五层龙提取物改善高果糖所致大鼠脂肪肝的重要分子靶点—肝脏固醇调节元件结合蛋白-1c

发布时间：2018-01-19 02:40

  本文关键词： 固醇调节元件结合蛋白-1c 五层龙 果糖 脂肪肝　出处：《重庆医科大学》2014年硕士论文　论文类型：学位论文

【摘要】：目的：五层龙提取物（SOR）为治疗肥胖和糖尿病的传统中草药，这两种疾病都与脂肪肝密切相关。已有研究证实SOR具有改善脂肪肝的作用，但是其机制尚不清楚。本文旨在研究SOR改善高果糖所致大鼠脂肪肝的分子机制。 方法：选取24只健康SD大鼠随机分为4组（n=6），饲养10周：(1)空白对照组:自由饮水；(2)果糖对照组:给予10%果糖水(w/v,每天配制)；(3)SOR低浓度药物组:果糖溶液+SOR提取物5mg/kg；(4)SOR高浓度药物组:果糖溶液+SOR提取物20mg/kg。每三天记一次体重，并根据体重变化调整用药组大鼠给药量。用药组起始果糖水浓度均为10%，每天记录摄食量与饮用果糖水量，并计算果糖摄入量，根据果糖对照组近三天果糖摄入量，调整果糖水浓度使用药组大鼠果糖摄入量与果糖对照组一致。于第10周末，夜间禁食14h后，称重，乙醚麻醉下，摘除眼球取血，分离血浆检测TC、TG、血糖、NEFA及胰岛素含量；处死大鼠，取肝脏称重，行肝脏H.E染色和油红O染色，，同时检测肝脏TG、TC含量；用TRIzol提取肝组织总RNA，逆转录获得cDNA，用Real-Time PCR方法检测肝脏脂质合成转录因子SREBP-1c、ChREBP及其下游相关基因FAS、ACC-1、SCD-1、LPK和脂质氧化相关基因PPAR-α、CPT-1α、ACO及PPAR-γ、CD36的mRNA水平；提取肝细胞核蛋白，用Western blot方法检测调节脂质合成的两个关键转录因子SREBP-1c和ChREBP的蛋白表达量。实验数据应用StatView统计软件进行分析，以均数±标准误（x±SEM）表示，t检验进行两组数据的差异比较。 结果： 1.生化指标：与空白对照组相比，果糖对照组的血浆TC、TG、血糖及胰岛素含量明显增加，而NEFA显著下降（P＜0.05）；SOR用药组相对于果糖对照组，血浆TC、TG、血糖、胰岛素及NEFA均无明显改变（P＞0.05）。 2．肝相关参数：各组大鼠肝重与肝TC含量无差异（P＞0.05）；但果糖对照组与空白对照组相比，肝重与体重比、肝TG含量明显升高（P＜0.05）；SOR（20mg/kg）显著下调果糖引起的肝TG含量的升高（P＜0.05）。 3.形态学显示：（1）H.E染色显示，果糖对照组肝细胞空泡化的程度较空白对照组明显增强，SOR（20mg/kg）明显改善肝细胞空泡化的程度。（2）ORO染色显示，与空白对照组比较，果糖对照组ORO染色阳性面积区域及阳性面积率明显增加（P＜0.05），SOR（20mg/kg）具有明显的改善作用（P＜0.05）。 4. Real-Time PCR结果：果糖对照组相对于空白对照组，显著上调肝脏SREBP-1c、FAS、ACC-1、SCD-1、ChREBP及LPK mRNA的表达并下调ACO与CPT-1α mRNA的表达（P＜0.05），对PPAR-γ、PPAR-α、及CD36mRNA无显著性影响（P＞0.05）；SOR（20mg/kg）明显抑制肝脏SREBP-1c、FAS、ACC-1、SCD-1及ChREBP mRNA的过表达（P＜0.05），但对LPK、PPAR-γ、PPAR-α、ACO、CPT-1α及CD36mRNA无明显改变（P＞0.05）。 5．Western blot分析显示：与空白对照组比较，果糖对照组细胞核SREBP-1c和ChREBP的蛋白含量增加（P＜0.05），SOR（20mg/kg）明显抑制SREBP-1c核蛋白的表达（P＜0.05），但没有显著改变ChREBP核蛋白含量（P＞0.05）。 结论 1. SOR（20mg/kg）能够改善高果糖所致大鼠脂肪肝； 2.肝脏SREBP-1c是SOR改善高果糖诱导所致大鼠脂肪肝的重要分子靶点。
[Abstract]:Objective: five layer (SOR) to extract the Dragon herbs for treatment of obesity and diabetes in the tradition, these two diseases are closely related to fatty liver. It has been proved that SOR can improve the fatty liver function, but the mechanism is not clear. This paper aims to study the molecular mechanism of SOR to improve the fatty liver of rats induced by high fructose.
Methods: 24 healthy SD rats were randomly divided into 4 groups (n=6), fed for 10 weeks: (1) control group: free drinking water; (2) the control group were given 10% fructose fructose water (w/v, daily preparation); (3) SOR of low concentration drug group: fructose solution extract of +SOR 5mg/kg; (4) SOR of high dose group: fructose solution extract of +SOR 20mg/kg. every three days on a weight, and the dosage is adjusted according to the weight changes of the rats in treatment group. Treatment group initial fructose water concentration was 10%, daily food intake and drinking water and fructose, fructose intake is calculated, according to the fructose in the control group three days of fructose intake, adjust the concentration of water use fructose and fructose fructose intake group rats in control group. In the tenth week, the night after fasting 14h, weighing, ether anesthesia, removal of the eye blood plasma separation detection of TC, TG, blood glucose, NEFA and insulin content; the rats were sacrificed and the liver weight Liver, H.E staining and oil red O staining, and detection of liver TG, TC content; the total RNA was extracted from liver tissue by TRIzol, reverse transcription cDNA, using Real-Time PCR method to detect the liver lipogenic transcription factor SREBP-1c, ChREBP and its downstream genes FAS, ACC-1, SCD-1, LPK and lipid oxidation related genes of PPAR- alpha, CPT-1 alpha, ACO and PPAR- CD36 gamma, mRNA levels; from the liver nuclear protein expression detection, regulation of lipid synthesis of two key transcription factor SREBP-1c and ChREBP protein by Western blot method. The experimental data using statistical software StatView analysis, mean standard error (x + SEM) said, t test for the comparison of data between the two groups.
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