解剖学报

解剖学报 ›› 2018, Vol. 49 ›› Issue (1): 49-55.doi: 10.16098/j.issn.0529-1356.2018.01.008

• 细胞和分子生物学 • 上一篇    下一篇

奥贝胆酸对非酒精性脂肪肝大鼠法尼醇X受体及细胞色素P450家族成员7A1基因表达的影响

林浩1 梁大伟1* 曾建峰2 李述剑3 王建福4 李飞1   

  1. 1.雅安职业技术学院药学与中药学教研室， 四川 雅安 625000; 2.四川省简阳市川空人民医院药剂科， 四川 简阳 641400; 3.四川省绵竹市土门镇卫生院内科， 四川 绵竹 618200; 4.雅安市河北医院药剂科 四川 雅安 625000
  • 收稿日期:2017-04-24 修回日期:2017-07-20 出版日期:2017-02-06 发布日期:2018-02-06
  • 通讯作者: 梁大伟 E-mail:qingyu152@sina.com

Effect of obeticholic acid on expression of farnesoid X receptor and cytochrome 7A1 in nonalcoholic fatty liver rats

LIN Hao1 LIANG Da-wei 1* ZNEG Jian-feng2 LI Shu-jian3 WANG Jian-fu4 LI Fei1   

  1. 1. Department of Pharmacy and Herbal Medicine, Ya’an Vocational and Technical College, Sichuan, Ya’an 625000, China; 2. Department of Pharmacy, Jianyang Chuan Kong People’s Hospital, Sichuan, Jianyang 641400, China;  3. Department of Internal Medicine, Tumen Town, Sichuan Mianzhu 618200, China; 4.Department of Pharmacy, Hebei Hospital,  Sichuan Ya’an 625000, China
  • Received:2017-04-24 Revised:2017-07-20 Online:2017-02-06 Published:2018-02-06
  • Contact: LIANG Da-wei E-mail:qingyu152@sina.com

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摘要：

目的 探讨奥贝胆酸对非酒精性脂肪肝（NAFLD）大鼠法尼醇X受体（FXR）蛋白、细胞色素P450家族成员7A1（CYP 7A1）蛋白及其基因表达的影响，及保肝作用。 方法 雄性SD大鼠52只通过高脂饲料喂养建立大鼠非酒精性脂肪肝模型；设置对照组11只，模型组、奥低组、奥中组和奥高组共41只，给药4周后，取大鼠相同部位肝脏，免疫组织化学法测定FXR蛋白表达、RT-PCR检测FXR mRNA，Western blotting法检测CYP 7A1蛋白、RT-PCR检测CYP 7A1 mRNA的表达水平；通过大鼠肝指数，外周血谷丙转氨酶（ALT）、谷草转氨酶（AST）和碱性磷酸酶（ALP）的浓度，及肝脏组织切片观察，评估肝功能。 结果 高脂饲料喂养10周后，大鼠具有典型的NAFLD特征，模型建立成功。模型组的FXR蛋白和FXR mRNA表达水平显著低于对照组（P＜0.05），奥低组和奥中组的FXR蛋白和FXR mRNA表达水平与模型组相比，差异无统计学意义（P＞0.05），奥高组的FXR蛋白和FXR mRNA表达水平显著高于模型组（P＜0.05）；模型组、奥低组和奥中组的吸光度值显著低于对照组（P＜0.05），奥高组显著升高，与对照组差异无统计学意义（P＞0.05）。模型组的CYP 7A1蛋白和CYP 7A1 mRNA表达水平显著高于对照组（P＜0.05），奥低组和奥中组的CYP 7A1蛋白和CYP 7A1 mRNA的表达水平与模型组相比，差异无统计学意义（P＞0.05），奥高组的CYP 7A1蛋白和CYP 7A1 mRNA表达水平显著低于模型组（P＜0.05）。与对照组相比，模型组的肝指数（%）显著增大（P＜0.05）；奥低组和奥中组的肝指数（%）与模型组相比，差异无统计学意义（P＞0.05），奥高组肝指数（%）显著低于模型组（P＜0.05）。与对照组相比，模型组的ALT、AST和ALP的浓度显著升高（P＜0.05）；奥贝胆酸组的ALT、AST和ALP浓度显著低于模型组（P＜0.05），且随着奥贝胆酸给药量的增加，ALT、AST和ALP的浓度逐渐降低；肝组织切片观察结果显示，奥贝胆酸呈剂量依赖性改善大鼠肝组织。 结论 奥贝胆酸高剂量能显著上调FXR蛋白及其基因的表达，并抑制CYP 7A1蛋白及其基因的表达，显著改善大鼠肝功能及肝组织病理状态。

