健脾化痰祛瘀方对动脉粥样硬化巴马小型猪心肌细胞氧化应激及凋亡的影响
|
摘要
目的:观察健脾化痰祛瘀方对脾虚痰浊动脉粥样硬化巴马小型猪心肌细胞氧化应激及凋亡的影响。方法:健康巴马小型猪15只,月龄6～8个月,随机分为正常对照组、模型组、中药治疗组3组,每组5只。造模:正常对照组予基础饲料喂饲,模型组、中药治疗组采用高脂高热量喂养。药物干预:高脂喂饲24周后健脾化痰祛瘀组予健脾化痰祛瘀方药搅拌于饲料中喂食。检测各组巴马小型猪的心肌ROS、CAT、MDA、GSH-PX的含量;Western Blot方法检测各组巴马小型猪心肌细胞Bax、Bcl-2、Cyto C、Caspase3的含量。结果:(1)与正常对照组相比,模型组心肌线粒体ROS、MDA含量明显上升(P<0.05、P<0.01),CAT、GSH-PX含量明显下降(P<0.01);与模型组相比,中药治疗组心肌线粒体ROS、MDA含量明显下降(P<0.01),CAT、GSH-PX含量明显上升(P<0.05)。(2)与正常对照组相比,模型组心肌细胞Bax、Cyto C、Caspase3蛋白表达明显上升(P<0.01),Bcl-2蛋白表达明显下降(P<0.01);与模型组相比,中药治疗组心肌细胞Bax、Cyto C、Caspase3蛋白表达明显下降(P<0.01、P<0.05),Bcl-2含量明显上升(P<0.05)。结论:(1)健脾化痰祛瘀方可以减少脾虚痰浊动脉粥样硬化巴马小型猪心肌线粒体ROS、MDA的生成,增加其抗氧化酶CAT、GSH-PX的表达,从而减少了心肌线粒体的氧化应激水平。(2)健脾化痰祛瘀法可以减少脾虚痰浊动脉粥样硬化巴马小型猪心肌细胞Bax、Cyto C,Caspase3表达,增加Bcl-2的表达,进而改善心肌细胞凋亡水平。
Objective:To observe the effect of invigorating spleen, removing phlegm and eliminating blood stasis recipe on oxidative stress and apoptosis of Myocardial cells in spleen deficiency and phlegm turbidity atherosclerosis Bama mini-pigs.Methods: There were 15 health Bama mini-pigs, 6～8 months of age, randomly divided into normal group, model group and invigorating spleen, removing phlegm and eliminating blood stasis groups. Each group had 5 pigs in it. Modeling: the normal control group were fed the basal diet, the other groups were fed high fat and high calorie fodder. The invigorating spleen, removing phlegm and eliminating blood stasis group started establish spleen deficiency and phlegm turbidity models. After the 24 th week feeding by fat invigorating spleen, removing phlegm and eliminating blood stasis group started to be fed by stirring the Chinese medicine in it. At the end of 48 th week, the myocardial tissue of the pigs were collected out Tested each mini-pig's myocardial mitochondrial ROS, CAT, MDA, GSH-PX levels. Detected each group Bama mini-pig cardiomyocytes Bax, Bcl-2, the content of Cytochrome C, Caspase3.Results:(1)Bama mini-pig myocardium ROS、MDA content detection: For Myocardial ROS、MDA content in each group comparison, compared with the normal control group,the content of ROS、MDAin model group was significantly increased than the normal group(P<0.05、P<0.01),CAT、GSH-PX in model group was significantly lower than the normal group(P<0.01);compared with the model group,the content of ROS、MDA in Chinese medicine treatment group was significantly lower than the model group( P<0.01),the content of CAT、GSH-PX in Chinese medicine treatment group was significantly increased than the model group(P<0.05).(2)For each group of Bax、Cyto C、Caspase 3 protein expression in comparison, compared with the normal control group,the express of Bax、Cyto C、Caspase 3protein in model group was significantly increased than the normal group(P<0.01),the express of Bcl-2 protein in model group was significantly lower than the normal group(P<0.01)compared with the modle group,the express of Bax、Cyto C、Caspase 3protein in Chinese medicine treatment group was significantly lower than the modle group(P<0.01、P<0.05).the express of Bcl-2 protein in Chinese medicine treatment group was significantly increased than the modle group(P<0.05).Conclusion:The recipe of invigorating spleen, removing phlegm and eliminating blood stasis can reduce the oxygen free radicals of the myocardial, MDA content and increase CAT, GSH-PX expression, reduce the oxidative stress of the mitochondrial, reduce the Bax, Cyto C, Caspase3 expression, increase the expression of Bcl-2 reduce the damage of the myocardial cells in spleen deficiency and phlegm turbidity atherosclerosis Bama mini pigs.
