目的 探讨胰岛素样生长因子结合蛋白(insulin-like growth factor binding protein,IGFBP)7在非酒精性病脂肪性肝病(non-alcoholic fatty liver disease,NAFLD)发病中的作用.方法 通过喂食高脂饮食(high-fat diet,HFD)建立NAFLD小鼠模型,并通过向肝脏注射腺相关病毒(adeno-associated virus,AAV)介导的短发夹(sh)-IGFBP7敲除IGFBP7.C57BL/6小鼠随机分为4组(每组n=6):正常饮食组(ND)、高脂饮食组(HFD)、HFD空载体注射液组(HFD/AAV-null)、HFD AAV-sh-IGFBP7注射液组(HFD/AAV-sh-IGFBP7).ND组给与正常饲料12周,HFD组、HFD/AAV-null组、HFD/AAV-sh-IGFBP7组给与高脂饲料4周,之后HFD/AAV-sh-IGFBP7组、HFD/AAV-null组小鼠分别注射2 × 1011 AAV-sh-IGFBP7或AAV-null的基因组拷贝,继续喂食8周.测定各组小鼠体重、肝湿重、肝指数,通过生化分析仪检测各组小鼠血清甘油三酯(TG)、总胆固醇(TC)、丙氨酸转氨酶(ALT)、天冬氨酸转氨酶(AST)、游离脂肪酸(FFA)、甘油及肝脏TG.采用实时定量PCR、Western blot检测IGFBP7在各组中mRNA,蛋白表达水平及脂肪生成相关的固醇调节元件结合蛋白(sterol regulatory element binding protein,SREBP)1c、脂肪酸合酶(fatty acid synthase,FASN)、乙醜辅酶 a 羧化酶(acetyl-coa carboxylase,ACC)1 和硬脂醜辅酶 a 去饱和酶(stearoyl coA desaturase,SCD)1的mRNA表达水平.结果 与ND组相比,HFD组体重、肝湿重、肝指数及血清中ALT,AST,FFA,TG,TC,甘油、肝TG水平水平显著增加,差异均具有统计学意义(P＜0.05),而HFD/AAV-sh-IGFBP7组体重、肝湿重、肝指数及血清中ALT,AST,FFA,TG,TC,甘油、肝TG水平增加均少于HFD/AAV-null组的小鼠,差异均具有统计学意义(P＜0.05).与ND组对比,HFD组小鼠肝脏中IGFBP7mRNA及蛋白的表达水平升高,差异具有统计学意义(P＜0.01).与 HFD/AAV-null组相比,HFD/AAV-sh-IGFBP7组小鼠肝脏中IGFBP7蛋白表达水平显著下降,差异具有统计学意义(P＜0.01).与ND组相比,HFD组小鼠肝脏组织中SREBP-1c,FASN,ACC1和SCD1的mRNA表达水平均明显升高,差异均具有统计学意义(P＜0.01).而与HFD/AAV-null组相比,HFD/AAV-sh-IGFBP7组SREBP-1c,FASN,ACC1和SCD1的mRNA表达水平均降低,差异具有统计学意义(P＜0.01).结论 IGFBP7在NAFLD的发展过程中参与了肝脏脂肪变性,下调IGFBP7可减轻肝脏脂质积累,对NAFLD的发病机制具有保护作用,这可能是一种新的治疗NAFLD的药物.
Effect of silencing IGFBP7 expression on nonalcoholic fatty liver disease
Objective This study aims to explore the role of Insulin-like growth factor binding protein(IGFBP)7in the pathogenesis of nonalcoholic fatty liver disease(NAFLD).Methods A mouse model of NAFLD was established by feeding a high-fat diet and IGFBP7 was knocked out by injecting adeno-associated virus(AAV)-mediated short hairpin(sh)-IGFBP7 into the liver.The mice were randomly divided into 4 groups(n=6 per group):normal diet group(ND),high fat diet group(HFD),HFD empty carrier injection group(HFD/AAV-null),and HFD AAV-sh-IGFBP7 injection group(HFD/AAV-sh-IGFBP7).ND group was given normal diet for 12 weeks,HFD group,HFD/AAV-null group and HFD/AAV-sh-IGFBP7 group were given high fat diet for 4 weeks.HFD/AAV-sh-IGFBP7 group and HFD/AAV-null group were injected with 2 × 1011 genome copies of AAV-sh-IGFBP7 or AAV-null group,respectively,and then continued to be fed for 8 weeks.Body weight,liver wet weight and liver index of mice in each group were determined.Serum triglyceride(TG),total cholesterol(TC),alanine aminotransferase(ALT),aspartate aminotransferase(AST),free fatty acid(FFA),glycerol and liver TG of mice in each group were detected by biochemical analyzer.Real-time quantitative PCR and Western blot were used to detect IGFBP7 mRNA in each group.Protein expression levels and mRNA expression levels of sterol regulatory element binding protein 1c(SREBP-1c),fatty acid synthase(FASN),acetyl-CoA carboxylase1(ACC1)and stearoyl-CoA desaturase 1(SCD1)related to lipogenesis.Results Compared with ND group,body weight,liver wet weight,liver index and serum ALT,AST,FFA,TG,TC,glycerol and liver TG levels in HFD group were significantly increased,and the differences were statistically significant(P＜0.05),while body weight,liver wet weight,liver index and serum ALT,AST,and serum TG levels in HFD/AAV-sh-IGFBP7 group were significantly increased(P＜0.05).FFA,TG,TC,glycerol and liver TG levels increased less than those of HFD/AAV-null group,and the differences were statistically significant(P＜0.05).Compared with ND group,the expression levels of IGFBP7mRNA and protein in liver of mice in HFD group were increased,and the difference was statistically significant(P＜0.01).Compared with HFD/AAV-null group,the expression level of IGFBP7 protein in liver of HFD/AAV-sh-IGFBP7 group was significantly decreased,and the difference was statistically significant(P＜0.01).Compared with ND group,mRNA expression levels of SREBP-1c,FASN,ACC1 and SCD1 in liver tissues of mice in HFD group were significantly increased,with statistical significance(P＜0.01).Compared with HFD/AAV-null group,mRNA expression levels of SREBP-1c,FASN,ACC1 and SCD1 in HFD/AAV-sh-IGFBP7 group were decreased,and the difference was statistically significant(P＜0.01).Conclusion IGFBP7 is involved in hepatic steatosis during the development of NAFLD.Down-regulation of IGFBP7 can reduce the accumulation of lipid in the liver and has a protective effect on the pathogenesis of NAFLD,which may be a new drug for the treatment of NAFLD.
NETL
NSTL
万方数据
维普
