参芪调肾方减轻慢阻肺肺肾气虚证大鼠气道炎症的机制:基于铁死亡途径

Shenqi Tiaoshen Formula alleviates airway inflammation in rats with chronic obstructive pulmonary disease and kidney qi deficiency syndrome by inhibiting ferroptosis via regulating the Nrf2/SLC7A11/GPX4 signaling pathway

目的基于核因子E2相关因子2(Nrf2)/溶质载体家族7成员11(SLC7A11)/谷胱甘肽过氧化物酶4(GPX4)途径探讨参芪调肾方(SQTSF)调控铁死亡减轻COPD肺肾气虚证病证结合大鼠气道炎症的作用。方法48只SD大鼠随机分为对照组(CON)、模型组(MOD)、SQTSF低、中、高剂量组(SQTSF-L、SQTSF-M、SQTSF-H)和氨茶碱组(APL),n=8。除对照组外,其余大鼠构建COPD肺肾气虚证模型,并从第30天开始灌胃SQTSF和APL。观察各组大鼠的体质量、抓力变化、肺功能、肺组织病理、肺泡灌洗液(BALF)炎性因子、肺组织氧化应激水平、铁离子代谢情况、肺组织细胞和线粒体超微结构以及Nrf2/SLC7A11/GPX4信号通路和铁死亡相关基因和蛋白的表达。结果与CON组比较,MOD组大鼠表现出皮毛枯黄脱落、对刺激反应迟钝,精神萎顿、嗜睡喜聚成堆闭目静卧等明显的气虚神疲表现;体质量和抓力明显下降(P<0.05);第0.1秒用力呼气量(FEV_(0.1))、用力肺活量(FVC)、FEV_(0.1)/FVC、Cdyn明显降低(P<0.05);肺组织表现出明显的炎性浸润和肺气肿,支气管、血管周围和肺泡炎症评分、肺组织平均内衬间隔(MLI)、肺泡破坏指数(DI)明显上升,平均肺泡数(MAN)明显降低(P<0.05);BALF中IL-1β、TNF-α、IL-6、IL-13明显上升(P<0.05);肺组织ROS、MDA明显上升,GSH明显下降(P<0.05);肺组织Fe^(2+)和总铁离子明显上升(P<0.05);透射电镜(TEM)观察发现MOD组大鼠肺组织细胞表现出线粒体外膜破裂,线粒体嵴消失等铁死亡特征性改变;肺组织Nrf2、GPX4、SLC7A11明显下降,酰基辅酶A合成酶长链家族成员4(ACSL4)明显上升(P<0.05);与MOD组比较,SQTSF能明显改善各项指标的病理改变,且作用效果优于APL组(P<0.05)。结论SQTSF能有效改善COPD肺肾气虚病证结合模型大鼠气道炎症和氧化应激,其机制可能与调控Nrf2/SLC7A11/GPX4信号通路抑制铁死亡有关。

Objective To investigate the effects of Shenqi Tiaoshen Formula(SQTSF)for alleviating airway inflammation in rats with both chronic obstructive pulmonary disease(COPD)and lung-kidney qi deficiency syndrome and explore its therapeutic mechanism.Methods Forty-eight SD rats were randomly divided into control group,model group,low-,medium-,and highdose SQTSF groups,and aminophylline(APL)group.In all but the control group,rat models of COPD with lung-kidney qi deficiency syndrome were established and treated with saline,SQTSF or APL via daily gavage as indicated(starting from day 30).The rats were observed for changes in body weight,grip strength,lung function,lung pathology,inflammatory cytokines in bronchoalveolar lavage fluid(BALF),oxidative stress levels,iron ion metabolism,cellular and mitochondrial ultrastructural changes in the lung tissue,and expressions of Nrf2/SLC7A11/GPX4 signaling pathway and ferroptosis-related proteins.Results The rats in the model group exhibited obvious symptoms of lung-kidney qi deficiency syndrome with significantly decreased body weight,grip strength,and lung function parameters.Examination of the lung tissue revealed showed significant inflammatory cell infiltration and emphysema with obvious bronchial,perivascular,and alveolar inflammation and alveolar destruction,significantly increased IL-1β,TNF-α,IL-6,and IL-13 levels in BALF,and elevated pulmonary oxidative stress levels and Fe^(2+)and total iron ion concentrations.The rat models also showed characteristic ultrastructural changes of ferroptosis in the lung tissue cells under transmission electron microscope and significantly decreased Nrf2,GPX4,and SLC7A11 and increased ACSL4 expressions in the lung tissue.Treatment with SQTSF significantly improved these pathological changes in the rat models with a better effect than APL.Conclusion SQTSF can effectively improve airway inflammation and oxidative stress in COPD rats with lung-kidney qi deficiency possibly by inhibiting ferroptosis via regulating the Nrf2/SLC7A11/GPX4 signaling pathway.