Du Qiwei, Xue Jingsong, Fan Dongdong, Chen Junling, Zhong Guibin, Chen Jianwei
Department of Orthopaedics, Baoshan Branch, Ren Ji Hospital, School of Medicine Shanghai Jiao Tong University, No.1058, Huan Zhen Bei Road, Shanghai, 200444, China.
Department of Orthopaedics, Ren Ji Hospital, School of Medicine Shanghai Jiao Tong University, Shanghai, 200127, China.
J Mol Histol. 2025 Jul 31;56(4):242. doi: 10.1007/s10735-025-10528-x.
Intervertebral disc degeneration (IDD) significantly contributes to back pain, impacting patients' quality of life, and the cartilaginous endplate (CEP) is crucial to the functioning of the disc in both normal and disease states. Furthermore, circular RNAs (circRNAs) have been implicated in the modulation of a variety of diseases, including IDD. Nonetheless, the specific involvement of circRNAs in the the intervertebral disc CEP degeneration is not yet fully understood. The objective of this research was to explore the function and potential mechanism of circZNF418 in the context of CEP degeneration. We employed Western blotting, immunofluorescence, and qRT-PCR to evaluate the expression levels of circZNF418, its binding protein human antigen R (HuR), and Sox9. The cell counting Kit-8 (CCK-8) assay was employed to assess the functional impact of circZNF418 on cell viability. Bioinformatics analysis was conducted to identify the interaction between the RNA-binding protein HuR and circZNF418. Our findings indicated that the levels of expression for both circZNF418 and Sox9 were notably reduced in degenerative CEP cells, as well as in CEP cells subjected to oxidative stress. Furthermore, oxidative stress was found to decrease cell viability in CEP cells, reduce the expression of extracellular matrix (ECM)-related proteins (collagen II and aggrecan), and increase the levels of matrix metalloproteinases (MMP3 and MMP13). Notably, overexpression of circZNF418 mitigated these oxidative stress-induced effects. CircZNF418 overexpression also significantly enhanced the proliferation of CEP cells. Additionally, the overexpression of circZNF418 resulted in an increase in Sox9 mRNA and protein levels, while simultaneously decreasing the protein levels of MMP3 and MMP13, and increasing those of collagen II and aggrecan. We found that circZNF418 promotes the expression and protein translation of the downstream gene Sox9 by interacting with HuR. Conversely, interference with Sox9 reversed the inhibitory effects of circZNF418 on oxidative stress-induced degeneration of CEP cells. In conclusion, circZNF418 alleviates the degeneration of CEP cells caused by oxidative stress by stabilizing Sox9 through HuR interaction. Therefore, a deeper exploration of its molecular mechanism is crucial for advancing the prevention and treatment of IDD.
椎间盘退变（IDD）是导致背痛的重要原因，影响患者生活质量，而软骨终板（CEP）在椎间盘正常及疾病状态下的功能中都起着关键作用。此外，环状RNA（circRNAs）已被证明参与多种疾病的调控，包括IDD。然而，circRNAs在椎间盘CEP退变中的具体作用尚未完全明确。本研究旨在探讨circZNF418在CEP退变中的功能及潜在机制。我们采用蛋白质免疫印迹法、免疫荧光法和qRT-PCR来评估circZNF418、其结合蛋白人抗原R（HuR）和Sox9的表达水平。使用细胞计数试剂盒-8（CCK-8）检测法评估circZNF418对细胞活力的功能影响。通过生物信息学分析确定RNA结合蛋白HuR与circZNF418之间的相互作用。我们的研究结果表明，circZNF418和Sox9在退变的CEP细胞以及遭受氧化应激的CEP细胞中的表达水平均显著降低。此外，发现氧化应激会降低CEP细胞的活力，减少细胞外基质（ECM）相关蛋白（胶原蛋白II和聚集蛋白聚糖）的表达，并增加基质金属蛋白酶（MMP3和MMP13）的水平。值得注意的是，circZNF418的过表达减轻了这些氧化应激诱导的效应。circZNF418的过表达还显著增强了CEP细胞的增殖。此外，circZNF418的过表达导致Sox9 mRNA和蛋白水平增加，同时降低MMP3和MMP13的蛋白水平，并增加胶原蛋白II和聚集蛋白聚糖的水平。我们发现circZNF418通过与HuR相互作用促进下游基因Sox9的表达和蛋白质翻译。相反，干扰Sox9可逆转circZNF418对氧化应激诱导的CEP细胞退变的抑制作用。总之，circZNF418通过与HuR相互作用稳定Sox9，减轻氧化应激引起的CEP细胞退变。因此，深入探究其分子机制对于推进IDD的防治至关重要。
