胃泌素对心脏缺血再灌注损伤的保护作用及机制研究

发布时间：2018-05-09 17:10

本文选题：胃泌素 + 心脏缺血再灌注损伤　；参考：《第三军医大学》2015年硕士论文

【摘要】：一、研究背景缺血性心脏病(ischemic heart disease,IHD)严重威胁人类健康安全,虽然医疗技术的发展使缺血的心肌尽早地实现了重建血运,拯救了许多缺血心肌,但同时带来了心肌细胞致命的损伤,造成梗死面积的增大,发生缺血再灌注(ischemia/reperfusion,I/R)损伤。当今世界已经步入缺血再灌注治疗的时代,如何减少缺血再灌注造成的心肌细胞损伤进一步加重,已成为基础研究者以及临床医生的研究热点。I/R损伤发生的机制具有复杂性,各机制间相互影响彼此促进。细胞凋亡和坏死是I/R损伤的重要的病理生理现象。研究发现减少心肌细胞凋亡可显著减少细胞死亡,可显著减少心肌梗死面积,因此减少心肌细胞凋亡对治疗I/R损伤具有重要意义。细胞凋亡是一种程序性死亡,受多种基因和凋亡信号严格调控,在细胞凋亡基因及信号激活的同时,机体的自我保护信号激活,如抑制凋亡的基因和促生存信号通路的激活(PI3K/Akt、ERK1/2、STAT3)。近年来对减少细胞凋亡的治疗策略上进行了大量的研究,取得了较为显著的成果,如缺血前预处理、缺血后处理以及远端缺血预处理,这些治疗措施中促生存途径或多或少参与其中,并有许多药物及治疗处理应用于临床,如依达拉奉,但这些成果应用于临床却受到安全性等很多因素的影响,目前仍感觉束手无策,因此急需要寻找出一些新的治疗措施。既往发现饱食可减少心脏缺血再灌注造成大鼠心肌的损伤,同时也发现Akt被激活。我们知道受进食影响最大的是胃肠道激素,其中胃泌素在进食后循环中的浓度升高高于其他胃肠道激素,此外胃泌素具有抗调亡、营养作用等生物活性,能激活机体的促生存途径,包含RISK通路(PI3K/Akt、ERK1/2)与SAFE通路(STAT3)。因此推测胃泌素可能在饱食对心脏缺血再灌注损伤的保护中发挥作用,这种保护作用可能激活了心脏的促生存途径(PI3K/Akt、ERK1/2、STAT3)而发挥的,以上问题的解决可能为我们寻找出一种新的治疗心脏缺血再灌注损伤的策略。二、研究目的明确胃泌素对心脏缺血再灌注损伤具有保护作用,并探讨胃泌素通过何种机制在心脏缺血再灌注损伤中发挥保护作用。三、研究内容1.第一部分我们对胃泌素在在体心脏I/R损伤中的保护作用进行研究,并对其机制进行初步探讨。将SD(Sprague Dawley)大鼠左前降支结扎,30min后剪开,制备在体的心脏I/R损伤模型,24小时后检测心肌(LDH、Tn T-HSST)改变,TTC染色观察心肌梗死面积,Tunel染色观察心肌细胞凋亡,同时采用Western Blotting方法检测凋亡蛋白Caspase-3、Akt、ERK1/2以及STAT3蛋白磷酸化表达水平,初步了解促生存途径在其中的作用。2.第二部分我们对胃泌素在离体心脏I/R损伤的保护作用进行研究,并对其作用机制进行进一步探讨。采用Langendorff建立离体心脏缺血再灌注损伤模型(心脏整体缺血40min,再灌注60min,在缺血前给予20min的药物干预),在该模型中去除了神经体液因素以及胃泌素对冠脉流量的影响。左心室球囊通过压力传感器和桥式放大仪持续记录心脏功能改变,收集缺血前以及再灌注60min时冠脉流出液检测心肌酶(LDH、Tn T-HSST)改变,TTC染色检测心肌梗死面积,TUNEL染色观察和心肌细胞凋亡情况,采用Western Blotting方法检测Caspase-3以及Akt、ERK1/2以及STAT3磷酸化水平,并应用Akt、ERK1/2以及STAT3拮抗剂分别阻断它们磷酸化改变,观察胃泌素预处理对心脏缺血再灌注损伤的影响。四、研究结果1.在在体缺血再灌注损伤中,胃泌素预处理后24小时心肌酶(LDH、Tn T-HSST)释放受到抑制,心梗面积缩小,心肌细胞凋亡下降,这些现象能被其受体拮抗剂CI 988所阻断。应用Western Blotting方法检测发现胃泌素预处理促进了Akt、ERK1/2以及STAT3蛋白磷酸化表达。2.在离体心脏缺血再灌注损伤中,胃泌素预处理可促进缺血后心功能的恢复(LVDP、LVEDP、+dp/dtmax),减少心肌梗死面积,抑制心肌酶(LDH、Tn T-HSST)释放以及减少心肌细胞的凋亡,这些作用能被其受体拮抗剂CI 988所阻断。此外应用Western Blotting方法检测发现胃泌素预处理促进了Akt、ERK1/2以及STAT3蛋白磷酸化表达,并应用Akt、ERK1/2以及STAT3拮抗剂分别阻断它们磷酸化后观察发现胃泌素对心脏I/R损伤的保护作用受到抑制,表现为心肌梗死面积增加,心肌酶释放增多。五、结论胃泌素预处理对心脏I/R损伤具有保护作用。该作用可通过胃泌素受体激活促生存途径即RISK信号通路(PI3K/Akt和ERK1/2)和SAFE信号通路(STAT3)来发挥。
[Abstract]:First, ischemic heart disease (IHD) is a serious threat to human health and safety. Although the development of medical technology makes the ischemic myocardium rebuild blood and save many ischemic myocardium, it also brings the fatal injury of myocardial cells, the increase of infarct size and the occurrence of ischemia reperfusion (ischem Ia/reperfusion, I/R) injury. The world has entered the era of ischemia reperfusion therapy. How to reduce the damage of myocardial