猪去氧胆酸对脂肪变性肝细胞活性的影响及其机制  

Effect of hyodeoxycholic acid on the activity of steatosis hepatocytes and its mechanism

作　　者：汪远远 邹艳[2] 刘朝霞[1,3] 阳学风 WANG Yuanyuan;ZOU Yan;LIU Zhaoxia;YANG Xuefeng(Department of Gastroenterology,Nanhua Hospital Affiliated to University of South China,Hengyang,Hunan 421002,China;Department of Hand and Foot Surgery,Nanhua Hospital Affiliated to University of South China,Hengyang,Hunan 421002,China;Hunan Provincial Clinical Research Center for Metabolic Associated Fatty Liver Disease,Hengyang,Hunan 421002,China)

机构地区：[1]南华大学附属南华医院消化内科,湖南衡阳421002 [2]南华大学附属南华医院手足外科,湖南衡阳421002 [3]湖南省代谢相关脂肪性肝病临床研究中心,湖南衡阳421002

出　　处：《临床肝胆病杂志》2024年第2期292-297,共6页Journal of Clinical Hepatology

基　　金：湖南省教育厅一般项目(20C1586);湖南省科技创新计划项目(2020SK51910,2021SK51902)。

摘　　要：目的探讨猪去氧胆酸(HDCA)在代谢相关脂肪性肝病(MAFLD)发展中的作用及机制,为进一步阐明MAFLD的发病机制提供新的理论依据。方法L02肝细胞作为实验细胞,利用棕榈酸诱导L02细胞发生脂肪变性。采用FXR siRNA干扰链技术,构建FXR低表达的肝细胞株。CCK8实验检测HDCA在不同浓度(0、100、200、300、400μmol/L)和时间(12、24、36、48 h)对L02脂肪变性肝细胞的影响。通过qRT-PCR检测法尼醇X受体(FXR)、增殖细胞核抗原(PCNA)、周期蛋白D1(Cyclin D1)、磷脂酰肌醇-3-激酶(PI3K)和蛋白激酶B(AKT)mRNA表达;Western Blot检测FXR、Cyclin D1、PCNA、PI3K、pPI3K、AKT和p-AKT蛋白表达。计量资料服从正态分布且方差齐时多组间比较采用单因素方差分析,进一步两两比较采用Tukey HSD检验;服从正态分布但方差不齐时采用Welch方差分析,进一步两两比较采用Games-Howell检验。两组间比较采用成组t检验。结果CCK8检测结果显示,300μmol/L HDCA处理的L02细胞和脂肪变性肝细胞活性明显下降(P值均<0.05);qRT-PCR检测结果显示,FXR mRNA表达增强,PCNA、Cyclin D1、PI3K、AKT的mRNA表达下降,差异均有统计学意义(P值均<0.05)。Western Blot检测结果显示,FXR蛋白表达明显上升(P<0.05);干扰L02细胞FXR的表达后,PCNA、PI3K、p-PI3K、AKT和p-AKT的蛋白表达均明显增加(P值均<0.05)。结论HDCA通过上调FXR表达抑制PI3K/AKT信号通路,从而造成脂肪变性肝细胞活性下降。Objective To investigate the role and mechanism of hyodeoxycholic acid(HDCA)in the progression of metabolic associated fatty liver disease(MAFLD),and to provide a new theoretical basis for further clarifying the pathogenesis of MAFLD.Methods L02 hepatocytes were used as experimental cells,and palmitic acid was used to induce steatosis in L02 cells.The farnesoid X receptor(FXR)siRNA interference chain technique was used to construct a hepatocyte cell line with low FXR expression.CCK8 assay was used to observe the effect of HDCA on L02 steatosis hepatocytes at different concentrations(0,100,200,300,and 400μmol/L)and time points(12,24,36,and 48 hours).The method of qRT-PCR was used to measure the mRNA expression levels of FXR,proliferating cell nuclear antigen(PCNA),Cyclin D1,phosphatidylinositol 3-kinase(PI3K),and protein kinase-B(AKT),and Western blot was used to measure the protein expression levels of FXR,Cyclin D1,PCNA,PI3K,phosphorylated PI3K(p-PI3K),AKT,and phosphorylated(p-AKT).A one-way analysis of variance was used for comparison of normally distributed continuous data with homogeneity of variance between multiple groups,and the Tukey HSD test was used for further comparison between two groups;the Welch analysis of variance was used for comparison of normally distributed continuous data with heterogeneity of variance between multiple groups,and the Games-Howell test was used for further comparison between two groups.The independent-samples t test was used for comparison between two groups.Results CCK8 assay showed a significant reduction in the viability of L02 cells and steatosis hepatocytes treated by 300μmol/L HDCA(P<0.05),and qRT-PCR showed a significant increase in the mRNA expression level of FXR and significant reductions in the mRNA expression levels of PCNA,Cyclin D1,PI3K,and AKT(all P<0.05).Western blot showed a significant increase in the protein expression level of FRX(P<0.05),and after interference of FXR expression in L02 cells,there were significant increases in the protein expression leve

关 键 词：代谢相关脂肪性肝病 猪去氧胆酸 法尼醇X受体 

分 类 号：R575.5[医药卫生—消化系统]