桃红四物汤调控Nrf2/Keap1/HO-1信号通路介导的铁死亡在膝骨关节炎中的作用机制

Mechanism of Taohong Siwu Decoction (桃红四物汤) in Regulating Ferroptosis Mediated by Nrf2/Keap1/HO-1 Signaling Pathway in Knee Osteoarthritis

目的:桃红四物汤调控核因子E2相关因子(Nrf2)/Kelch样ECH关联蛋白1(Keap1)/血红素氧合酶-1(HO-1)信号通路介导的铁死亡在膝骨关节炎(KOA)中的作用机制。方法:40只SD大鼠随机分为假手术对照组、模型对照组、桃红四物汤9 g/kg组、双醋瑞因胶囊0.0045 g/kg组,每组各10只;各组均采用内侧半月板失稳手术造模,造模成功1 w后各组灌胃给予相应的药物或无菌水,1次/d,连续8 w。干预完成后,检测各组大鼠血清白介素-1β(IL-1β)、肿瘤坏死因子-α(TNF-α)、IL-18含量及关节软骨组织丙二醛(MDA)含量、超氧化物歧化酶(SOD)活力、活性氧(ROS)含量;透射电镜观察大鼠软骨线粒体数量、形态、结构及分布情况;Western Blot法检测大鼠关节软骨组织Keap1、Nrf2、HO-1、谷胱甘肽过氧化物酶4(GPX4)、肿瘤抑制蛋白53(P53)、溶质载体家族7成员11重组蛋白(SLC7A11)的表达。结果:与假手术对照组比较,模型对照组Mankin和OARSI病理评分增加,IL-1β、TNF-α、IL-18、MDA、ROS含量显著升高,Keap1、P53蛋白表达显著上调,SOD活力显著降低,Nrf2、HO-1、GPX4、SLC7A11蛋白表达显著下调(P<0.01);与模型对照组比较,桃红四物汤组Mankin和OARSI病理评分降低,IL-1β、TNF-α、IL-18、MDA、ROS含量降低,Keap1、P53蛋白表达显著下调,SOD活力显著升高,Nrf2、HO-1、GPX4、SLC7A11蛋白表达显著上调(P<0.01)。结论:桃红四物汤可能通过激活Nrf2/Keap1/HO-1信号通路抑制大鼠关节软骨铁死亡。

Objective:To investigate the mechanism of Taohong Siwu Decoction(桃红四物汤) in regulating the Nrf2/Keap1/HO-1 signaling pathway-mediated ferroptosis in knee osteoarthritis(KOA). Methods:40 SD rats were randomly divided into a sham surgery group, a model control group, Taohong Siwu Decoction(9 g/kg) group, and a diacerein capsule(0.0045 g/kg) group, with 10 rats in each group. Rat models were established by using medial meniscus instability surgery. After successful modeling for one week, rats were given corresponding medication or the same dose of sterile water by gavage, once a day for eight weeks. After the intervention was completed, the levels of serum interleukin-1β(IL-1β),tumor necrosis factor-α(TNF-α),interleukin-18(IL-18),as well as the levels of malondialdehyde(MDA),superoxide dismutase(SOD),and reactive oxygen species(ROS) in articular cartilage tissue were detected. The number, morphology, structure, and distribution of cartilage mitochondria in each group were observed by transmission electron microscopy. Western blot was used to detect the expression of Keap1,Nrf2,HO-1,glutathione peroxidase 4(GPX4),tumor suppressor protein 53(P53),and solute carrier family 7 member 11 recombinant protein(SLC7A11) in rat articular cartilage. Results:Compared with the sham surgery group, the pathological scores of Mankin and OARSI in the model control group were increased, the levels of IL-1β,TNF-α,IL-18,MDA,and ROS were significantly increased, and the protein expressions of Keap1 and P53 were significantly up-regulated. The SOD activity was significantly decreased, and the protein expressions of Nrf2,HO-1,GPX4,and SLC7A11 were significantly down-regulated(P<0.01). Compared with the model control group, the pathological scores of Mankin and OARSI in the Taohong Siwu Decoction group were decreased, and the levels of IL-1β,TNF-α,IL-18,MDA,and ROS were decreased. The protein expressions of Keap1 and P53 were significantly down-regulated. The SOD activity was increased, and the protein expressions of Nrf2,HO-1,GPX4,and SLC7A11 were significantly up-regulated(P<0.01). Conclusion:Taohong Siwu Decoction may inhibit ferroptosis in rat articular cartilage by activating the Nrf2/Keap1/HO-1 signaling pathway.