Objective:To observe the effect of invigorating spleen, removing phlegm and eliminating blood stasis recipe on oxidative stress and apoptosis of Myocardial cells in spleen deficiency and phlegm turbidity atherosclerosis Bama mini-pigs.Methods: There were 15 health Bama mini-pigs, 6～8 months of age, randomly divided into normal group, model group and invigorating spleen, removing phlegm and eliminating blood stasis groups. Each group had 5 pigs in it. Modeling: the normal control group were fed the basal diet, the other groups were fed high fat and high calorie fodder. The invigorating spleen, removing phlegm and eliminating blood stasis group started establish spleen deficiency and phlegm turbidity models. After the 24 th week feeding by fat invigorating spleen, removing phlegm and eliminating blood stasis group started to be fed by stirring the Chinese medicine in it. At the end of 48 th week, the myocardial tissue of the pigs were collected out Tested each mini-pig's myocardial mitochondrial ROS, CAT, MDA, GSH-PX levels. Detected each group Bama mini-pig cardiomyocytes Bax, Bcl-2, the content of Cytochrome C, Caspase3.Results:(1)Bama mini-pig myocardium ROS、MDA content detection: For Myocardial ROS、MDA content in each group comparison, compared with the normal control group,the content of ROS、MDAin model group was significantly increased than the normal group(P<0.05、P<0.01),CAT、GSH-PX in model group was significantly lower than the normal group(P<0.01);compared with the model group,the content of ROS、MDA in Chinese medicine treatment group was significantly lower than the model group( P<0.01),the content of CAT、GSH-PX in Chinese medicine treatment group was significantly increased than the model group(P<0.05).(2)For each group of Bax、Cyto C、Caspase 3 protein expression in comparison, compared with the normal control group,the express of Bax、Cyto C、Caspase 3protein in model group was significantly increased than the normal group(P<0.01),the express of Bcl-2 protein in model group was significantly lower than the normal group(P<0.01)compared with the modle group,the express of Bax、Cyto C、Caspase 3protein in Chinese medicine treatment group was significantly lower than the modle group(P<0.01、P<0.05).the express of Bcl-2 protein in Chinese medicine treatment group was significantly increased than the modle group(P<0.05).Conclusion:The recipe of invigorating spleen, removing phlegm and eliminating blood stasis can reduce the oxygen free radicals of the myocardial, MDA content and increase CAT, GSH-PX expression, reduce the oxidative stress of the mitochondrial, reduce the Bax, Cyto C, Caspase3 expression, increase the expression of Bcl-2 reduce the damage of the myocardial cells in spleen deficiency and phlegm turbidity atherosclerosis Bama mini pigs.
引文
[1] 张璐,孙晴,郭书文,等.调脂通脉中药对动脉粥样硬化兔模型内皮细胞标志物的影响[J].中国中医急症,2018(10):1777-1780.
[2] 郭丛丛,黄力,董建军,等.冠状动脉粥样硬化性心脏病血瘀证与冠脉病变程度及血管内皮功能的相关性研究[J].上海中医药杂志,2018,52(9):15-18.
[3] 刘君花,欧阳恩鸿,贺志明,等.虎杖苷对动脉粥样硬化大鼠血脂的调节和CD36、VCAM-1表达的影响[J].邵阳学院学报(自然科学版),2018,15(4):99-104.
[4] 胡月,李潞,赵红丽.血管内皮功能与冠状动脉粥样硬化发生发展的研究进展[J].中西医结合心脑血管病杂志,2018,16(11):1525-1528.
[5] 许晓虹(Ruby).“心胆论治”针灸抗动脉粥样硬化斑块的临床与分子机制研究[D].广州:广州中医药大学,2017.
[6] 马艺鑫,张妮,贾连群,等.基于“痰浊血瘀”病机探讨自噬与动脉粥样硬化的关系[J].时珍国医国药,2016,27(9):2240-2242.
