基于NCOA4介导的铁自噬探讨大补脾汤对放射性肺损伤大鼠氧化应激的影响

Effects of Dabupi Decoction on Oxidative Stress of Rats with Radiation-induced Lung Injury Based on NCOA4-mediated Ferritinophagy

目的 观察大补脾汤通过核受体辅激活因子4(NCOA4)介导的铁自噬对放射性肺损伤大鼠氧化应激的影响,探讨其可能作用机制。方法 30只SPF级SD大鼠随机分为正常对照组、模型组和中药高、中、低剂量组,每组6只。中药高、中、低剂量组分别灌胃24.28、12.14、6.07 g/kg大补脾汤,正常对照组和模型组灌胃等体积生理盐水,连续14 d。给药后除正常对照组外,其余大鼠右肺予X射线8 Gy单次照射。HE染色观察大鼠右肺组织病理变化,试剂盒检测右肺组织亚铁离子(Fe2+)、谷胱甘肽(GSH)、丙二醛(MDA)、超氧化物歧化酶(SOD)、活性氧(ROS)含量,免疫组化、Western blot和qPCR检测右肺组织核因子E2相关因子2(Nrf2)、NCOA4、铁蛋白重链1(FTH1)、谷胱甘肽过氧化物酶4(GPX4)蛋白和mRNA表达。结果 与正常对照组比较,模型组大鼠肺泡壁断裂,肺泡塌陷变形,肺间质充血、出血、炎性细胞浸润并伴有少量纤维化,右肺组织Fe2+、MDA、ROS含量及NCOA4蛋白和mRNA表达明显升高,GSH、SOD含量及Nrf2、FTH1、GPX4蛋白和mRNA表达明显降低,差异有统计学意义(P<0.01);与模型组比较,中药各剂量组大鼠右肺组织病理损伤明显减轻,右肺组织Fe2+、MDA、ROS含量及NCOA4蛋白和mRNA表达明显降低,GSH、SOD含量及Nrf2、FTH1、GPX4蛋白和mRNA表达明显升高,差异有统计学意义(P<0.05,P<0.01)。结论 大补脾汤可能通过NCOA4介导的铁自噬抑制铁死亡,减轻放射性肺损伤氧化应激反应,改善肺部病理损害,发挥对放射性肺损伤的防护作用。

Objective To investigate the effect of Dabupi Decoction on oxidative stress in rats with radiationinduced lung injury by regulating ferroptosis through nuclear receptor coactivator 4(NCOA4)-mediated ferritinophagy;To discuss its possible mechanism.Methods Totally 30 SPF SD rats were randomly divided into normal control group,model group,and TCM high-,medium-and low-dosage groups,with 6 rats in each group.The TCM high-,medium-and low-dosage group were given Dabupi Decoction 24.28,12.14 and 6.07 g/kg by gavage respectively,the normal control group and model group were given equal saline solution for gavage for 14 days.Except for the normal control group,the right lungs of other rats were irradiated with 8 Gy single X-ray after administration.HE staining was used to observe the pathological changes in right lung tissue;the kit was used to detect the contents of Fe2+, GSH, MDA, SOD and ROS in right lung tissue;immunohistochemistry, Western blot andqPCR were used to detect the expressions of nuclear factor E2 related factor 2 (Nrf2), NCOA4, ferritin heavy chain 1(FTH1), glutathione peroxidase 4 (GPX4) protein and mRNA in right lung tissue. Results Compared with the normalcontrol group, the alveolar wall of the model group was broken, the alveoli were collapsed and deformed, thepulmonary interstitium was congested, bleeding, inflammatory cell infiltration and a small amount of fibrosis, thecontents of Fe2+ , MDA, ROS and the expression of NCOA4 protein and mRNA in right lung tissue increasedsignificantly, the contents of GSH, SOD and the expressions of Nrf2, FTH1, GPX4 protein and mRNA decreasedsignificantly, with statistical significance (P<0.01). Compared with the model group, the pathological damage of rightlung tissue of rats in each dosage of TCM group significantly alleviated, the contents of Fe2+ , MDA, ROS and theexpression of NCOA4 protein and mRNA in right lung tissue decreased significantly, the contents of GSH, SOD andthe expressions of Nrf2, FTH1, GPX4 protein and mRNA significantly increased, with statistical significance (P<0.05, P<0.01). Conclusion Dapbupi Decoction may reduce oxidative stress response, improve lung pathologicaldamage and play a protective role against radiation-induced lung injury by inhibiting ferroptosis through NCOA4-mediated ferritinophagy.