miR-23a-3p通过调控SIRT1/FOXO1通路影响NAFLD小鼠肝脏脂质代谢的研究  

miR-23a-3p regulates hepatic lipogenesis via SIRT1/FOXO1 pathway in mice with NAFLD

作　　者：孙雪莲 杨佳楠 姜同连 朱福彬 于闪闪 李响 李洪志 SUN Xue-lian;YANG Jia-nan;JIANG Tong-lian;ZU Fu-bin;YU Shan-shan;LIXiang;LI Hong-zhi(School of Basic Medicine,Beihua University,Jilin 132000,China;Jilin Provincial Science and Technology Innovation Cen-ter of Kidney Disease Precision Medicine Based on Gene Sequencing,Jilin 132000,China)

机构地区：[1]北华大学基础医学院,吉林吉林132000 [2]吉林省肾脏病基因测序精准医疗创新中心,吉林吉林132000

出　　处：《中国病理生理杂志》2022年第12期2175-2182,共8页Chinese Journal of Pathophysiology

基　　金：国家自然科学基金资助项目(No.81970644;No.81770856);吉林省教育厅科学技术研究项目(No.JJKH20200058KJ);吉林省肾脏病基因精准医疗科技创新中心项目(No.YDZJ202202CXJD039);北华大学研究生创新计划项目(No.2022-015)。

摘　　要：目的:探究微小RNA-23a-3p(miR-23a-3p)对非酒精性脂肪性肝病(NAFLD)小鼠肝脏脂质代谢的影响及沉默信息调节因子1(SIRT1)/叉头框蛋白O1(FOXO1)通路在其中的作用。方法:qPCR检测miR-23a-3p在脂肪肝细胞模型及NAFLD小鼠模型中的表达情况。高脂饮食(HFD)诱导miR-23a-3p基因敲除(miR-23a KO)小鼠,设置野生型(WT)C57BL/6小鼠正常饮食(ND)组和HFD组及miR-23a KO小鼠ND组和HFD组,每组10只,HFD组小鼠均接受HFD 12周。对各组小鼠体重和肝脏指数进行统计;HE染色观察肝组织病理损伤变化;油红O染色检测肝脏组织脂质蓄积程度;生化分析检测血清中总胆固醇(TC)、甘油三脂(TG)、丙氨酸转氨酶(AST)、天冬氨酸转氨酶(ALT)、高密度脂蛋白胆固醇(HDL-C)和低密度脂蛋白胆固醇(LDL-C)水平;qPCR检测肝脏脂代谢相关基因Srebp1、Fas、Pparγ、Pparα、Acc1和Cpt1的mRNA水平;双萤光素酶报告基因实验分析miR-23a-3p和SIRT1基因的靶向关系;Western blot检测AMP活化蛋白激酶(AMPK)、SIRT1、过氧化物酶体增殖物激活受体α(PPARα)及FOXO1蛋白的表达。结果:与对照组比较,脂肪肝细胞模型、动物模型及db/db小鼠肝组织中miR-23a-3p表达显著升高。与HFD(WT)组比较,HFD(miR-23a KO)组体重和肝脏脏器系数显著降低;HE和油红O染色结果显示,HFD(miR-23a KO)组肝脏脂肪蓄积减轻;血清TC、TG、AST和LDL-C水平显著降低(P<0.05),HDL-C水平显著升高(P<0.05);脂代谢相关基因Srebp1、Fas、Pparγ和Acc1的mRNA水平显著降低(P<0.05),Cpt1的mRNA水平显著升高(P<0.05)。双萤光素酶报告基因实验证明miR-23a-3p可以靶定SIRT1基因。Western blot结果显示,与HFD(WT)组比较,HFD(miR-23aKO)组SIRT1和PPARα表达显著增加(P<0.05),p-AMPK/AMPK比值显著升高(P<0.05),AcFOXO1/FOXO1比值显著降低(P<0.01)。结论:miR-23a-3p的缺失能够减轻HFD诱导的NAFLD小鼠肝脏脂质蓄积,改善肝脏的生理功能。miR-23a-3p的作用可能是通过靶定SIRT1基因而使FOXO1�AIM:To investigate the effect of microRNA-23a-3p(miR-23a-3p)on lipid metabolism in non-alcoholic fatty liver disease(NAFLD)mice,and to explore the role of silent information regulator 1(SIRT1)/forkhead box O1(FOXO1)signaling pathway in this process.METHODS:qPCR was used to detect the expression of miR-23a-3p in animal and cellular models.The NAFLD mouse model was established by feeding with high-fat diet(HFD).Mice were divided into 4 groups:wild-type(WT)C57BL/6 mice fed with normal diet(ND)group,WT mice fed with HFD group,miR-23a-3p knockout(miR-23a KO)mice fed with ND group,and miR-23a KO mice fed with HFD group,with 10 mice in each group.Body weight was recorded weekly.The liver tissue samples were collected at the end of the 12th week,and the liver index was also calculated.Pathological changes and accumulation of lipids in liver tissues were observed by HE and oil O red staining,respectively.Serum triglyceride(TG),total cholesterol(TC),aspartate aminotransferase(AST),alanine aminotransferase(ALT),high-density lipoprotein cholesterol(HDL-C)and low-density lipoprotein cholesterol(LDL-C)were detected.The mRNA expression levels of lipid metabolism-associated genes,including Srebp1,Fas,Pparγ,Pparα,Acc1 and Cpt1,were evaluated by qPCR.The dual-luciferase reporter assay was utilized to validate the targeting relationship between miR-23a-3p and SIRT1.The expression levels of AMP-activated protein kinase(AMPK),SIRT1,peroxisome proliferator-activated receptorα(PPARα)and FOXO1 proteins were measured by Western blot.RESULTS:The expression of miR-23a-3p was increased in the liver tissue of both HFD-fed mice and db/db mice.The body weight and liver index of HFD(miR-23a KO)mice were significantly reduced compared with HFD(WT)mice.The hepatocytes in HFD(WT)group were significantly swollen and degenerated,with a large number of fat vacuoles and orangered lipid droplets in the cytoplasm,which were ameliorated in HFD(miR-23a KO)group.Compared with HFD(WT)group,the serum TC,TG,AST and LDL-C levels were significantly decrease

关 键 词：微小RNA-23a-3p 非酒精性脂肪性肝病 SIRT1/FOXO1信号通路 脂质代谢 

分 类 号：R575.5[医药卫生—消化系统] R363.21[医药卫生—内科学]