电针对动脉粥样硬化家兔肝脏中CYP7A1表达的影响
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摘要
目的:观察电针"内关""关元""足三里"对动脉粥样硬化家兔肝脏中胆固醇7α-羟化酶(CYP7A1)表达的影响,探讨针刺防治动脉粥样硬化的机制。方法:将26只雄性家兔,分笼饲养,适应性喂养1周,随机选取7只为空白组,余下19只为造模组。空白组喂以普通饲料,造模组给予高脂饲料,高脂饲料喂养4周后行颈总动脉球囊损伤术,术后继续予高脂饲料喂养4周,从空白组和造模组各随机选1只家兔做颈动脉病理切片,采用HE染色法观察颈动脉粥样斑块病理形态,确定造模成功与否。造模成功后,将造模组18只家兔随机分为3组:模型组、电针组和西药组,每组6只。模型组不给予任何治疗;西药组将阿托伐他汀钙片混悬液灌胃给药;电针组选"内关""关元""足三里"进行电针治疗,每日1次,每次留针20 min,6d为一疗程,疗程间休息1 d,共4个疗程。治疗结束后,在家兔耳缘静脉采血,检测各组家兔胆固醇(CHO)、三酰甘油(TG)、低密度脂蛋白(LDL)、高密度脂蛋白(HDL)含量;取家兔肝脏组织,采用Western blot法检测家兔肝脏中CYP7A1蛋白表达水平;采用RT-PCR检测家兔肝脏中CYP7A1mRNA表达水平。结果:与空白组比较,模型组家兔血中CHO、TG及LDL含量显著升高,HDL含量明显降低,肝脏中CYP7A1蛋白及CYP7A1 mRNA表达量降低,差异有统计学意义(均P<0.01);干预后,与模型组比较,电针组和西药组家兔血中CHO、TG及LDL降低,HDL升高,CYP7A1蛋白及CYP7A1mRNA表达量均明显升高,差异有统计学意义(均P<0.01),电针组和西药组各指标比较,差异均无统计学意义(均P>0.05)。结论:电针能明显改善动脉粥样硬化家兔血脂水平,促进家兔肝脏中CYP7A1 mRNA的表达,这可能是电针治疗动脉粥样硬化的作用机制之一。
Objective To observe the effects of electroacupuncture(EA) at "Neiguan"(PC 6), "Guanyuan"(CV 4) and "Zusanli"(ST 36) on CYP7 A1 expression in liver of rabbits with atherosclerosis(AS), and to explore the mechanism of acupuncture for prevention and treatment of atherosclerosis. Methods A total of 26 male rabbits were adaptively fed for 1 week in different cages. Seven rabbits were randomly divided into a blank group, and the remaining 19 rabbits were divided into a model group. The blank group was fed with normal diet, while the model group was fed with high-fat diet. After high-fat diet for 4 weeks, the rabbits in the model group were treated with balloon injury surgery of the common carotid artery; after surgery, the rabbits were fed with high-fat diet for 4 weeks. One rabbit was randomly selected from the blank group and model group to obtain the pathological section of carotid artery; the HE staining was used to observe the pathomorphology of atheromatous plaque to determine the success of modeling or not. After successful establishment of modeling, 18 rabbits were randomly divided into a AS model group, an EA group and a medication group, 6 rabbits in each one. The rabbits in the AS model group received no treatment; the rabbits in the medication group were treated with intragastric administration of atorvastatin calcium tablets; the rabbits in the EA group were treated with EA at "Neiguan"(PC 6), "Guanyuan"(CV 4) and "Zusanli"(ST 36), once a day, 20 min per treatment; six-day EA treatment constituted one course, and totally 4 courses were given with an interval of 1 day between courses. After treatment, vein blood was collected from rabbit ear, and cholesterol(CHO), triglyceride(TG), low-density lipoprotein(LDL) and high-density lipoprotein(HDL) were measured in each group; the CYP7 A1 protein expression in rabbit liver was measured by Westernblot method, and CYP7 A1 mRNA expression in rabbit liver was measured by RT-PCR. Results Compared with the blank group, the contents of CHO, TG and LDL in the AS model group were significantly increased, but HDL was significantly decreased, and the expression of CYP7 A1 protein and CYP7 A1 mRNA in the liver were significantly decreased(all P<0.01).After treatment, compared with the model group, the contents of CHO, TG and LDL in the EA group and medication group were significantly reduced, but HDL was significantly increased, and the expression of CYP7 A1 protein and CYP7 A1 mRNA in the liver were significantly increased(all P<0.01). The significant difference of each outcome between the EA group and medication group was not observed(all P>0.05). Conclusion EA could significantly improve blood lipid and promote the expression of CYP7 A1 mRNA in rabbits with atherosclerotic, which may be one of the mechanisms of EA for atherosclerosis.