关键词: 奥贝胆酸, 非酒精性脂肪肝, 法尼醇X受体蛋白, 细胞色素7A1蛋白, 免疫印迹法, 大鼠

Abstract:

Objective To investigate the effect of obeticholic acid on protein expression and gene expression of farnesolid X receptor (FXR), cytochrome 7A1 (CYP 7A1) in non-alcohol fatty liver disease (NAFLD) rat and hepatocyteprotective effect. Methods Fifty two SD rats of the non-alcoholic fatty liver model was established by feeding with high-fat diet. The control group, model group,the experimental group(low), the experimental group(middle), the experimental group(high) were set up. After four weeks of administration, the same part of the liver was taken out. The expression of FXR protein was detected by immunohistochemistry method , the mRNA expression of FXR was detected by RT-PCR, the protein CYP7A1 was detected by Western blotting and the mRNA expression of CYP7A1 was detected by RT-PCR. The liver function was assessed by the rat liver index of the concentrations of peripheral blood alanine aminotransferase (ALT), aspartate transaminase(AST) and alkaline phosphatase(ALP), and the observation of liver sections. Results After fed with high-fat diet for 10 weeks, the rats had typical characteristics of NAFLD and the model was established successfully. The expression of FXR protein and FXR mRNA in the model group were significantly lower than that in the control group (P<0.05). There was no significant difference in the expression of FXR protein and FXR mRNA between the experimental group(low) and the experimental group(middle) (P> 0.05). The expression of FXR protein and FXR mRNA in the experimental group(high) was significantly higher than that in model group (P<0.05). The optical density values of model group, the experimental group(low), and the experimental group(middle) were significantly lower than those of control group (P<0.05). The optical density values of the experimental group(high) was significantly higher, and there was no significant difference compared with those of the control group (P>0.05). The CYP7A1 protein and CYP7A1 mRNA expression levels in the model group were significantly higher than those in the control group (P<0.05). Compared with those in the model group, the expression levels of CYP7A1 protein and CYP7A1 mRNA in the experimental group(low) and the experimental group(middle) had no significant difference (P>0.05). The levels of CYP 7A1 protein and CYP 7A1 mRNA in the experimental group(high) were significantly lower than those in the model group (P<0.05). Compared with the control group, the liver index (%) of the model group increased significantly (P<0.05), while the liver index (%) of the experimental group(low) and the experimental group(middle) had no significant difference compared with the model group (P> 0.05). The liver index (%) of the experimental group(high) was significantly lower than that of the model group (P<0.05). Compared with the control group, the concentrations of ALT, AST and ALP in the model group were significantly increased (P<0.05). The levels of ALT, AST and ALP in the experimental group were significantly lower than those in model group (P<0.05), and the concentrations of ALT, AST and ALP gradually decreased with the increase of acyclobacic acid dosage. The liver tissue sections were observed. The result showed that obeticholic acid improved the histopathology of rats in a dose-dependent manner. Conclusion High doses of acyclobacic acid can significantly upregulate the expression of FXR protein and its gene, inhibit the expression of CYP 7A1 protein and its gene, and significantly improve the liver function and pathological status of rats.

Key words: Obeticholic acid, Non-alcoholic fatty liver, Farnesoid X receptor, Cytochrome 7A1 protein, Western blotting, Rat