中国老年学杂志

目的 基于Kelch样ECH相关蛋白(Keap)1/核因子红细胞2相关因子(Nrf2)/抗氧化反应元件(ARE)信号通路探讨温针灸对膝骨关节炎(KOA)大鼠软骨组织氧化损伤和炎性反应的影响。方法 将雄性Sprague-Dawley大鼠随机分为假手术组、KOA组、温针灸治疗组、Nrf2抑制剂组、治疗+Nrf2抑制剂组，每组12只。测定各组膝关节厚度及被动活动度；苏木素-伊红(HE)染色观察膝关节组织病理学变化并进行Mankin评分；酶联免疫吸附试验(ELISA)检测各组血清超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-Px)活性及丙二醛(MDA)、白细胞介素(IL)-1β、肿瘤坏死因子(TNF)-α、IL-6水平；Western印迹检测Keap1、Nrf2、血红素加氧酶(HO)-1蛋白表达。结果 与假手术组相比，KOA组膝关节厚度显著增加、被动活动度显著降低，软骨层染色较轻较薄、关节软骨受损、软骨细胞减少、滑膜异常，Mankin评分、SOD、GSH-Px、Nrf2、HO-1水平显著降低，MDA、IL-1β、TNF-α、IL-6、Keap1表达显著升高(P<0.05)。与KOA组比较，温针灸治疗组软骨表面粗糙，出现部分缺损和骨折，膝关节被动活动度、Mankin评分、SOD、GSH-Px、Nrf2、HO-1水平显著增加，膝关节厚度、MDA、IL-1β、TNF-α和IL-6、Keap1表达显著降低(P<0.05);Nrf2抑制剂组膝关节厚度显著增加，膝关节被动活动度显著降低、关节软骨受损严重，Mankin评分、SOD、GSH-Px、Nrf2、HO-1水平显著降低，MDA、IL-1β、TNF-α、IL-6、Keap1表达显著升高(P<0.05)。与温针灸治疗组相比，治疗+Nrf2抑制剂组软骨表面粗糙，缺损骨折，膝关节被动活动度、Mankin评分、SOD、GSH-Px、Nrf2、HO-1水平显著降低，膝关节厚度、MDA、IL-1β、TNF-α和IL-6、Keap1表达显著升高(P<0.05)。结论 温针灸能够有效改善KOA大鼠关节软骨的病理形态，减轻氧化损伤和炎性反应，可能与激活Keap1/Nrf2/ARE信号通路有关。

Objective To investigate the effects of warm acupuncture on oxidative damage and inflammatory response in cartilage tissue of rats with knee osteoarthritis(KOA) based on the Kelch-like ECH-associated protein(Keap)1/nuclear factor erythroid 2-related factor(Nrf)2/antioxidant response element(ARE) signaling pathway.Methods Male Sprague-Dawley rats were randomly divided into the sham operation group, KOA group, warm acupuncture treatment group, Nrf2 inhibitor group, and treatment+Nrf2 inhibitor group, with 12 rats in each group.The knee joint thickness and passive range of motion in each group were measured.Hematoxylin-eosin(HE) staining was performed to observe the histopathological changes of knee joint tissues, and Mankin scoring was conducted for evaluation.Enzyme-linked immunosorbent assay(ELISA) was used to detect the activities of superoxide dismutase(SOD) and glutathione peroxidase(GSH-Px),as well as the levels of malondialdehyde(MDA),interleukin(IL)-1β,tumor necrosis factor(TNF)-α and IL-6 in serum of each group.Western blot was used to determine the protein expressions of Keap1,Nrf2 and heme oxygenase(HO)-1.Results Compared with the sham operation group, knee joint thickness was significantly increased, passive range of motion was significantly decreased, the staining of the cartilage layer was relatively light and thin, the articular cartilage was damaged, the chondrocytes were reduced, and the synovial membrane was abnormal, the Mankin score and the levels of SOD,GSH-Px, Nrf2,and HO-1 were significantly decreased, the expressions of MDA,IL-1β,TNF-α,IL-6 and Keap1 were significantly increased in the KOA group(P<0.05).Compared with the KOA group, the cartilage surface was rough with partial defects and fractures, the passive range of motion of the knee joint, the Mankin score and the levels of SOD,GSH-Px, Nrf2,and HO-1 were significantly increased, the knee joint thickness and the expressions of MDA,IL-1β,TNF-α,IL-6 and Keap1 were significantly decreased in the warm acupuncture treatment group(P<0.05);in the Nrf2 inhibitor group, the knee joint thickness was significantly increased, the passive range of motion of the knee joint was significantly decreased, the articular cartilage was severely damaged, the Mankin score and the levels of SOD,GSH-Px, Nrf2,and HO-1 were significantly decreased, and the expressions of MDA,IL-1β,TNF-α,IL-6 and Keap1 were significantly increased(P<0.05).Compared with the warm acupuncture treatment group, the cartilage surface was rough with defects and fractures, the passive range of motion of the knee joint, the Mankin score and the levels of SOD,GSH-Px, Nrf2,and HO-1 were significantly decreased, the knee joint thickness and the expressions of MDA,IL-1β,TNF-α,IL-6,and Keap1 were significantly increased in the treatment+Nrf2 inhibitor group(P<0.05).Conclusions Warm acupuncture could effectively improve the pathological morphology of articular cartilage in KOA rats, reduce oxidative damage and inflammatory response, which may be related to the activation of the Keap1/Nrf2/ARE signaling pathway.