【作者】 张瑞娟；

【导师】 杨继军；

【作者基本信息】 河北医科大学 ， 针灸推拿学， 2008， 硕士

【摘要】 目的:加味青麟丸由酒制大黄、党参、制首乌组成,具有益气化瘀降浊的功效,为导师杨继军教授多年临床观察筛选研制的效方,对高脂血症具有较好的治疗效果,为进一步验证加味青麟丸的药效并说明其作用机制,本研究采用高脂饲料复制高脂血症动物模型,同时施加药物干预,旨在通过观察对总胆固醇（TC）、甘油三酯（TG）、高密度脂蛋白（HDL-C）、低密度脂蛋白（LDL-C）的影响,探讨其调节脂质代谢的作用;通过观察对丙二醛（MDA）、超氧化物歧化酶（SOD）的影响,探讨其抗脂质过氧化损伤的作用机制;通过观察对内皮素-1（ET-1）、降钙素基因相关肽（CGRP）、血栓素B2（TXB₂）、6-酮-前列环素F_1α(6-Keto-PGF_1α)的影响,探讨其对血管舒缩状态和血液运行等方面的作用机制。方法:第一部分加味青麟丸对高脂血症小鼠脂代谢的影响选用健康雄性昆明小鼠50只,随机分为5组:⑴正常对照组（简称正常组）;⑵模型对照组（简称模型组）;⑶加味青麟丸低剂量组（简称低剂量组）;⑷加味青麟丸高剂量组（简称高剂量组）;⑸辛伐他汀对照组（简称辛伐他汀组）。除正常组喂饲普通饲料外,其余各组均喂饲高脂饲料。同时,给药组每日分别灌胃加味青麟丸低剂量1.5g/Kg,高剂量3.0g/Kg,辛伐他汀6.6mg/Kg,正常组和模型组每日给予同体积蒸馏水。每次用药体积均按0.2ml/10g计算,各组灌胃1次/天,连续灌胃4周。末次给药禁食不禁水12h后各组小鼠摘除眼球取血,将血样低温离心,分离血清,密封,-20℃冷存待检。血清TC、TG、HDL-C、LDL-C的含量测定均采用生化法,测定步骤和计算公式均严格按照试剂盒说明书操作。第二部分加味青麟丸对高脂血症小鼠血清MDA含量、SOD活力的影响实验动物分组、造模及给药方法同前。连续灌胃4周,于末次给药禁食不禁水12h后各组小鼠摘除眼球取血,将血样低温离心,分离血清,密封,-20℃冷存待检。血清MDA测定采用硫代巴比妥酸（TBA）比色分析法,SOD活力测定采用黄嘌呤氧化酶法,测定步骤和计算公式均严格按照试剂盒说明书操作。第三部分加味青麟丸对高脂血症小鼠血浆ET-1、CGRP及TXB₂、6-Keto-PGF_1α含量的影响实验动物分组、造模及给药方法同前。连续灌胃4周,于末次给药禁食不禁水12h后各组小鼠摘除眼球取血,将血样分成两份,分别装入加有不同抗凝剂的试管中混匀,低温离心,分离血浆,密封,-80℃冷存待检。ET-1、CGRP、TXB₂和6-Keto-PGF_1α均采用放射免疫法,测定步骤和计算公式均严格按照试剂盒说明书操作。结果:第一部分加味青麟丸对高脂血症小鼠脂代谢的影响加味青麟丸对高脂血症小鼠血清中TC、TG、HDL-C、LDL-C含量的影响高脂血症模型组（5.06±0.90、1.98±0.41、2.31±0.39）小鼠血清TC、TG、LDL-C含量均明显高于正常组（3.71±0.30、0.89±0.27、1.57±0.18）（P<0.01）,模型组（1.43±0.18）血清HDL-C含量明显低于正常组（1.81±0.14）（P<0.01）;药物干预各组TC、TG、LDL-C含量均明显低于模型组,HDL-C含量明显高于模型组（P<0.05或P<0.01）;高剂量组（3.95±0.41、1.78±0.12、1.69±0.14）,低剂量组（3.93±0.30、1.79±0.13、1.67±0.15）和辛伐他汀组（4.02±0.33、1.82±0.14、1.66±0.10）之间血清TC、LDL-C和HDL-C的含量无显著性差异（P>0.05）,而高剂量组（1.28±0.19）和低剂量组（1.26±0.45）血清TG的含量均低于辛伐他汀组（1.61±0.16）（P<0.05）,高、低剂量组之间TG的含量无显著性差异（P>0.05）。第二部分加味青麟丸对高脂血症小鼠血清MDA含量、SOD活力的影响1加味青麟丸对高脂血症小鼠血清MDA含量的影响高脂血症模型组（20.94±3.49）小鼠血清MDA含量明显高于正常组（11.94±1.44）（P<0.01）;药物干预各组血清MDA含量均明显低于模型组（P<0.01）;其中高剂量组（13.77±1.41）和低剂量组（15.87±1.51）均低于辛伐他汀组（17.95±1.24）（P<0.05或P<0.01）,而高剂量组低于低剂量组（P<0.05）。2加味青麟丸对高脂血症小鼠血清SOD活力的影响高脂血症模型组（160.6±10.64）小鼠血清SOD活力明显低于正常组（236.2±12.89）（P<0.01）;药物干预各组血清SOD活力均明显高于模型组（P<0.01）;其中高剂量组（205.7±11.82）和低剂量组（193.0±11.55）均高于辛伐他汀组（181.9±11.40）（P<0.05或P<0.01）,而高剂量组高于低剂量组（P<0.05）。第三部分加味青麟丸对高脂血症小鼠血浆ET-1、CGRP及TXB₂、6-Keto-PGF_1α含量的影响1加味青麟丸对高脂血症小鼠血浆ET-1含量的影响高脂血症模型组（126.95±15.46）小鼠血浆ET-1的含量明显高于正常组（67.78±7.04）（P<0.01）;药物干预各组血浆ET-1的含量均明显低于模型组（P<0.01）;其中高剂量组（74.29±6.65）和低剂量组（85.41±9.93）均低于辛伐他汀组（100.43±9.44）（P<0.01）,而高剂量组低于低剂量组（P<0.05）。2加味青麟丸对高脂血症小鼠血浆CGRP含量的影响高脂血症模型组（54.83±7.18）小鼠血浆CGRP的含量明显低于正常组（105.65±11.65）（P<0.01）;药物干预各组血浆CGRP的含量均明显高于模型组（P<0.01）;其中高剂量组（89.63±8.18）和低剂量组（80.49±8.28）均高于辛伐他汀组（67.54±6.79）（P<0.01）,而高剂量组高于低剂量组（P<0.05）。3加味青麟丸对高脂血症小鼠血浆TXB₂含量的影响高脂血症模型组（2704.24±370.59）小鼠血浆TXB₂的含量明显高于正常组（1514.64±342.93）（P<0.01）;药物干预各组血浆TXB₂的含量均明显低于模型组（P<0.05或P<0.01 ）;其中高剂量组（ 1883.46±332.30 ）和低剂量组（1893.38±237.39）均低于辛伐他汀组（2294.48±340.49）（P<0.05）,而高、低剂量组之间无显著性差异（P>0.05）。