Objective To observe the effects of electroacupuncture(EA) at "Neiguan"(PC 6), "Guanyuan"(CV 4) and "Zusanli"(ST 36) on CYP7 A1 expression in liver of rabbits with atherosclerosis(AS), and to explore the mechanism of acupuncture for prevention and treatment of atherosclerosis. Methods A total of 26 male rabbits were adaptively fed for 1 week in different cages. Seven rabbits were randomly divided into a blank group, and the remaining 19 rabbits were divided into a model group. The blank group was fed with normal diet, while the model group was fed with high-fat diet. After high-fat diet for 4 weeks, the rabbits in the model group were treated with balloon injury surgery of the common carotid artery; after surgery, the rabbits were fed with high-fat diet for 4 weeks. One rabbit was randomly selected from the blank group and model group to obtain the pathological section of carotid artery; the HE staining was used to observe the pathomorphology of atheromatous plaque to determine the success of modeling or not. After successful establishment of modeling, 18 rabbits were randomly divided into a AS model group, an EA group and a medication group, 6 rabbits in each one. The rabbits in the AS model group received no treatment; the rabbits in the medication group were treated with intragastric administration of atorvastatin calcium tablets; the rabbits in the EA group were treated with EA at "Neiguan"(PC 6), "Guanyuan"(CV 4) and "Zusanli"(ST 36), once a day, 20 min per treatment; six-day EA treatment constituted one course, and totally 4 courses were given with an interval of 1 day between courses. After treatment, vein blood was collected from rabbit ear, and cholesterol(CHO), triglyceride(TG), low-density lipoprotein(LDL) and high-density lipoprotein(HDL) were measured in each group; the CYP7 A1 protein expression in rabbit liver was measured by Westernblot method, and CYP7 A1 mRNA expression in rabbit liver was measured by RT-PCR. Results Compared with the blank group, the contents of CHO, TG and LDL in the AS model group were significantly increased, but HDL was significantly decreased, and the expression of CYP7 A1 protein and CYP7 A1 mRNA in the liver were significantly decreased(all P<0.01).After treatment, compared with the model group, the contents of CHO, TG and LDL in the EA group and medication group were significantly reduced, but HDL was significantly increased, and the expression of CYP7 A1 protein and CYP7 A1 mRNA in the liver were significantly increased(all P<0.01). The significant difference of each outcome between the EA group and medication group was not observed(all P>0.05). Conclusion EA could significantly improve blood lipid and promote the expression of CYP7 A1 mRNA in rabbits with atherosclerotic, which may be one of the mechanisms of EA for atherosclerosis.
引文
[1]焦艳,金蓉.动脉粥样硬化发病机制及影像学研究进展[J].国际生物医学工程杂志,2013,36(6):362-366.
[2]杜雪,李进军,卢立志,等.胆固醇7α-羟化酶基因研究进展[J].浙江农林大学学报,2013,30(5):755-760.
[3]钟一鸣,钟华平,廖伟,等.大鼠颈总动脉球囊损伤模型建立及其动态病理学和P27的变化[J].中国现代医学杂志,2014,24(13):13-17.
[4]沈琳,高海青,邱洁,等.葡萄籽原花青素对实验性动脉粥样硬化兔基质金属蛋白酶和氧化低密度脂蛋白的影响[J].中国心血管杂志,2006,11(5):331-333.
