雷公藤红素改善糖尿病小鼠非酒精性脂肪性肝病的初步机制研究  

Preliminary mechanism of celastrol in improving non-alcoholic fatty liver disease in diabetic mice

作　　者：季欣 王雅静 徐思佳 居梦娴 冯颖杰 方彭华 张真稳 Ji Xin;Wang Yajing;Xu Sijia;Ju Mengxian;Feng Yingjie;Fang Penghua;Zhang Zhenwen(Department of Endocrinology,Northern Jiangsu People's Hospital Affiliated to Yangzhou University,Yangzhou 225001,China;Experimental Research Center of Clinical Medicine,First Medical School of Nanjing University of Chinese Medicine,Nanjing 210023,China)

机构地区：[1]扬州大学附属苏北人民医院内分泌科,扬州225001 [2]南京中医药大学第一临床医学院临床医学实验研究中心,南京210023

出　　处：《中华糖尿病杂志》2024年第12期1402-1410,共9页CHINESE JOURNAL OF DIABETES MELLITUS

基　　金：江苏省中医药科技发展计划项目(YB2020087);国家自然科学基金(82374100)。

摘　　要：目的探讨雷公藤红素对糖尿病小鼠非酒精性脂肪性肝病(NAFLD)的影响及初步作用机制。方法(1)选取10只db/db雄性小鼠,采用随机数字表将其分为模型(db/db)组和雷公藤红素(Cel)组,每组5只,另选5只同周龄雄性db/m小鼠作为空白(db/m)组。Cel组连续腹腔注射雷公藤红素21 d,db/m组、db/db组腹腔注射等体积生理盐水。根据实验方案记录体重、摄食量,完善葡萄糖耐量实验及胰岛素耐量实验。采用酶联免疫吸附法测定小鼠血浆中白细胞介素-1β(IL-1β)、白细胞介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)、空腹胰岛素、甘油三酯(TG)、总胆固醇(TC)水平。计算稳态模型评估胰岛素抵抗指数(HOMA-IR)。采用实时聚合酶链反应(PCR)、Western blotting检测肝脏中沉默信息调节因子1(SIRT1)、过氧化物酶体增殖物激活受体γ共激活因子-1α(PGC-1α)、肉碱棕榈酰转移酶1A(CPT1A)mRNA和蛋白水平。(2)棕榈酸(PA)刺激人肝癌细胞系HepG2细胞建立肝脂肪变性模型,将脂肪变性肝细胞分为模型(PA)组、雷公藤红素(Cel)组,正常肝细胞作为空白(Con)组。通过CCK8实验明确雷公藤红素干预细胞的最佳剂量,然后以1μmol/L雷公藤红素干预脂肪变性肝细胞模型24 h。干预结束后观察肝细胞中TG水平变化,利用实时PCR、Western blotting检测肝细胞中SIRT1、PGC-1α、CPT1A mRNA和蛋白水平。组间比较采用t检验或单因素方差分析。结果(1)与db/m组比较,db/db组小鼠体重、摄食量、脂肪含量、空腹血糖、HOMA-IR明显升高(P<0.05)。与db/db组比较,Cel组小鼠体重、摄食量、脂肪含量、空腹血糖、HOMA-IR显著下降(P<0.05)。(2)与db/m组比较,db/db组小鼠血浆及肝脏中TG、TC含量明显上升(P<0.05),血浆IL-1β、IL-6、TNF-α水平明显上升(P<0.05)。与db/db组比较,Cel组小鼠血浆及肝脏中TG、TC含量显著下降(P<0.05),血浆IL-1β、IL-6、TNF-α水平显著下降(P<0.05)。(3)与db/m组比较,db/db组小鼠�Objective To investigate the effect of celastrol on non-alcoholic fatty liver disease(NAFLD)in diabetic mice and its preliminary mechanism.Methods(1)A total of 10 male db/db mice were divided into two groups:model(db/db)group and celastrol(Cel)group by using a random number table,with 5 mice in each group.Five male db/m mice of the same age were used as the blank(db/m)group.Celastrol was injected intraperitoneally into Cel group for 21 days,and an equal volume of saline was injected intraperitoneally into the db/m and db/db groups.Body weight and food intake were recorded,and glucose and insulin tolerance tests were performed.The levels of interleukin-1β(IL-1β),interleukin-6(IL-6),tumor necrosis factor-α(TNF-α),fasting insulin,triglycerides(TG),and total cholesterol(TC)in plasma were measured by enzyme-linked immunosorbent assay computational homeostasis modeling to assess insulin resistance(HOMA-IR)indices.Real-time polymerase chain reaction(PCR)and Western blotting were used to detect the mRNA and protein levels of sirtuin1(SIRT1),peroxisome proliferator-activated receptor γ coactivator-1α(PGC-1α)and carnitine palmityltransferase 1A(CPT1A)in the liver.(2)Human hepatoma cell line HepG2 cells were stimulated with palmitic acid to establish a model of hepatic steatosis.Steatosis hepatocytes were divided into model(PA)group and celastrol(Cel)groups,and normal hepatocytes were used as the blank(Con)group.The optimal dosage of celastrol for cell intervention was clarified by CCK8 assay.Then 1μmol/L celastrol was used for 24 hours.At the end of the culture,the level of TG in hepatocytes was detected,and real-time PCR and Western blotting were used to detect the mRNA and protein expression of SIRT1,PGC-1α,and CPT1A.Group comparisons were performed using t-test or one-way analysis of variance(ANOVA).Results(1)Compared with db/m group,the body weight,food intake,fat content,fasting blood glucose,and HOMA-IR were significantly increased in db/db group(P<0.05);compared with db/db group,the body weight,food intake,

关 键 词：非酒精性脂肪性肝病 雷公藤红素 脂肪酸氧化 胰岛素抵抗 

分 类 号：R285.5[医药卫生—中药学]