4加味青麟丸对高脂血症小鼠血浆6-Keto-PGF_1α含量的影响高脂血症模型组（924.37±153.28）小鼠血浆6-Keto-PGF_1α的含量明显低于正常组（1370.19±131.21）（P<0.01）;药物干预各组血浆6-Keto-PGF_1α均明显高于模型组（P<0.05或P<0.01）;其中高剂量组（1236.70±144.09）和低剂量组（1245.92±157.26）均高于辛伐他汀组（1093.73±135.86）（P<0.05）,而高、低剂量组之间无显著性差异（P>0.05）。5加味青麟丸对高脂血症小鼠血浆TXB₂/6-Keto-PGF_1α的影响高脂血症模型组（2.98±0.52）小鼠血浆TXB₂ / 6-Keto-PGF_1α明显高于正常组（1.10±0.23）（P<0.01）;用药干预各组血浆TXB₂/6-Keto-PGF_1α均明显低于模型组（P<0.01）;其中高剂量组（1.53±0.31）和低剂量组（1.47±0.24）血浆TXB₂/6-Keto-PGF_1α均低于辛伐他汀组（2.12±0.40）（P<0.01）,而高、低剂量组之间无显著性差异（P>0.05）。结论:1加味青麟丸可降低高脂血症模型小鼠血清TC、TG、LDL-C含量,升高HDL-C含量,具有较好的调节脂质代谢的作用。2该方可降低高脂血症模型小鼠血清MDA含量,提高SOD活力,抑制机体脂质过氧化反应,增加抗氧化能力,通过改善自由基代谢,以减轻高脂血症引发的氧化损伤。3该方可降低高脂血症模型小鼠血浆ET-1、TXB₂含量,升高CGRP、6-Keto-PGF_1α含量,降低TXB₂/6-Keto-PGF_1α,通过调节血管平滑肌收缩与舒张,抑制血小板聚集和血栓形成,以维持血管张力及血流畅通,减轻高脂血症引发的血管内皮损伤,从而保护血管内皮功能。更多还原

【Abstract】 Objective: Added Herbs’QingLin Pill （composed of jiuzhidahuang, dangshen, zhishouwu） has the function of benefiting vital energy, removing stasis and phlegm and is an effective formula through years of clinical observation by Professor YANG Ji-jun. And it has been proved significantly effect in treating hyperlipemia. In order to further probe its action mechanism, we duplicated the hyperlipemia animal model with high-fat forage, At the same time, we interfered the animal with medicine. And we studied the action mechanism of this formula in regulating lipid metabolism through observing its effect on TC,TG,HDL-C,LDL-C; We also studied its action mechanism in resisting lipid peroxidation injury through observing its effect on MDA,SOD; We studied the action mechanism of this formula in vasomotion state and blood flow through observing its effect on ET-1,CGRP,TXB₂ and 6-Keto-PGF_1α.Methods:PartⅠEffect of Added Herbs’QingLin Pill on lipid metabolism of hyperlipemia mice50 healthy male KM mice were divided into 5 groups at random, namely, （1） Normal control group（normal group for short）; （2） Model control group（model group for short）; （3） Low-dose group of Added Herbs’Qing-Lin Pill group（low-dose group for short）; （4） High-dose group of Added Herbs’Qing-Lin Pill group （high-dose group）; （5） Simvastatin control group （simvastatin group for short）. Except for normal group was fed with ordinary forage, the rest of groups were all fed with high-fat forage. At the same time, the treatment groups were given low-dose of Added Herbs’Qing-Lin Pill 1.5 g /Kg, high-dose of Added Herbs’Qing-Lin Pill 3.0 g /Kg and Simvastatin 6.6 mg /Kg per day. Normal and model group were given distilled water. Mice in each group were given stomach-perfusion once a day and every time the drug volume was calculated by 0.2 ml/10g. Stomach-perfusion lasted for 4 weeks. Then 12 hours after fasting and not prohibiting water of the last time, the mice were removed eyeball and their blood were drawn out, the blood sample was