【作者】 邹迪；

【导师】 隋殿军；

【作者基本信息】 长春中医药大学 ， 中医内科学， 2012， 博士

【摘要】 心肌缺血损伤是一种多因素参与的复杂病理过程，涉及到多个环节。近年来研究证明，在心肌缺血损伤的过程中炎性细胞因子大量释放并发挥重要的作用，心肌缺血是系统炎症反应和局部炎症反应共同参与的结果，炎症反应过度表达是心肌损伤的主要机制之一，而NF-κ B信号转导通路参与介导细胞增殖、炎症、凋亡等多种生理、病理过程，对于心肌缺血损伤的发生、发展具有关键作用。因此，本研究以三苦滴丸为观察药物，以拮抗炎症的过度表达在心肌中的损伤作用为研究的主要方向，以NF-κ B信号转导通路为切入点，从细胞及分子生物学水平揭示三苦滴丸抗实验性大鼠心肌缺血的作用及相关炎症机制，为三苦滴丸的临床疗效提供实验依据。目的：探讨三苦滴丸对实验性大鼠心肌缺血的保护作用及相关炎症机制。方法：健康Wistar大鼠随机分成空白组、模型组、三苦滴丸（SK）高剂量组、三苦滴丸中剂量组、三苦滴丸低剂量组、阳性对照药组（复方丹参滴丸），采用连续3天腹腔注射盐酸异丙肾上腺素,建立大鼠心肌缺血动物模型。观察心肌缺血模型大鼠服药后心电图改变及血清肌钙蛋白I、CK和CK-MB的含量；用光镜观察各组心肌的炎症改变；应用酶联免疫吸附试验检测服药后心肌缺血模型大鼠血浆中肿瘤坏死因子含量和白介素-1、白介素-6含量、血管细胞黏附因子1；应用蛋白印迹（Western-Blot）方法检测1κ B-α在心肌细胞胞浆中的表达，凝胶电泳迁移率（EMSA）变化检测NF-κ B活性，RT-PCR检测心肌NF-κ Bp65mRNA基因表达，免疫组化法检测心肌NF-κ Bp65蛋白表达，探讨三苦滴丸抗心肌缺血作用及相关机制。结果:1、三苦滴丸可改善心肌缺血大鼠的心电图S-T段、减少心肌缺血发生率、其中三苦滴丸低剂量组效果较好，其总体作用优于丹参滴丸组；三苦滴丸对肌钙蛋白、血清磷酸肌酸激酶（CK）、磷酸肌酸激酶同功酶(CK-MB)相关指标均有较好的改善效果，且呈量效依赖关系，其中尤以SK高剂量组作用显著，其总体作用优于丹参滴丸组。2、三苦滴丸能使心肌缺血大鼠模型受损心肌组织的显微结构明显得到改善，其中尤以三苦滴丸高、中剂量组作用显著，其总体作用优于丹参滴丸组。提示三苦滴丸可减轻心肌缺血组织损伤，可保护心肌细胞。3、三苦滴丸可抑制白介素-1、白介素-6、肿瘤坏死因子-α、血管细胞黏附因子-1等炎性因子，其中以三苦滴丸高、中剂量组显著，总体作用优于丹参滴丸组。4、三苦滴丸可上调1κ B-α在心肌细胞胞浆中的表达，降低NF-κ B的活性，抑制心肌NF-κ Bp65mRNA基因表达，抑制心肌NF-κ Bp65蛋白表达，其中以三苦滴丸高、中剂量组显著，总体作用优于丹参滴丸组。结论：三苦滴丸能有效的抗大鼠急性心肌缺血。其作用机制可能与三苦滴丸抑制NF-κ B信号转导通路上游激酶的活化，可上调1κ B-α在心肌细胞胞浆中的表达，从而阻止NF-κ B核移位，降低NF-κ B的活性，减少其转录调控的炎性因子的释放，如白介素-1、白介素-6、肿瘤坏死因子-α、血管细胞黏附因子-1等，抑制炎症反应，因此，三苦滴丸可通过多种综合作用机制起到抗心肌缺血损伤的作用。更多还原

【Abstract】 The injury of myocardial ischemia is a complex pathological mechanism which linked tomany factors. Recently research showed that the surge of inflammatory cytokines play animportant role in the process of myocardial ischemia, myocardial ischemia is the result ofboth system inflammatory reaction and local inflammatory reaction, and the over expressionof inflammatory reaction is one of the most important mechanisms of myocardial ischemia.As the key of pathology of myocardial ischemia, the signal transduction pathway of NF-kappaB participated in all kinds of physiology and pathology like cell proliferation, inflammation,apoptosis, and so on. Therefore, to study the mechanism of antagonism to myocardialischemia in experimental rats and provide the evidence of clinical efficacy for sankudiwan，this study used sankudiwan as the research medicine and focused on the antagonist ofinflammation over expression in myocardial injury, also the NF-kappa B signal transductionpathways.Purpose: This study sought to present the mechanism of sankudiwan in antagonism tomyocardial ischemia and related inflammatory reaction in experimental rats.Method:Using modern scientific technology to study Wistar rats,including pharmacy，pharmacology and phathology.The healthy Wistar rats were divited into normal control,negative control,danshendiwan positive control and sankudiwan high, medium,low dosagegroup. The myocardial ischemia rats model were made by intraperitoneal injection ofhydrochloric acid isoproterenol for three days. After the therapy of myocardial ischemiarats,we can observed the electrocardiogram change and the concentration of serum cardiactroponin I, CK, CK-MB, as well as the myocardial inflammation under electron microscop.Using the ELISA methord, we can observed the concentration of plasma TNF,IL1,IL6andvascular cell adhesion factor1.Also, we can detected the1κ B-alpha in myocardialcell,NF-kappa B activity, the NF-kappa B p65mRNA gene expression, the N-kappa B p65protein expression by Western-Blot, electrophoresis migration rate (EMSA), RT-PCR,immunohistochemical method,respectively.This study sought to present the mechanism ofsankudiwan in antagonism to myocardial ischemia. Result:1.Sankudiwan has obvious does-dependent effect on model ratsS-T segment deviation,incidence of myocardial ischemia，CK, CK-MB and CTN, especially the sankudiwan high,low group, and the effect is better than danshendiwan.2. Sankudiwan can obviously improve the micro-struture and ultra micro-structure onmodel rats,especialy the sankudiwan high and medium groups, and the effect is better thandanshendiwan group. Which indicated that sankudiwan can protect the cardiac muscleorganize.3．Sankudiwan may decrease the release of IL-1, IL-6,TNF-α, Vascular cell adhesionfactor-1, especially the SK high, medium group, and the effect is better than danshendiwa.4．sankudiwan may upregulate the expression of1κB-α in myocardial cell cytoplasm，decrease the activity of NF-κB, restrain the nf-kappa Bp65mRNA gene and the nf-kappaBp65protein expression, especially the SK high, medium group, and the effect is better thandanshendiwa.Conclusion: Sankudiwan has effective anti-myocadrial ischemia function.Themechanism relate to inhibiting the activity of NF-κB，upregulating the expression of1κB-αand then decrease the release of IL-1, IL-6,TNF-α,Vascular cell adhesion factor-1,which can protect myocardial cells，resistance of myocardial ischemia injury through a varietyof comprehensive mechanism.更多还原