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目的探讨白藜芦醇(resveratrol,RSV)对脂肪变性肝细胞线粒体功能的影响,以及沉默信息调节因子3(sirtuin 3,Sirt3)在其中的重要作用。方法用0.2 mmol/L棕榈酸(palmitic acid,PA)处理Hep G2细胞24 h建立肝细胞脂肪变性模型,再用40μmol/L RSV和/或50μmol/L Sirt3抑制剂3-TYP处理24 h。实验分为对照组、PA组、PA+RSV组、PA+RSV+3-TYP组、PA+3-TYP组。用相关试剂盒检测甘油三酯(TG)含量、沉默调节蛋白3(sirtuin 3,Sirt3)活性、ATP合酶活性、ATP生成量以及线粒体基础呼吸率,使用DCFH-DA荧光探针检测细胞活性氧(reactive oxygen species,ROS)含量,Western blot检测Sirt3蛋白表达。Sirt3 siRNA处理Hep G2细胞后再用PA和/或RSV处理,分析ROS和甘油三酯含量。结果相对于PA组,PA+RSV组Hep G2细胞的甘油三酯、ROS含量明显降低(P<0.05),ATP合酶活性、ATP生成量和基础呼吸率明显提高(P<0.05)。而3-TYP或Sirt3 siRNA处理后,RSV的上述作用被削弱(P<0.05)。结论 RSV通过上调脂肪变性肝细胞Sirt3的表达与活性而改善其线粒体功能。
Objective To investigate the effects of resveratrol (RSV) on the mitochondrial function of steatosis hepatocytes and the role of sirtuin 3 (Sirt3) in the pathogenesis of steatohepatitis. Methods Hep G2 cells were treated with 0.2 mmol / L palmitic acid (PA) for 24 hours to establish a hepatocellular steatosis model and treated with 40 μmol / L RSV and / or 50 μmol / L Sirt3 inhibitor 3-TYP for 24 hours. The experiment was divided into control group, PA group, PA + RSV group, PA + RSV + 3-TYP group and PA + 3-TYP group. The triglyceride (TG) content, sirtuin 3 (Sirt3) activity, ATP synthase activity, ATP production and mitochondrial respiration rate were detected by using the kit. DCFH-DA fluorescent probe was used to detect the cellular reactive oxygen species (reactive oxygen species, ROS) content, Western blot detection of Sirt3 protein expression. Sirt3 siRNA treatment of Hep G2 cells followed by PA and / or RSV treatment, ROS and triglyceride content analysis. Results Compared with PA group, the content of triglyceride and ROS in Hep G2 cells in PA + RSV group was significantly lower than that in PA group (P <0.05). The ATP synthase activity, ATP production and basic respiration rate were significantly increased (P <0.05). The above effects of RSV were weakened after 3-TYP or Sirt3 siRNA treatment (P <0.05). Conclusion RSV improves mitochondrial function by up-regulating the expression and activity of Sirt3 in steatosis hepatocytes.