【作者】 李明；

【导师】 鲁卫星；

【作者基本信息】 北京中医药大学 ， 中西医结合内科学， 2009， 博士

【摘要】 研究背景及目的:随着社会的发展,冠心病已经成为成年人死亡的重要原因,在经济和工业化发达的国家,它在死亡原因中位居前列。我国的冠心病发病率和死亡率虽然远较这些国家为低,但是在大部分地区,特别是在北方和经济发达地区,都发现冠心病的发病率呈逐年上升趋势。因此防治冠心病成为各国医药学界重要研究课题。心肌缺血再灌注损伤(Myocardium ischemic reperfusion injury,MIRI)是临床常见的病理现象,是冠心病心肌梗死的病理生理学基础。粘附分子与再灌注损伤有着密切关系,各种粘附分子在心肌细胞的缺血/再灌注损伤作用已经愈来愈受到重视。其中P选择素、CD40/CD40L是近年来研究的热点。作为世界医药的一个重要组成部分,利用天然植物资源治疗疾病,研制有效的防治冠心病的新药物是目前中医药研究的重要方向。冠心病属中医学“胸痹”、“心痛”范畴。其病机以本虚标实,虚实错杂为主,实则寒凝、气滞、血瘀、痰浊;虚则气血阴阳亏虚,而心气虚则为发病之根本、病机之关键。心气亏虚,无以行血,则血脉瘀滞,气虚血瘀为冠心病最常见病机。本课题针对这一病机,研究具有益气活血功效的龙牙枢木(Aralia elata)对心肌细胞的缺血/再灌注损伤的作用。龙牙楤木其主要有效成分龙牙楤木总苷(aralosides)具有抗炎、抗自由基损伤和抗心肌缺血作用。本研究通过建立心肌缺血再灌注模型,模拟急性心肌IRI,探讨龙牙楤木总苷及普奈洛尔对心肌缺血再灌注过程中粘附分子表达的影响。方法与结果:第一部分:观察龙牙楤木总苷对心肌缺血再灌注损伤P选择素表达的影响。方法:龙牙楤木总苷等药物预处理,然后可逆性冠脉左前降支结扎缺血45min再灌注5分钟复制心肌缺血再灌注(MI/R)模型,HE染色观察心肌病理学变化,免疫组化法测量心内膜P选择素的表达。结果表明龙牙楤木组与模型组比较无显著差异。波立维组面密度、积分光密度(IOD)值与模型组比较有统计学显著性差异。第二部分:观察龙牙楤木总苷对心肌缺血再灌注损伤ICAM-1表达及MDA、GSH-PX的影响。方法:龙牙楤木总苷等药物预处理,然后可逆性冠脉左前降支结扎缺血45min再灌注6h复制MI/R模型,HE染色观察心肌病理学变化,检测血清LDH含量,测定心肌MDA、GSH-PX活性,,ELIsA法测定血清sVCAM-1的表达。结果表明:与模型组比较,龙牙楤木组、波立维组血清LDH含量显著降低(P＜0.05),龙牙楤木组、波立维组SOD活性显著升高(P＜0.05),龙牙楤木组、波立维组与模型组ICAM-1表达没有统计学差异(P＞0.05)。第三部分:本研究旨在研究龙牙楤木总苷对急性心肌梗死引发的室性心律失常及生存率的改善作用。方法:龙牙楤木总苷等药物预处理,然后冠脉左前降支结扎复制心梗模型,计算心律失常评分、室颤发生率、死亡率、除颤成功率。结果:心律失常评分龙牙楤木组和普萘洛尔组与模型组比较无统计学差异(P＞0.05),合心爽组与模型组比较有显著统计学差异(P＜0.05);室颤发生率龙牙楤木组和普萘洛尔组与模型组比较无统计学差异(P＞0.05),合心爽组与模型组比较有显著统计学差异(P＜0.05);死亡率龙牙楤木组和合心爽组与模型组比较有显著统计学差异(P＜0.05)。除颤成功率龙牙楤木组与模型组比较有显著统计学差异(P＜0.05)。第四部分:观察普奈洛尔对心肌缺血再灌注损伤CD40L表达的影响。方法:普奈洛尔等药物预处理,然后可逆性冠脉左前降支结扎缺血45min再灌注5分钟复制MI/R模型,HE染色观察心肌病理学变化,免疫组化法半定量测量心内膜CD40L的表达。结果表明普奈洛尔组面密度、积分光密度(IOD)值与模型组比较有统计学显著性差异。结论:1、龙牙楤木总苷保护心肌的作用与P选择素、ICAM-1表达无直接关系,其机制可能是通过增加GSH-PX的活力,减少MDA含量,对抗氧自由基对心肌细胞的毒害作用,减轻心肌细胞的损伤而实现的。龙牙楤木总苷的作用机制有待进一步研究。2、普奈洛尔可有效地减少缺血再灌注过程中心内膜CD40L的表达,提示普奈洛尔可以改善内皮细胞功能及再灌注微血管功能失调,是其药理作用机制的新发现。3、龙牙楤木总苷大大降低急性心梗大鼠的死亡率,改善大鼠急性心梗出现的恶性心率失常的预后,提高生存率。其作用机理可能与龙牙楤木抗心肌缺血、稳定心肌细胞电生理及升高室颤阈有关。更多还原

