脱氧胆酸对HBV复制及肝癌细胞自噬影响的研究  

Study on the effects of deoxycholic acid on HBV replication andautophagy in hepatocellular carcinoma cells

作　　者：侯冰 高玉雪 杨鹏翔 关媛月 刘芃祥 杨静[1] 陈德喜[2] Hou Bing;Gao Yuxue;Yang Pengxiang;Guan Yuanyue;Liu Pengxiang;Yang Jing;Chen Dexi(School of Basic Medicine and Forensic Medicine,Baotou Medical College,Baotou 014040,Inner Mongolia,China;Beijing Institute of Hepatology,Beijing You An Hospital,Capital Medical University,Beijing 100069,China)

机构地区：[1]包头医学院基础医学与法医学院,内蒙古包头014040 [2]首都医科大学附属北京佑安医院北京肝病研究所,北京100069

出　　处：《右江民族医学院学报》2023年第4期545-551,共7页Journal of Youjiang Medical University for Nationalities

基　　金：国家自然科学基金项目(82073676);北京市自然科学基金-北京市教委联合资助项目(KZ202010025037);包头医学院研究生科研创新项目(bycx2021001)。

摘　　要：目的观察脱氧胆酸(deoxycholic acid,DCA)对乙型肝炎病毒(hepatitis B virus,HBV)复制的影响,及其对肝癌细胞自噬的影响程度,探讨其可能的作用机制。方法观察不同浓度DCA对HepG2及HepG2.215细胞活性的影响,进而测定不同浓度DCA对HepG2.215细胞HBV DNA的载量及HBsAg、HBeAg和HBcAg表达水平的影响,蛋白免疫印迹检测自噬水平;将HepG2.215细胞转染乙型肝炎病毒X(HBX)蛋白突变表达质粒后,观察HBX对DCA处理的HepG2及HepG2.215细胞的HBV DNA载量以及HBsAg、HBeAg和HBcAg表达水平的影响,蛋白免疫印迹检测细胞自噬水平。结果①CCK-8实验结果显示随着DCA浓度的升高,细胞活力均呈现梯度下降趋势(P<0.01);②HepG2.215细胞的HBV DNA载量以及HBsAg、HBeAg、HBcAg水平均随着DCA浓度的提高而上升(P<0.01);③HepG2与HepG2.215细胞的LC3-Ⅱ/LC3-Ⅰ比值升高(P<0.01),HepG2细胞的p62蛋白水平随着DCA浓度的升高而下降(P<0.01),HepG2.215细胞的p62蛋白水平随着DCA浓度的升高而上升(P<0.01);④未经DCA处理的对照组、经100μmol/L DCA处理的对照组、未经DCA处理的HBX突变表达质粒转染组、经100μmol/L DCA处理的HBX突变表达质粒转染组的HBV DNA载量、HBsAg、HBeAg、HBcAg水平、HBX蛋白水平以及LC3-Ⅱ/LC3-Ⅰ比值均呈现依次上升趋势(P<0.01),p62蛋白水平呈现依次下降趋势(P<0.01)。结论①DCA处理可以降低肝癌细胞的活力;②DCA可以促进HBV的复制;③DCA可以促进HepG2细胞的自噬,在HepG2.215细胞中促进不完全自噬;④DCA可以通过促进HBX蛋白的合成来加强HBV的复制,进而促进不完全自噬。Objective To observe the effects of deoxycholic acid(DCA)on the hepatitis B virus(HBV)replication and the extent of autophagy in hepatocellular carcinoma cells,and explore the possible underlying mechanisms.Methods The study assessed the effects of different DCA concentrations on the viability of HepG2 and HepG2.215 cells.Additionally,the impact of different DCA concentrations on the HBV DNA load and expression levels of HBsAg,HBeAg,and HBcAg were evaluated in HepG2.215 cells.Western Blot analysis was performed to assess the levels of autophagy.Moreover,after HepG2.215 cells were transfected with a plasmid of mutant expression in hepatitis B virus X(HBX)protein,the effects of HBX on HBV DNA load and expression levels of HBsAg,HBeAg,and HBcAg were observed in both HepG2 and HepG2.215 cells treated with DCA.Western Blot was used to test cellular autophagy levels.Results①The results of the CCK-8 experiment showed a gradient decrease in cell viability with increasing concentrations of DCA(P<0.01).②The HBV DNA load and the levels of HBsAg,HBeAg and HBcAg were elevated with increasing concentrations of DCA in HepG2.215 cells(P<0.01).③The ratio of LC3-Ⅱ/LC3-Ⅰwas increased in both HepG2 and HepG2.215 cells(P<0.01).The protein levels of p62 were decreased with increased DCA concentrations in HepG2 cells(P<0.01),while it was increased in HepG2.215 cells(P<0.01).④The HBV DNA load,HBsAg,HBeAg,HBcAg levels,HBX protein levels,and the ratio of LC3-Ⅱ/LC3-Ⅰshowed graded increase(an increasing trend in turn)(P<0.01),and the protein level of p62 showed a sequential decrease(P<0.01)in the control group without DCA treatment,the control group treated with 100μmol/L DCA,the group transfected with the HBX mutant plasmid without DCA treatment and the group transfected with the HBX mutant plasmid treated with 100μmol/L DCA.Conclusion①DCA treatment can reduce the viability of hepatocellular carcinoma cells;②DCA can promote HBV replication;③DCA can promote autophagy in HepG2 cells and facilitate incomplete autophagy

关 键 词：脱氧胆酸 乙型肝炎病毒 自噬 肝肿瘤 

分 类 号：R373.21[医药卫生—病原生物学]