非诺贝特缓解非酒精性脂肪性肝病的体外机制研究

发布时间：2018-10-24 20:44

【摘要】：目的利用细胞模型研究非诺贝特在缓解非酒精性脂肪性肝病(NAFLD)中的作用及其分子机制。方法首先以0~0.9mmol/L梯度浓度的油酸处理正常肝细胞LO2 48h建立脂肪肝细胞模型;同时,以非诺贝特处理正常细胞或模型组细胞48h,然后用CCK-8试剂盒测定细胞活力,用油红O染色观察细胞内脂滴形成或抑制情况,利用试剂盒检测细胞中的三酰甘油(TG)以及上清液中谷丙转氨酶(ALT)、谷草转氨酶(AST)水平,蛋白质免疫印迹法检测细胞中蛋白激酶B(AKT)、p-AKT、FSAN及Nrf2蛋白水平,并使用2′,7′-二氯荧光黄双乙酸盐(DCFH-DA)检测细胞内的活性氧自由基(ROS)含量。结果油酸浓度超过0.5mmol/L后细胞活力显著降低,非诺贝特可抑制高浓度油酸对细胞的损伤。油酸处理后细胞中出现大量脂滴,细胞内TG含量以及上清液中的ALT、AST水平显著升高,给予非诺贝特处理后细胞中脂滴减少,TG、ALT和AST含量均显著降低。油酸处理后细胞中AKT、p-AKT、FSAN和Nrf2蛋白水平升高,给予非诺贝特可抑制上述蛋白水平的升高。同时,非诺贝特逆转了由油酸导致的细胞内ROS累积。结论非诺贝特可以抑制油酸引起的脂肪积累,并缓解脂肪累积导致的细胞损伤,其机制是通过下调AKT/FSAN信号通路,从而降低ROS水平,进而减少ROS导致的细胞损伤。
[Abstract]:Objective to study the role and molecular mechanism of fenofibrate in relieving (NAFLD) in patients with non alcoholic fatty liver disease. Methods Fatty liver cell model was established by treating normal hepatocytes with oleic acid at gradient concentration of 0~0.9mmol/L for 48h, and normal cells or model cells treated with fenofibrate for 48h, then the cell viability was measured by CCK-8 kit. Oil red O staining was used to observe the formation or inhibition of lipid droplets in cells. The levels of triglyceride (TG) and (ALT), (ALT), in supernatant were detected by Elisa kit. The protein levels of protein kinase B (AKT), p-AKTN FSAN and Nrf2 were detected by Western blot, and the content of reactive oxygen free radical (ROS) in cells was determined by using 2-chloride-dichlorofluorescein diacetate (DCFH-DA). Results when the concentration of oleic acid exceeded 0.5mmol/L, the cell viability was significantly decreased, and fenofibrate could inhibit the damage caused by high concentration of oleic acid. After oleic acid treatment, a large number of lipid droplets appeared in cells, and the content of TG and ALT,AST in supernatant increased significantly. After fenofibrate treatment, the lipid droplets in cells decreased, and the contents of TG,ALT and AST decreased significantly. The protein levels of AKT,p-AKT,FSAN and Nrf2 increased after oleic acid treatment, and fenofibrate inhibited the increase. At the same time, fenofibrate reversed the accumulation of ROS in cells induced by oleic acid. Conclusion fenofibrate can inhibit fat accumulation induced by oleic acid and alleviate cell damage induced by lipid accumulation. The mechanism of fenofibrate is to decrease the level of ROS by down-regulating the AKT/FSAN signaling pathway and then to reduce the cell damage induced by ROS.
【作者单位】：华中科技大学同济医学院药学院;华中科技大学同济医学院附属同济医院儿科;
【分类号】：R575.5

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