清脂通脉颗粒对动腺硬化模型家兔血脂及SR-BI mRNA和蛋白表达影响的实验研究

发布时间：2018-03-07 17:18

  本文选题：动脉粥样硬化　切入点：清脂通脉颗粒　出处：《辽宁中医药大学》2017年硕士论文　论文类型：学位论文

【摘要】：目的:通过观察清脂通脉颗粒对动脉粥样硬化(AS)模型家兔的血脂代谢及胆固醇逆转运受体SR-BI mRNA和蛋白表达方面的影响,深入研究在防治动脉粥样硬化方面清脂通脉颗粒的作用机制。材料与方法:1.本实验选取的实验动物为健康雄性家兔,在实验中的动物的分组均采取随机数字表的方法,我们将22只家兔,随机选取6只为空白组,剩余16只为造模组。使各组家兔每天都自由饮水,室温保持在20?C-25?C,适应性饲养一周后,对模型组进行造模,造模方法即采用传统的高脂饲料喂养加维生素D3注射法进行动脉粥样硬化模型的复制。模型组将维生素D3一次性注射于腹腔,注射剂量为(60万IU/kg)。空白组给予正常饲料颗粒喂养,并同样适应性饲养一周后,在实验第一天在腹腔注射同等剂量的生理盐水。高脂饲料喂养,时间为4周,从空白组和模型组各随机抽取1只家兔进行颈动脉病理切片,采用HE染色法观察颈动脉粥样斑块病理改变,评价造模成功与否。造模成功后,按照随机数字表法将造模组15只家兔,按照每组5只随机分为模型组、辛伐他汀组(西药组)、清脂通脉颗粒组(中药组)然后开始对辛伐他汀组(西药组)、清脂通脉颗粒组(中药组)进行药物治疗,两组分别每日一次经口灌饲辛伐他汀、清脂通脉颗粒,同样连续灌饲4周。2.至实验第9周末禁食,对各组家兔进行经耳缘静脉采集血液样本,对血清进行分离,目的是检测各组血清中血脂四项指标,分别包括总胆固醇(TC)、甘油三酯(TG)、低密度脂蛋白(LDL-C)和高密度脂蛋白(HDL-C)的含量采用全自动生化仪,并检测各组家兔氧化型低密度脂蛋白(OX-LDL)的含量通过酶联免疫吸附法(ELISA)。至实验9周末禁食经耳缘静脉麻醉后处死动物,剖开腹部,摘取肝脏尾状叶,检测各组动物肝脏组织中SR-BImRNA及SR-BI蛋白的表达水平分别通过RT-PCR方法和Western-blot方法。3.对四组实验数据进行统计分析,分析软件采用SPSS17.0,其中各组的计量资料以均数±标准差((?)±S)表示,多个组间的数据进行比较运用单因素方差分析,统计结果以P0.01有统计学意义,P0.05则无统计学意义。结果:1.血脂四项检测结果得出:与空白组比较,模型组、西药组、中药组TG、TC、LDL-C含量均升高(P0.01);与模型组比较,西药组,中药组TG、TC、LDL-C含量均降低(P0.01);与西药组比较,中药组TG、TC、LDL-C含量均降低(P0.05);与空白组比较,模型组、西药组、中药组HDL-C含量均降低(P0.01);与模型组比较,西药组、中药组HDL-C含量均升高(P0.01);与西药组比较,中药组HDL-C含量降低(P0.05)。2.氧化型低密度脂蛋白检测结果:与空白组比较,模型组、西药组、中药组OX-LDL含量均升高(P0.01);与模型组比较,西药组、中药组OX-LDL含量均降低(P0.01);与西药组比较,中药组OX-LDL含量升高(P0.05)。3.Western-blot结果显示:SR-BI蛋白的目的条带大约在36k Da处,统计积分光密度值的结果可以得出,与空白组比较,模型组、西药组、中药组SR-BI表达降低(p0.01)。与模型组比较,西药组、中药组SR-BI表达升高(p0.01)。与西药组比较,无显著差异(p0.05)。4.RT-PCR结果显示:SR-BI mRNA的目的条带大约在457bp处,统计积分光密度值的结果可以得出,与空白组比较,模型组、西药组、中药组SR-BImRNA表达降低(p0.01)。与模型组比较,西药组、中药组SR-BImRNA表达升高(p0.01)。与西药组比较,无显著差异(p0.05)。结论:1.通过高脂饲料喂养家兔,发现模型组中含量显著升高的有TC、TG、LDL-C及OX-LDL,含量显著降低的为HDL-C,说明高脂血症动脉硬化兔模型造模成功。2.清脂通脉颗粒使动脉粥样硬化模型家兔肝细胞组织中SR-BI mRNA的表达升高。3.清脂通脉颗粒使动脉粥样硬化模型家兔肝细胞组织中SR-BI蛋白的表达升高。4.清脂通脉颗粒实现其对胆固醇的逆向转运是通过上调肝细胞中SR-BI mRNA及蛋白的表达。5.肝细胞中SR-BI mRNA及蛋白的表达升高,清脂通脉颗粒防治动脉粥样硬化的可能机制是通过促进胆固醇逆转运实现。
[Abstract]:Objective: through the observation of Qingzhi Tongmai Granule on atherosclerosis (AS) metabolism and cholesterol transport reversal rabbit model of receptor SR-BI mRNA and protein expression of the influence of in-depth study of the mechanism of Qing fat Tongmai granule in treating atherosclerosis. Materials and methods: the experimental animal experiment for the selected 1. healthy male rabbits. The packet in the experiment animal adopt the method of random number table, we will be 22 rabbits, 6 rabbits were randomly selected as control group, the