骨细胞凋亡、炎症反应、氧化应激和细胞外基质降解，为支持WTAP和CA12作为OA治疗潜在靶点的可能性提供了证据。