[7] 陈丝,杨关林,王群,等.基于“异病同治”探讨“从脾论治”高脂血症及动脉粥样硬化共同理论基础及临床应用[J].中华中医药学刊,2018,36(9):2200-2202.
[8] 陈宁,宋囡,贾连群,等.基于“脾-线粒体”相关理论谈痰瘀互结所致动脉粥样硬化的微观变化[J].中华中医药学刊,2018,36(4):849-851.
[9] 程岩岩,贾连群,宋囡,等.基于PCRarray技术探讨脾虚痰浊AS对心肌线粒体能量代谢通路相关基因mRNA表达的影响[J].辽宁中医杂志,2016,43(5):1069-1071,1078.
[10] 吕晓明,宋囡,王德山,等.脾气虚证与肾气虚证大鼠小肠组织cAMP-PKA信号转导通路变化的比较研究[J].北京中医药大学学报,2018,41(7):579-584.
[11] 陈宁,宋囡,贾连群,等.化瘀祛痰方对AS家兔肝脏线粒体呼吸链酶复合物的影响[J].中国实验方剂学杂志,2018,24(7):171-176.
[12] 王英,贾连群,宋囡,等.健脾化痰祛瘀方药对脾虚痰浊动脉粥样硬化巴马猪肝脏线粒体能量代谢基因表达的影响[J].辽宁中医杂志,2017,44(11):2427-2429.
[13] 尹妮,杨关林,姜钧文,等.巴马小型猪冠状动脉粥样硬化模型的评价方法[J]. 山东大学学报(医学版),2017,55(7):1-5+48.
[14] 杨茗茜.巴马小型猪脾虚痰浊模型建立与评价[D].沈阳:辽宁中医药大学,2014.
[15] Elahi MM, Kong YX, Matata BM. Oxidative stress and glucose metabolism disorders[J]. Current opinion in clinical nutrition and metabolic care, 2010, 13:439-446.
[16] Hulsmans M, Van dooren E, Holvoet P. Mitochondrial reactive oxygen species and risk of atherosclerosis[J]. Current atherosclerosis report, 2012, 2:259-269.
[17] 郑荣梁,黄中洋.自由基生物学[M].3版.北京:高等教育出版社,2007:187.
[18] Warner DS, Sheng H, Batinic-Haberle I. Oxidants, antioxidants and the ischemic brain[J]. J Exp Biol, 2004, 207(Pt 18): 3221-3231.
[19] Silva JP, Gomes AC and Coutinho OP. Oxidative DNA damage protection and repair by polyphenolic compounds in PC12 cells[J]. Eur J Pharmacol, 2008, 601(1-3): 50-60.
[20] 曹国君.健脾化痰法治疗痰湿中阻型冠心病心绞痛的临床研究[D].哈尔滨:黑龙江中医药大学,2010.
[21] 张晓丹,终欣,刘琳,等.党参及黄芪对实验性心肌缺血大鼠心电图影响的比较[J].中草药,2003,34(1):1018-1020.
[22] 李岩,农一兵,林谦.益气药对慢性心力衰竭心气虚证模型大鼠总肌酸激酶活性、肌酸激酶同工酶及腺苷酸转位酶mRNA表达的影响[J].中华中医药杂志,2011,26(5):1216-1221.
[23] 王志强,李炳超.半夏药理作用研究进展[J].山西医药志,2009,38(1):68-69.
[24] 龙全江,徐雪琴,胡昀.白术的化学、药理与炮制研究进展[J].中国中医药信息杂志,2004,11(11):1033-1034.
[25] 曲丹.复方桂枝茯苓丸对动脉粥样硬化大鼠ICAM1、VCAM1表达影响的实验研究[D].咸阳:陕西中医学院,2012.
[26] 王姗姗.瓜萎皮的药理作用及临床应用[J].西医药杂志,2009,38(1):67-68.
[2] 郭丛丛,黄力,董建军,等.冠状动脉粥样硬化性心脏病血瘀证与冠脉病变程度及血管内皮功能的相关性研究[J].上海中医药杂志,2018,52(9):15-18.
[3] 刘君花,欧阳恩鸿,贺志明,等.虎杖苷对动脉粥样硬化大鼠血脂的调节和CD36、VCAM-1表达的影响[J].邵阳学院学报(自然科学版),2018,15(4):99-104.
