异甘草素对醋氨酚诱导的急性肝细胞损伤的保护作用

Cytoprotective effect and its mechanisms of isoliquiritigenin on acetaminophen induced acute injury of hepatocytes

目的：研究异甘草素（isoliquiritigenin，ISL）对醋氨酚（AAP）诱导的肝细胞损伤的保护作用及机制。方法：以AAP 20mmol／L培养原代肝细胞12h造成急性化学性肝损伤模型。肝细胞随机分为正常组，AAP损伤组，ISL预处理组和ISL加N-硝基-L-精氨酸甲酯（L-NAME）预处理组。酶学测定培养液ALT、AST活性，肝细胞CYP2E1活性和谷胱甘肽（GSH）含量，RT-PCR测定肝细胞CYP2E1 mRNA表达水平，放射免疫分析测定肝细胞cAMP和cGMP含量，三明治ELISA方法测定核转录调节因子（NF-κB等）活性。结果：ISL浓度（5～20μmol／L）依赖性抑制AAP引起的ALT和AST升高；下调肝细胞CYP2E1活性和mRNA表达，最大抑制率分别可达75．7％和78．7％，同时明显抑制AAP引起的肝细胞GSH含量下降；加入L-NAME 2．0mmol／L可阻断ISL 20μmol／L对肝细胞的保护作用；ISL不能逆转AAP引起的肝细胞cAMP含量下降，但可浓度依赖性提高cGMP含量（最高可达4、75倍）；ISL浓度依赖性抑制AAP所致NF-κB活性的提高，抑制率分别为24、8％、37．3％和64．1％。结论：ISL对CYP2E1介导的AAP肝细胞损伤具有保护作用，可能与通过NO-cGMP通路，抑制NF-κB激活和下调CYP2E1表达有关。

AIM： To study the cytoprotective action and mechanisms of isoliquiritigenin （ISL） on acute injury of hepatocytes induced by acetaminophen. METHODS： Rat hepatocytes in sandwich cultures were randomly divided into control group, model group, ISL group and ISL plus L-NAME groups. The acute injury of hepatocytes was induced by being exposed into acetaminophen （20 mmol/L） for 12 h. In the pretreatment groups, L-NAME and/or ISL were given before administration of acetaminophen. The activities of ALT and AST in medium were measured by using enzyme assays. The CYP2E1 expression in hepatocytes was determined by using enzyme assays and semiquantitative RT-PCR. The contents of cGMP and cAMP in hepatocytes were measured by radioimmunoassay. The activites of nuclear transcription regulation factors such as NF-κB were determined by sandwich ELISA. RESULTS： ISL （5- 20 μmol/L） reduced the increase of the ALT and AST levels dose-dependently. And the cytochrome P450 （CYP） 2El （aniline hydroxylase, ANH） activity was decreased by 75.7% maximumly and the mRNA expression was inhibited in a dose （50- 200 μg/mL） dependent manner, the maximum inhibitory rate was 78.7 %. In addition, ISL remarkably increased the glutathione content decreased by acetaminophen of hepatocytes. L-NAME （2 mmol/L） could reverse the protective effect of ISL （20 μmol/L）. ISL could not reverse the decrease of the level of cAMP induced by acetaminophen of hepatocytes but ISL could dose-dependently enhance the intracellular cGMP content （to 4.75 times maximumly） and suppress the NF-κB activation, the inhibitory rates were 24.8%,37.3% and 64.1%. CONCLUSION： ISL can protect the injury of hepatocytes induced by acetaminophen. It may be relate to the inhibition of NF-κB activation and down regulation of CYP2E1 expression via NO -cGMP pathways.