[5]管耘园,叶炳华,卢辉和,等.球囊损伤加高脂喂养建立兔颈动脉粥样硬化模型[J].实验动物科学,2007,24(1):6-9.
[6]田飞,石怀瑞,张新平.兔颈动脉球囊损伤动物模型的构建改良[J].中国医学创新,2012,9(11):141-142.
[7]李忠仁.实验针灸学[M].北京:中国中医药出版社,2003:147-148.
[8]赵莹,付军.肠道菌群与心血管疾病的研究进展[J].中国老年病学杂志,2014,34(5):1443-1444.
[9]Lange Y,Steck TL.The role of intracellular cholesterol transport in cholesterol homeostasis[J].Trends Cell Biol,1996,6(6):205-208.
[10]刘浩,余金明,陈芳,等.冠心病患者血脂代谢异常的流行病学研究[J].中国实用内科杂志,2007,27(12):965-967.
[11]Lu TT,Makishima M,Repa JJ,et al.Molecular basis for feedback regulation of bile acid synthesis by nuclear receptors[J].Mol Cell,2000,6(3):507-515.
[12]Vlahcevic Z,Heuman DM,Hylemon PB.Regulation of bile acid synthesis[J].Hepatology,1991,13(3):590-600.
[13]Hubacek JA,Bobkova D.Role of cholesterol 7α-hydroxylase(CYP7A1)in nutrigenetics and pharmacogenetics of cholesterol lowering[J].Mol Diagn Ther,2006,10(2):93-100.
[14]Zong C,Yu Y,Song G,et al.Chitosan oligosaccharides promote reverse cholesterol transport and expression of scavenger receptor BI and CYP7A1 in mice[J].Exp Biol Med,2012,237(2):194-200.
[15]Myant NB,Mitropoulos KA.Cholesterol 7α-hydroxylase[J].JLipid Res,1977,18(2):135-153.
[16]Post SM,de Crom R,van Haperen R,et al.Increased fecal bile acid excretion in transgenic mice with elevated expression of human phospholipid transfer protein[J].Arterioscler Thromb Vasc Biol,2003,23(5):892-897.
[17]Norlin M,Wikvall K.Enzymes in the conversion of cholesterol into bile acids[J].Curr Mol Med,2007,7(2):199-218.
[18]张文来,王伟志.针灸对颈动脉粥样硬化患者颈动脉血管和血流的影响[J].中国针灸,2011,31(9):783-785.
[19]张莉,刘雨英,张锴.针刺治疗下肢闭塞性动脉硬化症29例疗效观察[J].黑龙江中医药,2008,37(2):41.
[20]陶文剑,詹冬梅,艾炳蔚,等.针刺对ApoE-/-小鼠主动脉窦斑块病理变化因素的影响[J].南京中医药大学学报,2014,30(1):41-45.
[21]肖颖,乐薇,黄浩,等.电针“丰隆”穴对高脂血症大鼠血脂及巨噬细胞肿瘤坏死因子-α、白介素-6的影响[J].针刺研究,2013,38(6):459-464.
[22]封笑.针刺左侧内关穴对健康青年大脑中动脉血流动力学影响的研究[D].哈尔滨:黑龙江中医药大学,2013.
[23]夏春梅.针刺内关穴调节延髓NO/NOS系统及其有关神经递质改善大鼠心肌缺血作用机制探讨[D].上海:复旦大学,2008.
[24]姜小英.艾灸足三里、关元穴对血液流变性的影响[J].四川中医,2001,19(3):75.
[25]宋少军,李学玉,韩秀华,等.灸关元、大椎改善阳虚证血液流变指标的临床研究[J].辽宁中医杂志,2011,38(8):1641-1642.
[2]杜雪,李进军,卢立志,等.胆固醇7α-羟化酶基因研究进展[J].浙江农林大学学报,2013,30(5):755-760.