made centrifugation with low temperature and the blood serum was isolated. It was sealed up and keep temperature of under -20℃to examine them later. Finally the contents of TC,TG,HDL-C,LDL-C were measured with biochemistry method. The contents of each item were measured strictly according to the directions of reagent box.PartⅡEffect of Added Herbs’QingLin Pill on the content of MDA and the activity of SOD in serum of hyperlipemia miceThe group distribution, duplication of the hyperlipemia animal model and drug-given were exactly same as above. After 4 weeks continuous stomach-perfusion, then 12 hours after fasting and not prohibiting water of the last time, the mice were removed eyeball and their blood were drawn out. The blood sample was made centrifugation with low temperature and the blood serum was isolated. It was sealed up and keep temperature of under -20℃to examine them later. The content of MDA was examined by the method of TBA colorimeter analysis and activity of SOD was examined by the method of xanthine oxidase. The content of each item was measured strictly according to the directions of reagent box.PartⅢEffect of Added Herbs’QingLin Pill on the contents of ET-1, CGRP, TXB₂ and 6-Keto-PGF_1α in blood plasma of hyperlipemia miceThe group distribution, duplication of the hyperlipemia animal model and drug-given are exactly same as above. After 4 weeks continuous stomach-perfusion, then 12 hours after fasting of the last time, the mice were removed eyeball and their blood were drawn out. The blood sample was divided two portions and builded in sample tubes of containing different decoagulant and misced bene. The blood sample was made centrifugation with low temperature and the plasma was isolated. It was sealed up and keep temperature of under -80℃to examine them later.Finally the contents of ET-1, CGRP, TXB₂ and 6-Keto-PGF_1α were all measured with radio-immuno method. The content of each item was measured strictly according to the directions of reagent box.Results: PartⅠEffect of Added Herbs’QingLin Pill on lipid metabolism of hyperlipemia miceEffect of Added Herbs’QingLin Pill on the contents of TC, TG, HDL-C and LDL-C in serum of hyperlipemia miceThe contents of TC, TG, LDL-C in serum of model group （5.06±0.90, 1.98±0.41, 2.31±0.39） were all higher than normal group （3.71±0.30, 0.89±0.27, 1.57±0.18） （P < 0.01）, while HDL-C of model group （1.43±0.18） was lower than normal group （1.81±0.14） （P<0.01）; After the intervention of medicine, the contents of TC, TG, LDL-C in serum of each treatment group were all lower than the model group, while HDL-C was higher than the model group （P<0.05 or P<0.01）; The contents of TC, LDL-C and HDL-C in serum were no significant difference in high-dose group （3.95±0.41, 1.78±0.12, 1.69±0.14）, low-dose