【Abstract】 Back Ground and Objective:In nowadays,coronary heart disease(CHD) was the important cause of adult’s death.It is in the first place of all kinds of dead causes in developed countries of economics and industrialization.The incidence rate and mortality of CHD are lower in our country than those countries,but with the development and changes of the society and economics,the incidence rate of CHD is in rising tendency year by year in most of regions,espicaliy in north and developed regions.So the prevention and cure of cardiovascular disease has been the important researchful subject of medicine in each country.MIRI are the common clinical pathology phenomenon,the pathophysiology foundation of the myocardial infarction in coronary heart disease.Adhesion molecule has closely related to MIRI.Variety of adhesion molecules have been more and more attetion in myocardial ischemic/reperfusion injury.And CD40 / C D40L P-selectin are the hot research points in recent year.As a important constitution of world-medicine,it is the main researchful direction of traditional Chinese medicine(TCM) of using wildings to treat disease and developing effective new drugs to prevent and cure chest pain,at present. CHD belongs to the"chest pain"or"heartache"category in TCM.Deficiency in origin and excess in superficiality and simultaneous occurrence of deficiency and excess syndromes is its main pathogenesis.Excess in superficiality are cold retained,qi stagnation,blood stasis and obstruction by phlegm.Deficiency in origin are deficiency of qi,blood,yin and yang.Deficiency of heart-qi is the essence of onset of disease and the key of pathogenesis.Qi can’t promot circulation of blood because of deficiency of heart-qi,which leads to blood vessels stasis.Blood stasis due to qi deficiency is the most common pathogenesis of CHD. This subject aims at this pathogenesis,uses the method of supplementing qi and activating blood to treat CHD,observes the effect of Aralosides on myocardium ischemic reperfusion injury.Aralosides possess the capabilities of anti-inflammatory、anti oxygen free radicals and anti myocardial ischemic.The study was performed by using myocardial I/R model to investigate the effects of Aralosides and Propranolol on expression of adhesion molecules in rats after myocardial ischemia/reperfusion injury.Methods and Results:PartⅠ:To investigate effects of Aralosides to the quantity of P-sel expression on endocardium during myocardial ischemia reperfusion.Methods:A model of myocardial ischemia 45min followed by 5min reperfusion was made by reversibly ligating coronary left descending artery.P-sel were observed.As a result, Aralosides has no significant effect of the quantity of P-sel expression.PartⅡ:To investigate the effects of Aralosides on expression of inflammatory molecule ICAM-1 and the activity of Malcic Dialdchyde(MDA) & Glutathione Peroxidase(GSH-PX) in rats after myocardial ischemia/reperfusion(MI/R) injury. Methods:A model of myocardial ischemia 45min followed by 6h reperfusion was made by reversibly ligating coronary left descending artery.30 Wistar rats were divided randomly into 3 groups:Ichemia reperfusion Group,Aralosides Goup, Plavix Group.HE staining was used to observe the pathological changes of myocardial issues.The activity of MDA and GSH-PX was measured,too.ELISA was used to determine ICAM-1 expression in myocardium.Results:In rats treated by Aralosides or Plavix,the lactate dehydrogenase LDH) activity was significantly decreased.Meamwhile,Aralosides and Plavix raised the activity of Malcic Dialdchyde(MDA) & Glutathione Peroxidase(GSH-PX).ICAM-1 expression was detected in myocardium after ischemia 45min followed by 6h reperfusion.In rats treated by Aralosides or Plavix,ICAM-1 expression remained unchanged.PartⅢ:To investigate the effects of Aralosides on Mortality of rat model with myocardial infarction Methods.Mcthods:43 Wistar rats were divided randomly into 4 groups:ischemia reperfusion group(n=11),Aralosidcs group(n=10), Propranolol group(n=12),Diltiazem group(n=10).The left anterior coronary descending artcry was ligated for 45 min.ECGs werc recorded incessantly during myocardial ischemia.Thc ECG was analysed and given score in Lambeth regulation.The occurrence rate of repcrfusion arrhythemia(VF) was analyscd.Rcsults:In rats treated by Aralosides,the occurrence rate of repcrfusion arrhythemia(VF) showd no difference.However,mortality was significantly decreased.PartⅣ:To investigate effects of Propranolol to the quantity of CD40Lexpression on endocardium during myocardial ischcmia reperfusion.Methods:A model of myocardial ischemia 45min followed by 5min reperfusion was made by reversibly ligating coronary left descending artery.As a result,Propranolol reduced the quantity of CD40L expression significantly.Conclusion:1.Aralosides could protect myocardium from inflammatory injury during MI/R. Aralosides has no significant effect of P-sel、ICAM-1.The mechanism is related to the enhancement of antioxidative effect on cells,and the reduction membrane damage caused by free radical and lipid peroxide.2.Propranolol reduced the quantity of CD40L expression significantly during MI/R.This was a new mechanism of their pharmacologic action and as a proof for them to be applied to ACS.3.Mortality of rat model with myocardial infarction could significantly decreased in rats treated by Aralosides.The mechanism may be related to that Aralosides could protect myocardium from injury during MI/R.更多还原