基于应力调控探讨摇法诱导滑膜细胞炎症信号通路调节膝骨关节炎作用机制

Mechanistic study on stress-modulated regulation of knee osteoarthritis via shaking manipulation-induced synovial cell inflammatory signaling pathways

关键词:  摇法  膝骨关节炎  滑膜炎  FLSs  信号通路  应力

Key words:  shaking manipulation  KOA  synovitis  FLSs  signaling pathway  stress

摘要:

[目的] 探讨推拿摇法通过调控滑膜细胞αVβ3整合素（αVβ3）/黏着斑激酶（FAK）/细胞外信号调节激酶（ERK）/转录因子C-Jun（C-Jun）信号通路改善膝骨关节炎滑膜炎症的作用机制。[方法] 采用随机数字表法将100只雄兔分为5组：正常组（Normal）、假手术组（Sham）、模型组（Model）、摇法组（Shaking）和塞来昔布对照组（Celecoxib），每组20只。关节腔注射木瓜蛋白酶法建立KOA模型，摇法组进行频率0.5 Hz，每次5 min，每日1次，摇法治疗，塞来昔布组药物水溶液灌胃，每周期7 d，治疗个3周期。透射电镜观察滑膜细胞超微结构变化，蛋白质免疫印迹法（Western blot）检测αVβ3、FAK、ERK、C-Jun蛋白表达，实时荧光定量逆转录聚合酶链式反应（RT-qPCR）分析相关基因转录水平，组织块培养法分离原代滑膜成纤维细胞（FLSs），放射免疫法测定细胞培养上清基质金属蛋白酶-1（MMP-1）、基质金属蛋白酶-13（MMP-13）含量，染色法观察细胞骨架纤维状肌动蛋白（F-actin）重构。[结果] 摇法干预可显著改善细胞超微结构，显著下调αVβ3、FAK、ERK、C-Jun蛋白表达（P<0.01），相关基因转录水平同步降低（P<0.05），MMP-1、MMP-13分泌量减少（P<0.01），细胞骨架重构与炎症因子（MMP-1、MMP-13）分泌呈负相关（P<0.05）。[结论] 摇法通过调控αVβ3/FAK/ERK/C-Jun信号通路诱导滑膜细胞力学重构，抑制MMPs分泌机制改善膝骨关节炎滑膜炎症，为推拿治疗膝骨关节炎提供了分子水平的实验依据，证实了机械应力刺激在滑膜炎症调控中的重要作用。

Abstract:

[Objective] This study aimed to investigate the mechanism by which shaking manipulation alleviates synovial inflammation in knee osteoarthritis（KOA） by regulating the αVβ3/FAK/ERK/C-Jun signaling pathway. [Methods] A total of 100 male New Zealand white rabbits were randomly divided into five groups（Normal，Sham，Model，Shaking，and Celecoxib；n=20 per group）. The KOA model was induced by intra-articular injection of papain. The Shaking group received shaking manipulation treatment at a frequency of 0.5 Hz for 5 min once daily，and the Celecoxib group received intragastric administration of celecoxib solution，7 days per cycle for 3 cycles. Transmission electron microscopy was used to observe ultrastructural changes of synovial cells. Protein expression of αVβ3，FAK，ERK，and C-Jun was detected by Western blot，and mRNA expression was analyzed by RT-qPCR. Primary fibroblast-like synoviocytes（FLSs） were isolated using the tissue explant method. Levels of MMP-1 and MMP-13 in the culture supernatant were determined by radioimmunoassay，and F-actin cytoskeletal remodeling was observed by fluorescence staining. [Results] Shaking manipulation significantly improved synovial cell ultrastructure，downregulated the protein expression of αVβ3，FAK，ERK，and C-Jun（P<0.01），and reduced their mRNA levels（P<0.05）. The secretion of MMP-1 and MMP-13 was significantly decreased（P<0.01）. F-actin cytoskeletal remodeling was negatively correlated with inflammatory mediator secretion（P<0.05）. [Conclusion] Shaking manipulation improves KOA synovitis by regulating the αVβ3/FAK/ERK/C-Jun signaling pathway，inducing cytoskeletal remodeling in synovial cells，and suppressing MMP secretion. This study provides molecular-level evidence for the efficacy of manipulation therapy in KOA and highlights the role of mechanical stress in synovial inflammation regulation.