He Wen-Xi, Zhang Cheng-Liang, Xiang Dong, Yang Jin-Yu, Xu Yan-Jiao, Ren Xiu-Hua, Liu Dong
Department of Pharmacy of Tongji Hospital,Tongji Medical School,Huazhong Science and Technology University Wuhan 430030,China.
Zhongguo Zhong Yao Za Zhi. 2019 Sep;44(17):3780-3785. doi: 10.19540/j.cnki.cjcmm.20190416.402.
The aim of this paper was to investigate the molecular mechanism of Calculus Bovis Sativus( CBS) in alleviating lipid accumulation in vitro by serum pharmacology. The CBS-containing serum of mice was obtained by serum pharmacology method to evaluate its effect on the proliferation of LO2 hepatocytes. The lipid reducing effects of CBS-containing serum through Nrf2 was evaluated by fructose-induced LO2 hepatocyte steatosis model,nuclear factor erythroid 2 related factor 2( Nrf2) agonist oltipraz combined intervention,cell oil red O staining and intracellular triglyceride( TG) content. The effects of CBS-containing serum on lipid peroxidation and hepatocytes apoptosis were evaluated by reactive oxygen species( ROS) and apoptosis assay,respectively. Real-time quantitative polymerase chain reaction( PCR) was used to detect the relative expression of lipid synthesis-related genes and apoptosis-related genes.RESULTS:: showed that CBS drug-containing serum had no significant effect on LO2 hepatocyte proliferation. As compared with the model group,CBS-containing serum could effectively reduce the formation of lipid droplets in fructose-induced LO2 hepatocytes,significantly reduce intracellular TG and ROS levels,and significantly reduce hepatocyte apoptosis rate( P < 0. 05). As compared with the model group,carbohydrate responsive element binding protein( ChREBP),sterol regulatory element binding protein-1 c( SREBP-1 c),fatty acid synthase( FAS),acetyl-CoA carboxylase 1( ACC1),stearoyl-CoA desaturase 1( SCD1),Bax and caspase-3 mRNA levels were significantly reduced in CBS drug-containing serum treatment group( P<0. 05). All of the above effects could be reversed by oltipraz.In conclusion,CBS-containing serum can significantly inhibit the fructose-induced LO2 liver fat deposition,and the mechanism may be related to reducing intracellular ROS level through the Nrf2 pathway and improving intracellular peroxidation state to reduce apoptosis.
本文旨在通过血清药理学方法研究牛黄（CBS）体外减轻脂质蓄积的分子机制。采用血清药理学方法制备含CBS的小鼠血清，评价其对LO2肝细胞增殖的影响。通过果糖诱导的LO2肝细胞脂肪变性模型、核因子红细胞2相关因子2（Nrf2）激动剂奥替普拉联合干预、细胞油红O染色及细胞内甘油三酯（TG）含量，评价含CBS血清通过Nrf2发挥的降脂作用。分别通过活性氧（ROS）检测和凋亡检测，评价含CBS血清对脂质过氧化和肝细胞凋亡的影响。采用实时定量聚合酶链反应（PCR）检测脂质合成相关基因和凋亡相关基因的相对表达。结果显示，含CBS药物血清对LO2肝细胞增殖无显著影响。与模型组相比，含CBS血清可有效减少果糖诱导的LO2肝细胞中脂滴的形成，显著降低细胞内TG和ROS水平，并显著降低肝细胞凋亡率（P<0.05）。与模型组相比，含CBS药物血清治疗组中碳水化合物反应元件结合蛋白（ChREBP）、固醇调节元件结合蛋白-1c（SREBP-1c）、脂肪酸合酶（FAS）、乙酰辅酶A羧化酶1（ACC1）、硬脂酰辅酶A去饱和酶1（SCD1）、Bax和caspase-3的mRNA水平显著降低（P<0.05）。上述所有作用均可被奥替普拉逆转。综上所述，含CBS血清可显著抑制果糖诱导的LO2肝脏脂肪沉积，其机制可能与通过Nrf2途径降低细胞内ROS水平、改善细胞内过氧化状态以减少凋亡有关。
