【作者】 刘继烈；

【导师】 阎文泰；

【作者基本信息】 郑州大学 ， 心血管内科， 2004， 硕士

【摘要】 硝酸甘油（Nitroglycerin，NTG）及其它硝酸酯类药物目前仍是治疗冠心病的经典药物，但由于其持续应用48～72小时后NTG耐药性的快速形成是限制其临床效应的主要因素。NTG的耐药性是多因素、多环节相互作用的复杂结果。近十余年的研究认为NTG持续应用时在血管内皮、神经内分泌及氧自由基之间导致了复杂的相互作用。NTG耐药时，血管组织内超氧化阴离子（superoxide anion，O₂^-）生成增多，氧化应激状态加剧，一氧化氮（nitric oxide，NO）灭活增多；一氧化氮合酶（nitric oxide synthase，NOS）功能障碍，使NO／O₂^-平衡失调，NO合成减少，O₂^-净生成增多；还有研究表明NTG耐药时血管组织中内皮素-1（endthelin-1，ET-1）生成增多，使血管组织对N／G的舒张反应减弱。以往预防NTG耐药性的措施都是从某单一环节去改善其耐药性，临床效应较差。目前研究发现3-羟基-3-甲基戊二酰辅酶A还原酶抑制剂（3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors，HCRIs）不但具有降脂作用，而且还具有改善血管内皮功能的作用，它可通过抑制NADPH膜氧化酶的激活，减少血管组织内O₂^-的生成，改善氧化应激状态；通过对geranylgeranyl磷酸酶（geranylgeranylpyrophosphate，GGPP）的抑制，上调NOS的表达，提高NOS的活性，使NO生成增多；还可通过抑制ET-1前体物质及其受体mRNA的表达，减少ET-1的合成，并降低其免疫学活性，从理论上可能更好的改善NTG的耐药性。本文将阿托伐他汀（Atorvastatin）与NTG合用，观察阿托伐他汀对NTG耐药性的影响，并对其作用机制进行初步探讨。郑州大学2004年硕士学位论文方法 实验选用64只健康雄性阿托伐他汀对硝酸甘油耐药性的影响及作用机制探讨J么嘟几护姿厂夕位沪矛Fj^沪SD大鼠，体重300一3509。随机分成4组，每组16只，其中8只用于动脉环实验和检测血管组织ET一1含量，另8只用于检测血管组织SOD活性、MDA含量、N0含量、NOS活性及血浆ET一含量。I组:对照组;H组:NTG耐药组;m组:阿托伐他汀（Atorvastat加）组;W组:阿托伐他汀+NTG组。I组和H组给予2一3ml的生理盐水灌胃，每天一次，共4周，最后3天分别连续给予不含有、含有NTG的贴膜（0.05mg·h一‘）贴敷;111组给以含阿托伐他汀（smg·kg一‘·d一‘）的生理盐水2一3ml灌胃，每天一次，共4周，最后3天给以不含NTG的贴膜;IV组Atorvastatin用法同111组，最后3天连续给以NTG贴膜（0.Osmg·h一‘）贴敷。动物模型制备成功后将动物处死，开胸取出胸主动脉，剪成3一Slnln的血管环，悬挂于3ml的恒温浴槽中，标本与张力换能器相连，连续通入95%压+5%C02的混合气体，平衡45一60分钟，加入10--6mol·L一L的去甲肾上腺素预收缩，（10刁一10一‘mol当标本收缩达最大值曲线平稳后加入浓度递增的NTGL一’）绘出浓度一效应曲线，观察血管对NTG的舒张效应，测出最大舒张反应百分率，测出舒张50%时NTG的浓度值。统计学处理:所有数据均采用SPSS 10.0工具软件处理。一结果1.动脉环的最大舒张反应:H组与I组相比明显减弱（尸<0.01）组相比明显增强（尸<0 .01），W组与I组相比差异也有显著性（尸<0.01）I组相比差异无统计学意义（尸> .05）。W组与1工，111组与 2.舒张50%时的NTG的浓度值:n组与I组相比明显增大（尸<0.01），IV组与11组相比明显减小（尸<0.01），W组与I组相比差异也有显著性（P<0.01），111组与I组相比差异无统计学意义（尸> .05）。3.浓度一效应曲线:n组与I组相比明显右移，m组与I组相比浓度一效应曲线无明显偏移，W组与I右移。W组与H组相比明显左移，、11工组相比浓度一效应曲线4.血管组织SOD活性、MDA含量:11组与I组相比SOD活性下降（尸<0.01）MDA含量升高（P<0.01），而IV组SOD活性又高于11组（P<0.01），MDAH组石P（0.01），IV组与工组相比SOD活性无差别（尸>.05），而MDA含量低于含量差别郑州大学2004年硕士学位论文阿托伐他汀对硝酸甘油耐药性的影响及作用机制探讨有统计学意义（尸<0.01），111组与I组、W组相比SOD活性、MDA含量虽有差别，但差异无统计学意义（尸）0.05)。 5.血管组织N0含量、NOS活性:11组血管壁组织内N0含量与I组相比显著升高（P<0.0一），而NOS活性与I组相比活性降低(P（0.01），IV组与I组、11组相比血管组织N0含量、NOS活性显著升高（尸<0 .01），111组血管壁组织内NO含量、NOS活性与I组相比无差异（尸> .05），111组血管壁组织内N0含量、NOS活性与W组相比差别有统计学意义（尸<0 .01）。 6.血浆及血管组织ET一1含量:H组血管壁组织内ET一1浓度及血浆内ET一1浓度与I组、m组和IV组相比明显升高（尸<0 .01），而I组、W组间相比差异有统计学意义（尸<0.01），I组、111组间相比差异无统计学意义（尸> .05）。结论 1.大鼠在NTG贴膜治疗3天后可以建立NTG耐药模型。 2.阿托伐他汀通过减少血管壁组织内盯的生成，减轻NTG耐药时血管壁组织的氧化应激状态。 3.阿托伐他汀通过提高NOS的酶活性，可改善NTG治疗时NOS功能的不相匹配，从而改善NTG耐药时血管的舒张反应。 4.阿托伐他汀通过减少血浆及血管壁组织内ET一1的生成，恢复血管组织?更多还原

