泽泻提取物改善游离脂肪酸诱导的HepG2脂肪变性细胞模型的脂质代谢及氧化应激异常  

Alisma orientale extract alleviates lipid metabolism and oxidative stress abnormalities in free fatty acid-induced HepG2 steatosis cell model

作　　者：谢燕 李贵平 段月阳[2,3] 李良 吕耀中 温建辉 曹亮[2,3] 李旭 杨昊[2,3,4] 肖伟 王振中[1,2,3] XIE Yan;LI Guiping;DUAN Yueyang;LI Liang;LYU Yaozhong;WEN Jianhui;CAO Liang;LI Xu;YANG Hao;XIAO Wei;WANG Zhenzhong(Nanjing University of Chinese Medicine,Nanjing 210023,China;State Key Laboratory on Technologies for Chinese Medicine Pharmaceutical Process Control and Intelligent Manufacture,Lianyungang 222001,China;Jiangsu Kanion Pharmaceutical Co.,Ltd.,Lianyungang 222001,China;Institute of Chinese Materia Medica,Shanghai University of Traditional Chinese Medicine,Shanghai 201203,China)

机构地区：[1]南京中医药大学,江苏南京210023 [2]中药制药过程控制与智能制造技术全国重点实验室,江苏连云港222001 [3]江苏康缘药业股份有限公司,江苏连云港222001 [4]上海中医药大学中药研究所,上海201203

出　　处：《中草药》2024年第6期2002-2012,共11页Chinese Traditional and Herbal Drugs

基　　金：江苏省工信厅工业和信息产业转型升级专项-多组分中药研究关键技术。

摘　　要：目的研究泽泻提取物(Alisma orientale extract,AE)及其3种主要单体成分对游离脂肪酸(free fatty acid,FFA)诱导的HepG2脂肪变性细胞模型的脂质代谢及氧化应激异常的改善作用及机制。方法利用FFA(油酸-棕榈酸2∶1)建立稳定的HepG2脂肪变性细胞模型,CCK-8法测定AE、泽泻醇A、泽泻醇A-23-乙酸酯及泽泻醇A-24-乙酸酯对HepG2细胞的安全剂量;在FFA诱导同时给予AE(25.00、12.50、6.25 mg/L)、泽泻醇A(50.0、25.0、12.5μmol/L)、泽泻醇A-23-乙酸酯(25.00、12.50、6.25μmol/L)、泽泻醇A-24-乙酸酯(50.0、25.0、12.5μmol/L)处理24 h,检测细胞内脂滴的形成情况,利用生化试剂盒测定各组细胞内三酰甘油(triglyceride,TG)、总胆固醇(total cholesterol,TC)水平,DCFH-DA检测药物对细胞内活性氧(reactive oxygen species,ROS)生成的影响,采用试剂盒检测细胞内氧化应激关键指标丙二醛(malondialdehyde,MDA)、总谷胱甘肽(glutathione,GSH)水平以及超氧化物歧化酶(superoxide dismutase,SOD)活性;采用Western blotting检测过氧化物酶体增殖物激活受体α(peroxisome proliferator-activated receptorα,PPARα)、PPAR共激活因子-1α(peroxisome proliferator-activated receptor gamma coactivator-1α,PGC-1α)、下游脂肪酸氧化和胆固醇代谢相关蛋白及核因子红细胞2相关因子2/血红素加氧酶-1(nuclear factor red blood cell 2 associated factor 2/hemeoxygenase-1,Nrf2/HO1)通路蛋白表达。结果与对照组比较,HepG2脂肪变性细胞模型细胞内TC、TG、ROS及MDA水平显著升高(P<0.001),SOD活性及GSH水平降低(P<0.01、0.001);同时,细胞中PPARα、PGC-1α、肉碱棕榈酰转移酶1A(carnitine palmitoyl transferase 1A,CPT1A)、酰基辅酶A氧化酶1(acyl coenzyme A oxidase 1,ACOX1)、Nrf2及HO-1蛋白表达水平显著降低(P<0.05、0.01、0.001)。与模型组比较,AE、泽泻醇A及泽泻醇A-23-乙酸酯给药组细胞内脂滴、TG和TC水平显著下降(P<0.05、0.01、0.001),AE、泽泻醇A及泽泻醇A-24-乙酸酯�Objective To explore the ameliorative effect and mechanisms of Alisma orientale extract(AE)and its three primary components on lipid metabolism abnormalities and oxidative stress in free fatty acids(FFA)-induced HepG2 steatosis cell model.Methods A HepG2 steatosis cell model was established using FFA[oleic acid-palmitic acid(2∶1)].Safe dosages of AE,alisol A,alisol A-23-acetate,and alisol A-24-acetate for HepG2 cells were determined using CCK-8 kit.Simultaneously administering AE(25.00,12.50,6.25 mg/L),alisol A(50.0,25.0,12.5μmol/L),alisol A-23-acetate(25.00,12.50,6.25μmol/L),and alisol A-24-acetate(50.0,25.0,12.5μmol/L)treatment for 24 h under FFA induction,the formation of intracellular lipid droplets was detected;Levels of triglyceride(TG)and total cholesterol(TC)were assessed using biochemical kits.DCFH-DA assay was utilized to measure intracellular reactive oxygen species(ROS)generation.Malondialdehyde(MDA),glutathione(GSH)levels and superoxide dismutase(SOD)activity as indicators of oxidative stress,were also determined.Western blotting was employed to detect the expressions of peroxisome proliferator-activated receptorα(PPARα),PPAR coactivator-1α(PGC-1α),and proteins involved in fatty acid oxidation,cholesterol metabolism,and nuclear factor erythrocyte 2-associated factor 2/heme oxygenase-1(Nrf2/HO-1)pathway.Results Compared with control group,HepG2 steatosis cell model exhibited significantly increased intracellular TC,TG,ROS production and MDA leve ls(P<0.001),alongside reduced SOD activity and GSH level(P<0.01,0.001).There was also a decrease in PPARα,PGC-1α,carnitine palmitoyl transferase 1A(CPT1A),acyl coenzyme A oxidase 1(ACOX1),Nrf2 and HO-1 protein expressions(P<0.05,0.01,0.001).Compared with model group,AE,alisol A and alisol A-23-acetate led to a significant reduction in intracellular lipid droplets,TG,and TC levels(P<0.05,0.01,0.001);AE,alisol A and alisol A-24-acetate decreased ROS and MDA levels(P<0.05,0.001),increased SOD activity and GSH level(P<0.05,0.01).Furthermore,the expressi

关 键 词：泽泻 泽泻醇A 泽泻醇A-23-乙酸酯 泽泻醇A-24-乙酸酯 游离脂肪酸 非酒精性脂肪肝 脂质代谢 氧化应激 过氧化物酶体增殖物激活受体Α 核因子红细胞2相关因子2/血红素加氧酶-1通路 

分 类 号：R285.5[医药卫生—中药学]