[4] 胡月,李潞,赵红丽.血管内皮功能与冠状动脉粥样硬化发生发展的研究进展[J].中西医结合心脑血管病杂志,2018,16(11):1525-1528.
[5] 许晓虹(Ruby).“心胆论治”针灸抗动脉粥样硬化斑块的临床与分子机制研究[D].广州:广州中医药大学,2017.
[6] 马艺鑫,张妮,贾连群,等.基于“痰浊血瘀”病机探讨自噬与动脉粥样硬化的关系[J].时珍国医国药,2016,27(9):2240-2242.
[7] 陈丝,杨关林,王群,等.基于“异病同治”探讨“从脾论治”高脂血症及动脉粥样硬化共同理论基础及临床应用[J].中华中医药学刊,2018,36(9):2200-2202.
[8] 陈宁,宋囡,贾连群,等.基于“脾-线粒体”相关理论谈痰瘀互结所致动脉粥样硬化的微观变化[J].中华中医药学刊,2018,36(4):849-851.
[9] 程岩岩,贾连群,宋囡,等.基于PCRarray技术探讨脾虚痰浊AS对心肌线粒体能量代谢通路相关基因mRNA表达的影响[J].辽宁中医杂志,2016,43(5):1069-1071,1078.
[10] 吕晓明,宋囡,王德山,等.脾气虚证与肾气虚证大鼠小肠组织cAMP-PKA信号转导通路变化的比较研究[J].北京中医药大学学报,2018,41(7):579-584.
[11] 陈宁,宋囡,贾连群,等.化瘀祛痰方对AS家兔肝脏线粒体呼吸链酶复合物的影响[J].中国实验方剂学杂志,2018,24(7):171-176.
[12] 王英,贾连群,宋囡,等.健脾化痰祛瘀方药对脾虚痰浊动脉粥样硬化巴马猪肝脏线粒体能量代谢基因表达的影响[J].辽宁中医杂志,2017,44(11):2427-2429.
[13] 尹妮,杨关林,姜钧文,等.巴马小型猪冠状动脉粥样硬化模型的评价方法[J]. 山东大学学报(医学版),2017,55(7):1-5+48.
[14] 杨茗茜.巴马小型猪脾虚痰浊模型建立与评价[D].沈阳:辽宁中医药大学,2014.
[15] Elahi MM, Kong YX, Matata BM. Oxidative stress and glucose metabolism disorders[J]. Current opinion in clinical nutrition and metabolic care, 2010, 13:439-446.
[16] Hulsmans M, Van dooren E, Holvoet P. Mitochondrial reactive oxygen species and risk of atherosclerosis[J]. Current atherosclerosis report, 2012, 2:259-269.
[17] 郑荣梁,黄中洋.自由基生物学[M].3版.北京:高等教育出版社,2007:187.
[18] Warner DS, Sheng H, Batinic-Haberle I. Oxidants, antioxidants and the ischemic brain[J]. J Exp Biol, 2004, 207(Pt 18): 3221-3231.
[19] Silva JP, Gomes AC and Coutinho OP. Oxidative DNA damage protection and repair by polyphenolic compounds in PC12 cells[J]. Eur J Pharmacol, 2008, 601(1-3): 50-60.
[20] 曹国君.健脾化痰法治疗痰湿中阻型冠心病心绞痛的临床研究[D].哈尔滨:黑龙江中医药大学,2010.
[21] 张晓丹,终欣,刘琳,等.党参及黄芪对实验性心肌缺血大鼠心电图影响的比较[J].中草药,2003,34(1):1018-1020.
[22] 李岩,农一兵,林谦.益气药对慢性心力衰竭心气虚证模型大鼠总肌酸激酶活性、肌酸激酶同工酶及腺苷酸转位酶mRNA表达的影响[J].中华中医药杂志,2011,26(5):1216-1221.
[23] 王志强,李炳超.半夏药理作用研究进展[J].山西医药志,2009,38(1):68-69.
[24] 龙全江,徐雪琴,胡昀.白术的化学、药理与炮制研究进展[J].中国中医药信息杂志,2004,11(11):1033-1034.
[25] 曲丹.复方桂枝茯苓丸对动脉粥样硬化大鼠ICAM1、VCAM1表达影响的实验研究[D].咸阳:陕西中医学院,2012.
[26] 王姗姗.瓜萎皮的药理作用及临床应用[J].西医药杂志,2009,38(1):67-68.