【作者】 唐昭；

【导师】 石刚刚；

【作者基本信息】 汕头大学 ， 药理学， 2005， 硕士

【摘要】 氟哌啶醇（Haloperidol,Hal）为抗精神病药。以往的一些研究表明Hal有扩张冠状动脉的作用。鉴于Hal有较严重的锥体外系不良反应,将Hal结构改造得到其季铵盐衍生物F₂（碘化 N-正丁基氟哌啶醇,N-n-butyl haloperidoliodide）,它不易通过血脑屏障,避免了中枢的不良反应,又保持了良好的心血管活性。本文拟从基因水平研究F₂对在体模型心肌缺血再灌注过程中重要相关功能基因转录及表达的影响及对炎性损伤的保护作用,进一步探讨F₂抗心肌缺血再灌注损伤的分子机制。一、F₂对大鼠心肌组织Egr-1表达及炎性反应的影响 大鼠称重,麻醉后固定,手术结扎大鼠冠状动脉左前降支造成心肌缺血60min后解除结扎再灌注180min。大鼠随机分成四组:i假手术组（sham组）、ii对照组（I/R组）、iii溶剂组（I/R+PEG）、iv给药组（F₂,2mg/kg）。实验结束后取结扎线下厚约1.5—2.0mm的心肌组织经10%多聚甲醛固定,梯度脱水后心肌组织经石蜡包埋,切片,HE染色,光镜下观察。同时取结扎线下缺血部分心肌组织-70℃保存备用,分别应用RT-PCR方法及Western-blot方法检测缺血更多还原

【Abstract】 Haloperidol (Hal) is an antipsychotic agent. Our previous research showed that Hal could noncompetitively antagonize the contraction of coronary artery induced by phencyclidine, agonists of opioid receptor, and noradrenaline, etc. However, it was not used as an anti-myocardial ischemia drug due to its extrapyramidal adverse reactions. In consideration of this, we synthesized a series of quaternary ammonium salt derivatives of Hal, with polarity increased so that they do not pass the blood-brain barrier. One of these compounds with serial number F2 (N-n-butyl haloperidol iodide) was screened, and was found to maintain the vasodilating effects but have no side reactions. In this article, To investigate the effect of F2 on Egr-1 mRNA and protein expression in the model of rats with myocardial ischemia-reperfusion and to perfect the molecular mechanism of protective effect of F2 on ischemia-repurfusion injury. The effect of F2on Egr-1 expression and myocardial inflammationSprague-Dawley rats were anaesthetized with ethyl carbamate(40%).MI was produced by temporarily exteriorizing the heart by means of placing a silk suture slipknot around the left anterior descending coronary artery (LAD). After 60 minof MI, the slipknot was released, and the myocardium was reperfused (R) for 3 h . The rats were randomly assigned to one of the following groups: (i) sham; (ii)MI/R (0.9% NaCl, control group); (iii) MI/R+vehicle (PEG400, 2ml/kg); (iv) MI/R+F2 (2mg/kg); After the 3 h reperfusion period, the ischemia portions of the myocardium under suture were then stored at -70°C for later assay of RT-PCR, Western-blot, myeloperoxidase activity (MPO) as described. Myocardial tissue under silk 1.5-2.0mm were cut and fixed in 10% formalin, hemotoxylineosin stained.The results of experiment were as follows:1. Expression of Egr-1 in the rat ischemia-reperfusion modelWe first analyzed Egr-1 mRNA expression in myocardial tissues of each group by RT-PCR. We detected high levels of Egr-1 mRNA in control and PEG groups, compared with very low levels of Egr-1 mRNA detected in sham and F2 groups. We found PEG did not decrease Egr-1 expression measurably but F2 inhibit level of Egr-1 mRNA based on densitometric analysis. We observed a increase markedly in Egr-1 protein along according with levels of Egr-1 mRNA in each groups. Expression of Egr-1 protein was increased in control and PEG groups, based on densitometric analysis of multiple blots.2. Cardiac MPO ActivitiesThe mean values of MPO activity obtained in the area not at risk of left ventricle wall were low in the sham group. However, these values were increased remarkedly in the area at risk of vehicle-treated animals subjected to MI/R. Treatment of the rats with F2 caused inhibition of MPO activity in the area at riskof these hearts. 3. Pathological sectionMany neutrophils adhered to endothelium of micrangium and migrated into myocardial tissue in portion of ischemia-reperfused myocardium. And pathological characteristic of exudation? hemorrhage > dropsy were observed. However neutrophils counts adhereing or migrating to ischemia-reperfused myocardium were decreased markedly in F2 group compared with I/R and PEG groups.The effect of F2 on hemodynamics and myocardial enzymes Sprague-Dawley rats were anaesthetized with sodium pentobarbital (30 mg/kg, ip.) and were ventilated with a small animal respirator. The arterial blood pressure was measured via a polyethylene catheter cannulated to the right femoral artery, and the left ventricular pressure was measured via a polyethylene catheter, which was inserted into the left ventricular cavity through the right carotid artery. MI was produced by temporarily exteriorizing the heart by means of a left thoracic incision and placing a 6-0 silk suture slipknot around the left anterior descending coronary artery (LAD), 2-3 mm from its origin. After 60 min of MI, the slipknot was released, and the myocardium was reperfused (R) for 3 h. After the 3 h reperfusion period, 2 ml blood was taken from the carotid artery and centrifuged at 6000 rpm for 3 min. the plasma CK> CK-MB> LDH levels were analyzed by automatic biochemical apparatus.The results of experiment were as follows:1. Hemodynamics.-9-At the end of ischemia the HR^ SP > ± dp/dt max and LVSP values of the rats in the I/Rn PEG and F2 groups consistently decreased significantly contrasted with sham group. After 180 min reperflision, in I/R and PEG groups, the SP > ± dp/dt max and LVSP values fell gradually, which were significantly different from those measured in F2 and sham rats, indicating that, F2 could ameliorate the hemodynamics of ischemia-repefusion injured myocardium. 2. Analysis of serum CK> CK-MEk LDH concentration.The values of plasma CK> CK-MBk LDH concentration obtained in I/R and PEG rats increased intensely at the end of 180 min of reperflision compared with sham group. The administration of 2mg/kg F2 caused a marked inhibition in serum CK> CK-MB -> LDH concentration compared with I/R and PEG groups.The studies suggested that F2 may relieve inflammatory injury and protect myocardial function in rats with myocardial ischemia-reperfusion, the mechanisms may be correlation with attenuating the transcription and expression of Egr-1.更多还原