remaining 16 rats were model group. The rabbits of each group every day free drinking water, room temperature maintained at 20? C-25? C, afteradaptivebreedingforoneweek, the model group was made die, mold method that using the traditional model of atherosclerosis in high fat diet with vitamin D3 injection in the model group. The replication of vitamin D3 injection in abdominal cavity injection (600 thousand IU/kg) blank. Group with normal feed particle feeding, and the same afteradaptivebreedingforoneweek, saline in the first day in the same dose intraperitoneal injection. High fat diet for 4 weeks, from the blank group and the model group were randomly selected 1 Rabbits Carotid artery pathological sections, the pathological changes of carotid atherosclerotic plaque HE to observe the staining, the evaluation model is successful or not. After the success of the model, according to the random number table method model of 15 rabbits, with 5 rats in each group were randomly divided into model group, simvastatin group (Western medicine group), Qingzhi Tongmai granule group (Chinese medicine group) and simvastatin group (Western medicine group) on Qingzhi, Tongmai granule group (Chinese medicine group) for drug treatment, two groups were once daily oral intragastric administration of simvastatin, Qingzhi Tongmai Granules, the same continuous feeding of 4 weeks.2. to the ninth weekend fasting by ear vein blood samples were collected and analyzed in each group, Separation of serum, the aim is to detect four indexes of blood lipid in serum, including total cholesterol (TC), triglyceride (TG), low density lipoprotein (LDL-C) and high density lipoprotein (HDL-C) content by automatic biochemical analyzer, and were observed by oxidation of low density lipoprotein (the content of OX-LDL) by enzyme linked immunosorbent assay (ELISA). To the 9 weekend of fasting by ear vein anesthesia after the death of animal, open abdomen, liver caudate lobe, the expression level of SR-BImRNA and SR-BI protein in liver tissue were detected in animal respectively by RT-PCR method and Western-blot method of.3. of four groups of experimental data statistics the analysis, using SPSS17.0 analysis software, the measurement data were the mean standard deviation ((?) + S) said that many groups of data were compared using single factor analysis of variance, statistical results in P0.01 have statistical significance, P0. 05鍒欐棤缁熻�″�︽剰涔，

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