[3]钟一鸣,钟华平,廖伟,等.大鼠颈总动脉球囊损伤模型建立及其动态病理学和P27的变化[J].中国现代医学杂志,2014,24(13):13-17.
[4]沈琳,高海青,邱洁,等.葡萄籽原花青素对实验性动脉粥样硬化兔基质金属蛋白酶和氧化低密度脂蛋白的影响[J].中国心血管杂志,2006,11(5):331-333.
[5]管耘园,叶炳华,卢辉和,等.球囊损伤加高脂喂养建立兔颈动脉粥样硬化模型[J].实验动物科学,2007,24(1):6-9.
[6]田飞,石怀瑞,张新平.兔颈动脉球囊损伤动物模型的构建改良[J].中国医学创新,2012,9(11):141-142.
[7]李忠仁.实验针灸学[M].北京:中国中医药出版社,2003:147-148.
[8]赵莹,付军.肠道菌群与心血管疾病的研究进展[J].中国老年病学杂志,2014,34(5):1443-1444.
[9]Lange Y,Steck TL.The role of intracellular cholesterol transport in cholesterol homeostasis[J].Trends Cell Biol,1996,6(6):205-208.
[10]刘浩,余金明,陈芳,等.冠心病患者血脂代谢异常的流行病学研究[J].中国实用内科杂志,2007,27(12):965-967.
[11]Lu TT,Makishima M,Repa JJ,et al.Molecular basis for feedback regulation of bile acid synthesis by nuclear receptors[J].Mol Cell,2000,6(3):507-515.
[12]Vlahcevic Z,Heuman DM,Hylemon PB.Regulation of bile acid synthesis[J].Hepatology,1991,13(3):590-600.
[13]Hubacek JA,Bobkova D.Role of cholesterol 7α-hydroxylase(CYP7A1)in nutrigenetics and pharmacogenetics of cholesterol lowering[J].Mol Diagn Ther,2006,10(2):93-100.
[14]Zong C,Yu Y,Song G,et al.Chitosan oligosaccharides promote reverse cholesterol transport and expression of scavenger receptor BI and CYP7A1 in mice[J].Exp Biol Med,2012,237(2):194-200.
[15]Myant NB,Mitropoulos KA.Cholesterol 7α-hydroxylase[J].JLipid Res,1977,18(2):135-153.
[16]Post SM,de Crom R,van Haperen R,et al.Increased fecal bile acid excretion in transgenic mice with elevated expression of human phospholipid transfer protein[J].Arterioscler Thromb Vasc Biol,2003,23(5):892-897.
[17]Norlin M,Wikvall K.Enzymes in the conversion of cholesterol into bile acids[J].Curr Mol Med,2007,7(2):199-218.
[18]张文来,王伟志.针灸对颈动脉粥样硬化患者颈动脉血管和血流的影响[J].中国针灸,2011,31(9):783-785.
[19]张莉,刘雨英,张锴.针刺治疗下肢闭塞性动脉硬化症29例疗效观察[J].黑龙江中医药,2008,37(2):41.
[20]陶文剑,詹冬梅,艾炳蔚,等.针刺对ApoE-/-小鼠主动脉窦斑块病理变化因素的影响[J].南京中医药大学学报,2014,30(1):41-45.
[21]肖颖,乐薇,黄浩,等.电针“丰隆”穴对高脂血症大鼠血脂及巨噬细胞肿瘤坏死因子-α、白介素-6的影响[J].针刺研究,2013,38(6):459-464.
[22]封笑.针刺左侧内关穴对健康青年大脑中动脉血流动力学影响的研究[D].哈尔滨:黑龙江中医药大学,2013.
[23]夏春梅.针刺内关穴调节延髓NO/NOS系统及其有关神经递质改善大鼠心肌缺血作用机制探讨[D].上海:复旦大学,2008.
[24]姜小英.艾灸足三里、关元穴对血液流变性的影响[J].四川中医,2001,19(3):75.
[25]宋少军,李学玉,韩秀华,等.灸关元、大椎改善阳虚证血液流变指标的临床研究[J].辽宁中医杂志,2011,38(8):1641-1642.