【作者】 张舵舵；

【导师】 高振平；

【作者基本信息】 吉林大学 ， 生理学， 2010， 硕士

【摘要】 目的:本文以体表心电图为指标,观察心梗模型大鼠心律失常发生率,运用BL-420E+记录仪观察LPA对心梗模型大鼠离体乳头肌收缩力的影响,应用膜片钳技术观察LPA对心梗模型大鼠心室肌细胞L-Ca2+电流的影响,应用Western Blot检测心梗模型大鼠心肌Cx43蛋白的表达,研究LPA对心梗模型大鼠心肌的作用,探讨溶血磷脂酸在心梗并发心律失常的作用和机理。方法:采用冠状动脉结扎方法制备大鼠心梗模型,观察心律失常发生率;制备大鼠离体乳头肌标本,观察心梗大鼠心肌收缩力;采用Langendorff离体心脏灌流法分离单个心室肌细胞,应用全细胞膜片钳技术,观察不同浓度的LPA对心梗大鼠心室肌细胞L-Ca2+电流的影响;应用Western Blot检测Cx43蛋白的表达,观察不同浓度的LPA对心室肌细胞缝隙连接蛋白43表达的影响。结果:我们采用结扎大鼠左冠状动脉前降支的方法发现急性心肌梗塞模型大鼠心率明显增快,并有70%的实验动物出现室性期前收缩,证明心梗模型大鼠可发生心律失常。本实验观察到LPA可使心梗大鼠心肌收缩力增强,应用膜片钳实验技术从离子通道水平进一步证明其正性肌力作用的机理,是通过激活L型钙通道,增加钙通道电流实现的。我们对急性心梗模型大鼠的心脏,应用免疫印迹杂交(Western blot)方法观察心肌Cx43蛋白的表达情况,发现与空白对照组比较,心梗模型组和心梗模型+LPA组Cx43蛋白表达明显减少,而且,心梗模型+LPA组较心梗模型组减少更明显。心梗模型组和心梗模型+LPA组Cx43蛋白表达强度与空白对照组比差异均具有显著性。而LPA特异性阻断剂Ki16425+心梗模型组Cx43蛋白表达较心梗模型组明显增强,表达增强几乎接近空白对照组。结论:1..心梗模型大鼠可发生心律失常。2.溶血磷脂酸可明显增强心梗大鼠心肌收缩力。3.溶血磷脂酸能明显增强心梗大鼠心室肌细胞L型钙通道电流的幅值,且具有剂量依赖性。4.溶血磷脂酸可使心梗大鼠心室肌Cx43蛋白表达明显减少,参与心律失常的发生。5.溶血磷脂酸使心梗大鼠Cx43蛋白表达减少这一作用,可被其特异性阻断剂Ki16425所阻断,进一步证明溶血磷脂酸可通过影响Cx43蛋白的表达而引发心律失常。创新点:1.观察到溶血磷脂酸可降低心梗模型大鼠心室肌细胞Cx43蛋白的表达。2 .观察到溶血磷脂酸增加心梗模型大鼠心室肌细胞L-Ca2+电流的幅值,并呈剂量依赖性。更多还原

【Abstract】 Lysophosphatidic acid (LPA) is an intermediate product of membranous phospholipid contents. It can also be released from the activated platelets. During the acute myocardial infarction, the increase of phospholipid enzymatic activity facilitates the produce of lysophospholipids acid, which means that LPA may be related with arrhythmia occurrence. Connexin 43(Cx43) is the predominant protein in gap junction and the main gap junction protein of the ventricular myocytes. The low-resistance electrical couplings comprising with Cx43 make the ventricular myocytes contracting synchronously, which can be modulated through ion exchanges. If Cx43 reduced by 50%, the ventricular conduction velocity decrease obviously, resulting in arrhythmia.In this paper, through ligating the left lower branch of the rat left coronary artery, we find the heart rate of acute myocardial infarction of a rat model increases significantly. Seventy percent of experimental animal shows premature ventricualr contraction, which means that arrhythmia can take place in the rats with myocardial infarction. We observe through experiments that LPA can enhance the contraction of rats with myocardial infarction. Using the patch clamp recording technique, it is proved the positive inotropic mechanism from the ion channel level, which take functions through activating calcium channel L and increasing calcium channel currents, resulting in the occurrence of arrhythmia.The cardiac function is based on the law of myocardial cell signaling, which has been proved to be mediated by gap junction(GJ). Using the western blot for measuring the expression of Cx43 in the rats with acute myocardial infarction, we find that the Cx43 expression of ventricular myocytes of the rat with myocardial infarction and LPA decreased more than that of the normal rats (p<0.05). The Cx43 expression of LPA specificity blocker Ki16425 and AMI groups are enhanced obviously compared with the myocardial infarction group, which is almost same as the normal group. It is not significant for the group LPA and the normal group(p>0.05), while it is significant compared with the myocardial infarction group(p<0.05). Takingβ-actin as the parameter, every group show no significant difference(p>0.05). LPA specificity blocker can reduce the expression of Cx43. The expression of the group AMI+ Ki16425 is improved compared with the myocardial infarction group, which prove LPA can reduce the expression of Cx43. It is an important way of LPA inducing arrhythmia.The above results suggest that:(1) LPA involved in the occurrence of arrhythmia after acute myocardial infarction.(2)LPA can induce the open of the intracellular calcium ion channel, which result in the intracellular calcium overload and the generation of EADs and DADs. All of these factors can induce arrhythmia.(3)LPA can reduce the expression of Cx43 in rats with myocardial infarction, which can increase the occurrence of arrhythmia.更多还原