橙皮素调控Nrf2/ARE通路改善载脂蛋白E基因敲除小鼠动脉粥样硬化的实验研究  

Hesperedin Regulates Nrf2/ARE Pathway to Improve Atherosclerosis in Apolipoprotein E Knockout Mice

作　　者：来利红[1] 汪晓东 杨兰 杜来景[1] LAI Lihong;WANG Xiaodong;YANG Lan;DU Laijing(The First Affiliated Hospital of Henan University of Science and Technology,Luoyang 471000,Henan,China)

机构地区：[1]河南科技大学第一附属医院,河南洛阳471000

出　　处：《中西医结合心脑血管病杂志》2023年第5期821-827,共7页Chinese Journal of Integrative Medicine on Cardio-Cerebrovascular Disease

基　　金：2019年度河南省医学科技攻关计划联合共建项目(No.LHGJ20190561)。

摘　　要：目的:基于核因子E2相关因子2(Nrf2)/抗氧化反应原件(ARE)通路探讨橙皮素对载脂蛋白E基因敲除(ApoE^(-/-))小鼠动脉粥样硬化(AS)的影响。方法:将48只ApoE^(-/-)小鼠用高脂饮食喂养12周后,采用左颈总动脉缩窄性套管法建立ApoE^(-/-)小鼠AS模型。模型制备成功后将ApoE^(-/-)小鼠随机分为模型组、橙皮素低剂量组、橙皮素高剂量组、鸦胆子苦醇组,每组12只。另取12只雄性C57BL/6J小鼠作为对照组,给予普通饲料喂养。橙皮素低、高剂量组小鼠分别给予50 mg/kg、100 mg/kg橙皮素灌胃,每日1次;鸦胆子苦醇组小鼠灌胃100 mg/kg橙皮素,隔天腹腔注射Nrf2抑制剂鸦胆子苦醇2 mg/kg,连续给药12周;对照组和模型组给予等量0.5%羧甲基纤维素钠灌胃。检测各组血清血脂水平以及丙二醛(MDA)、超氧化物歧化酶(SOD)和谷胱甘肽过氧化物酶(GSH-Px)水平;油红O染色观察主动脉斑块形成和主动脉窦脂质沉积情况,并计算损伤面积百分比;荧光探针测定主动脉中活性氧(ROS)的生成;蛋白免疫印迹法(Western Blot)检测主动脉中Nrf2/ARE通路相关蛋白表达。结果:与对照组相比,模型组小鼠主动脉ROS增加,血清总胆固醇(TC)、三酰甘油(TG)、低密度脂蛋白胆固醇(LDL-C)、氧化型低密度脂蛋白(ox-LDL)、MDA、主动脉斑块损伤和主动脉窦脂质损伤面积百分比、Kelch样环氧氯丙烷相关蛋白1(Keap1)的表达均明显升高,血清SOD、GSH-Px活性、主动脉Nrf2、还原型辅酶/醌氧化还原酶(NQO1)、血红素加氧酶1(HO-1)的表达明显降低,差异均有统计学意义(P<0.05);与模型组相比,橙皮素低、高剂量组小鼠ROS降低,血清TC、TG、LDL-C、ox-LDL、MDA水平、主动脉斑块损伤和主动脉窦脂质损伤面积百分比、Keap1均明显下降,HDL-C、SOD、GSH-Px活性、Nrf2、NQO1、HO-1的表达均明显升高,差异均有统计学意义(P<0.05);鸦胆子苦醇可逆转高剂量橙皮素减轻ApoE^(-/-)小鼠AS病变的作用。Objective:To explore the effect of hesperetin on atherosclerosis in apolipoprotein E(ApoE)knockout mice based on nuclear factor erythroid 2 related factor 2(Nrf2)/antioxidant response factor(ARE)pathway.Methods:Forty-eight ApoE^(-/-)mice were fed with high-fat diet for 12 weeks,and then the atherosclerosis(AS)model of ApoE^(-/-)mice was established by left common carotid artery constriction cannula method.After successful model preparation,ApoE^(-/-)mice were randomly divided into model group,low-and high-dose hesperedin groups(50 mg/kg and 100 mg/kg),and Brucea Javanica alcohol group(100 mg/kg hesperedin+Nrf2 inhibitor Brucea Javanica alcohol 2 mg/kg),with 12 mice in each group.Another 12 male C57BL/6J mice were selected as control group and fed with normal diet.The mice in low-dose and high-dose hesperedin groups were given 50 mg/kg and 100 mg/kg hesperedin by gavage,once a day.While the mice in Brucea Javanica group were given 100 mg/kg hesperetin by gavage,and 2 mg/kg Brucea Javanica by intraperitoneal injection every other day,for 12 weeks.The mice in control group and model group were given 0.5%sodium carboxymethylcellulose by gavage.The levels of serum lipid,malondialdehyde(MDA),superoxide dismutase(SOD),and glutathione peroxidase(GSH-Px)were detected.The formation of aortic plaque and lipid deposition in aortic sinus were observed by oil red O staining,and the percentage of damage area was calculated.The production of reactive oxygen species(ROS)in aorta was measured by fluorescence probe,and the expression of Nrf2/ARE pathway related proteins in aorta was detected by Western Blot.Results:Compared with control group,ROS increased in the aorta of model group,serum total cholesterol(TC),triacylglycerol(TG),low density lipoprotein cholesterol(LDL-C),oxidized low density lipoprotein(ox-LDL),MDA,percentages of aortic plaque injury and aortic sinus lipid injury,and expression of Kelch-like ECH-associated protein 1(Keap1)were significantly increased(P<0.05),serum SOD and GSH-Px activities,aortic Nrf2,reductive c

关 键 词：动脉粥样硬化 橙皮素 氧化应激 核因子E2相关因子2 NRF2 抗氧化反应原件 ARE 载脂蛋白E 实验研究 

分 类 号：R285.5[医药卫生—中药学]