血管生成素-2 通过 HIF-1α/NF-κB 信号通路促进纤维环的机械应激诱导的细胞外基质降解。
Angiopoietin-2 Promotes Mechanical Stress-induced Extracellular Matrix Degradation in Annulus Fibrosus Via the HIF-1α/NF-κB Signaling Pathway.
机构信息
Division of Spine Surgery, Department of Orthopaedics, Nanfang Hospital of Southern Medical University, Guangzhou, Guangdong, China.
Department of Spine Surgery, Center for Orthopedic Surgery, The Third Affiliated Hospital of Southern Medical University, Guangzhou, Guangdong, China.
出版信息
Orthop Surg. 2023 Sep;15(9):2410-2422. doi: 10.1111/os.13797. Epub 2023 Jul 21.
OBJECTIVE
Mechanical stress is an important risk factor for intervertebral disc degeneration (IVDD). Angiopoietin-2 (ANG-2) is regulated by mechanical stress and is widely involved in the regulation of extracellular matrix metabolism. In addition, the signaling cascade between HIF-1α and NF-κB is critical in matrix degradation. This study aims to investigate the role and molecular mechanism of ANG-2 in regulating the degeneration of annulus fibrosus (AF) through the HIF-1α/NF-κB signaling pathway.
METHODS
The bipedal standing mice IVDD model was constructed, and histological experiments were used to evaluate the degree of IVDD and the expression of ANG-2 in the AF. Mouse primary AF cells were extracted in vitro and subjected to mechanical stretching experiments. Western blot assay was used to detect the effect of mechanical stress on ANG-2, and the role of the ANG-2-mediated HIF-1α/NF-κB pathway in matrix degradation. In addition, the effect of inhibiting ANG-2 expression by siRNA or monoclonal antibody on delaying IVDD was investigated at in vitro and in vivo levels. One-way ANOVA with the least significant difference method was used for pairwise comparison of the groups with homogeneous variance, and Dunnett's method was used to compare the groups with heterogeneous variance.
RESULTS
In IVDD, the expressions of catabolic biomarkers (mmp-13, ADAMTS-4) and ANG-2 were significantly increased in AF. In addition, p65 expression was increased while HIF-1α expression was significantly decreased. The results of western blot assay showed mechanical stress significantly up-regulated the expression of ANG-2 in AF cells, and promoted matrix degradation by regulating the activity of HIF-1α/NF-κB pathway. Exogenous addition of Bay117082 and CoCl inhibited matrix degradation caused by mechanical stress. Moreover, injection of neutralizing antibody or treatment with siRNA to inhibit the expression of ANG-2 improved the matrix metabolism of AF and inhibited IVDD progression by regulating the HIF-1α/NF-κB signaling pathway.
CONCLUSION
In IVDD, mechanical stress could regulate the HIF-1α/NF-κB signaling pathway and matrix degradation by mediating ANG-2 expression in AF degeneration.
目的
机械应力是椎间盘退变（IVDD）的一个重要危险因素。血管生成素-2（ANG-2）受机械应力调节，广泛参与细胞外基质代谢的调节。此外，HIF-1α和 NF-κB 信号级联在基质降解中至关重要。本研究旨在通过 HIF-1α/NF-κB 信号通路探讨 ANG-2 在调节纤维环（AF）退变中的作用和分子机制。
方法
构建双足站立小鼠 IVDD 模型，组织学实验评估 IVDD 程度和 AF 中 ANG-2 的表达。体外提取小鼠原代 AF 细胞，进行机械拉伸实验。Western blot 检测机械应力对 ANG-2 的影响，以及 ANG-2 介导的 HIF-1α/NF-κB 通路在基质降解中的作用。此外，在体外和体内水平研究通过 siRNA 或单克隆抗体抑制 ANG-2 表达对延缓 IVDD 的作用。同方差组间采用最小显著差法进行单向方差分析，异方差组间采用 Dunnett 法进行比较。
结果
在 IVDD 中，AF 中 catabolic 生物标志物（mmp-13、ADAMTS-4）和 ANG-2 的表达明显增加。此外，p65 表达增加，而 HIF-1α 表达明显降低。Western blot 结果显示，机械应力显著上调 AF 细胞中 ANG-2 的表达，并通过调节 HIF-1α/NF-κB 通路的活性促进基质降解。外源性添加 Bay117082 和 CoCl 抑制机械应力引起的基质降解。此外，注射中和抗体或用 siRNA 抑制 ANG-2 的表达，通过调节 HIF-1α/NF-κB 信号通路改善 AF 的基质代谢，抑制 IVDD 进展。
结论
在 IVDD 中，机械应力可通过调节 AF 退变中 ANG-2 的表达来调节 HIF-1α/NF-κB 信号通路和基质降解。
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