group （3.93±0.30, 1.79±0.13, 1.67±0.15） and simvastatin group （4.02±0.33, 1.82±0.14, 1.66±0.10） （P>0.05）. But the contents of TG in low-dose group （1.26±0.45） and high-dose group （1.28±0.19） were lower than simvastatin group （1.61±0.16） （P<0.05）. And there was no significant difference between low-dose and high-dose group （P>0.05）.PartⅡEffect of Added Herbs’QingLin Pill on the content of MDA and the activity of SOD in serum of hyperlipemia mice1 Effect of Added Herbs’QingLin Pill on the content of MDA in serum of hyperlipemia miceThe content of MDA in serum of model group （20.94±3.49） was higher than the normal group （11.94±1.44） （P<0.01）; After the intervention of medicine, the content of MDA in serum of each treatment group was lower than the model group （P<0.01）; But the contents of MDA in low-dose group （15.87±1.51） and high-dose group （13.77±1.41） were lower than simvastatin group （17.95±1.24） （P<0.05 or P<0.01）. The high-dose group was lower than the low-dose group （P<0.05）.2 Effect of Added Herbs’QingLin Pill on the activity of SOD in serum of hyperlipemia miceThe activity of SOD in serum of model group （160.6±10.64） was lower than the normal group （236.2±12.89） （P<0.01）; After the intervention of medicine, the activity of SOD in serum of each treatment group was higher than the model group （P<0.01）; But the activity of SOD in low-dose group （193.0±11.55） and high-dose group （205.7±11.82） were higher than simvastatin group （181.9±11.40） （P<0.05 or P<0.01）. The high-dose group was higher than the low-dose group （P<0.05）.PartⅢEffect of Added Herbs’QingLin Pill on the contents of ET-1, CGRP,TXB₂ and 6-Keto-PGF_1α in blood plasma of hyperlipemia mice1 Effect of Added Herbs’QingLin Pill on the content of ET-1 in plasma of hyperlipemia miceThe content of ET-1 in plasma of model group （126.95±15.46） was higher than the normal group （67.78±7.04） （P<0.01）; After the intervention of medicine, the content of ET-1 in plasma of each treatment group was lower than the model group （P<0.01）; But the contents of ET-1 in low-dose group （85.41±9.93） and high-dose group （74.29±6.65） were lower than simvastatin group （100.43±9.44） （P<0.01）. The high-dose group was lower than the low-dose group （P<0.05）.2 Effect of Added Herbs’QingLin Pill on the content of CGRP in plasma of hyperlipemia miceThe content of CGRP in plasma of model group （54.83±7.18） was lower than the normal group （105.65±11.65） （P<0.01）; After the intervention of medicine, the content of CGRP in plasma of each treatment group was higher than the model group （P<0.01）; But the contents of CGRP in low-dose group （80.49±8.28） and high-dose group （89.63±8.18） were higher than simvastatin group （67.54±6.79） （P<0.01）. The high-dose group was higher than the low-dose group （P<0.05）.3 Effect of Added