【Abstract】 Nitroglycerin (NTG) and other nitrovasodilators have been widely used to treat ischemic heart disease, but its long-term hemodynamic and anti-ischemic efficacy of NTG is rapidly attenuated by the development of tolerance. The mechanisms involved hi the tolerance of NTG are not completely understood yet and probably are multifactorial and multilink. In the past decade, studies have demonstrated that nitroglycerin therapy leads to complex interactions among the vasculature, neurohormones and free oxygen radicals.. There is evidence that nitroglycerin tolerance is associated with an increased bioavailability of superoxide anion (Qj") and the nitric oxide synthase (NOS) uncoupling. The increased 02" takes a bad turn of vascular oxidative stress and reduces the bioavailability of nitroglycerin-derived NO. The NOS uncoupling makes the balance of NO/O2~ to the right, resulting in the net generation of 02". There is also evidence that long-term nitroglycerin therapy leads to the increased production of endthelin-l(ET-l) hi vasculature, decreasing the effects of nitroglycerin because of these changes in the balance of vasomotor tone. Those preventive measures of tolerance can partially restore nitroglycerin response by affecting a piece of mechanisms in the past, however, clinical efficacy is worse. Studies manifested that 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reduc-tase inhibitors (HCRIs) can not only lower the lipid load of the vessel wall, but also ameliorate vascular endothelial dysfunction. Several explanatory theories that HCRIs leads to increased NO have prevailed, including decreased ability to product 02" and improved oxidative stress in vessel wall, improved eNOS expression by inhibiting the activity of geranylgeranylpyrophosphate, and increased activity of NOS. HCRIs canalso inhibite the expression of pre-pro ET-lmRNA and reduce the levels of immunoreactive ET-1. Abstractly, these agents have a better function for nitroglycerin tolerance. This study was designed to assess the effects of atorvastatin on the development of tolerance during continuous transdermal nitroglycerin therapy and to investigate the mechanism of its action. Method64 health male Sprague-Dawley rats, weighing from 300 to 350 grams, were randomly divided into four groups: group I , group II, groupIII and group IV, 16 rats in every group. In every group, 8 rats were used for vascular rings experiment and measured the contents of ET-1 in vasculature, another 8 were only used for measuring the activity of SOD, the contents of MDA, the contents of NO, the activity of NOS in vessel organization and blood plasma ET-1 contains. The rat models of nitroglycerin tolerance were established by nitroglycerin patches. The animals were handled as follows: (1) group I and group II were administered the same dosage of physiological saline 2~3ml, not or nitroglycerin patches were administered in the last 3 days;(2) groupIII: atorvastatin (5mg-kg-1-d-1) was performed for 4 consecutive weeks, not nitroglycerin patches were administered in the last 3 days; (3) group IV: the administration of atorvastatin was the same as that of groupIII, nitroglycerin patches were administered in the last 3 days. The chest aorta were took out after the animals were, sentenced to death, and parts of the aortas were cut into vascular rings, suspended in a constant temperature organ chamber. The specimens were connected to a force transducer, the organ chamber was aerated with a mixture of 95% O2+5% CO2, equilibrium 45-60 minutes, acceded to noradrenalin (10-6mol-L"1). When the contractions had reached a stable plateau, relaxation responses to nitroglycerin(10"9~ 10-4mol-L-1) were obtained, drawing the density- effect curve, testing the density value of 50% of its own maximal response. The data were processed with SPSS10.0. Results1. The maximal relaxation responses in group II were significantly weakener than that in group I (P<0.01). The maximal relaxation responses in groupIV were significantly stronger than that in group II (P<0.01). There was significant diffe更多还原