Herbs’QingLin Pill on the content of TXB₂ in plasma of hyperlipemia miceThe content of TXB₂ in plasma of model group （2704.24±370.59） was higher than the normal group （1514.64±342.93） （P<0.01）; After the intervention of medicine, the content of TXB₂ in plasma of each treatment group was lower than the model group （P<0.05 or P<0.01）; But the contents of TXB₂ in low-dose group （1893.38±237.39） and high-dose group （1883.46±332.30） were lower than simvastatin group （2294.48±340.49） （P<0.05）. And there was no significant difference between low-dose and high-dose group（P>0.05）.4 Effect of Added Herbs’QingLin Pill on the content of 6-Keto-PGF_1α in plasma of hyperlipemia mice The content of 6-Keto-PGF_1α in plasma of model group （924.37±153.28） was lower than the normal group （1370.19±131.21） （P<0.01）; After the intervention of medicine, the content of 6-Keto-PGF_1α in plasma of each treatment group was higher than the model group （P<0.05 or P<0.01）; But the contents of 6-Keto-PGF_1α in low-dose group （1245.92±157.26） and high-dose group （1236.70±144.09） were higher than simvastatin group （1093.73±135.86） （P<0.05）. And there was no significant difference between low-dose and high-dose group （P>0.05）.5 Effect of Added Herbs’QingLin Pill on TXB₂/ 6-Keto -PGF_1α in plasma of hyperlipemia miceThe TXB₂/6-Keto-PGF_1α in plasma of model group （2.98±0.52） was higher than the normal group （1.10±0.23） （P<0.01）; After the intervention of medicine, the TXB₂/6-Keto-PGF_1α in plasma of each treatment group was lower than the model group （P<0.01）; But the TXB₂/6-Keto-PGF_1α in low-dose group （1.47±0.24） and high-dose group （1.53±0.31） were lower than simvastatin group （2.12±0.40） （P<0.05）. And there was no significant difference between low-dose and high-dose group （P>0.05）.Conclusion:1 Added Herbs’QingLin Pill has decreased the contents of TC,TG,LDL-C and increased the content of HDL-C in serum. The result shows that it has marked function of regulating lipid metabolism. 2 It obviously increased the activity of SOD, and decreased the content of MDA in serum. The result shows that it has marked function of inhibiting lipid peroxidation reaction and raising anti-oxidative ability. It could reduce oxidative damage of hyperlipemia through amending free radical metabolism.3 It obviously decreased the contents of ET-1 with TXB₂ and increases the contents of CGRP with 6-Keto-PGF_1α in plasma. And it obviously regulated the proportion of TXB₂ and 6-Keto-PGF_1α in plasma. The results shows that it has marked function of regulating the normal contraction and dilation state of vascular smooth muscle , which in order to remove the platelet coagulation with thrombosis. It could prevent the blood vessel endothelium damage of hyperlipemia and protect blood vessel endothelium function.更多还原