cells caused by ischemia-reperfusion has become a research hotspot of basic researchers and clinicians. The mechanism of.I/R damage is complex, and the mechanisms of each mechanism interact to promote each other. And necrosis is an important pathophysiological phenomenon of I/R damage. It is found that reducing apoptosis can significantly reduce cell death and significantly reduce the area of myocardial infarction. Therefore, the reduction of myocardial apoptosis is of great significance in the treatment of I/R damage. At the same time that the apoptosis genes and signals are activated, the self protection signal of the body is activated, such as the inhibition of apoptosis gene and the activation of the survival signal pathway (PI3K/Akt, ERK1/2, STAT3). In recent years, a large number of studies have been carried out on the treatment strategies to reduce apoptosis, such as pre ischemic preconditioning and post ischemia. There are many drugs and treatment treatments used in clinical, such as edaravone, but these results are applied to a lot of factors such as safety, but they still feel helpless, so we need to find some new treatment urgently. Therapeutic measures. Previously found that full eating can reduce myocardial injury in rats with myocardial ischemia and reperfusion, and also found that Akt is activated. We know that gastrointestinal hormones are most affected by eating, and the concentration of gastrin in the circulation is higher than that of other gastrointestinal hormones, and gastrin has anti adjustment, nutrition, and so on. Activity, which activates the survival pathway of the body, including the RISK pathway (PI3K/Akt, ERK1/2) and the SAFE pathway (STAT3). Therefore, it is presumed that gastrin may play a role in the protection of cardiac ischemia reperfusion injury, which may activate the cardiac stimulating pathway (PI3K/Akt, ERK1/2, STAT3). The solution may find a new strategy for the treatment of ischemic reperfusion injury in the heart. Two. The purpose of this study is to clarify the protective effect of gastrin on cardiac ischemia reperfusion injury and to explore the mechanism of gastrin to protect the heart from ischemia reperfusion injury. Three. The first part of the study is to study the gastro secretion of the stomach. The protective effect of vegetarian on I/R injury in body heart was studied and its mechanism was preliminarily discussed. The left anterior descending branch of SD (Sprague Dawley) rats was ligated and 30min was cut open after 30min, and the I/R damage model of the heart was prepared. After 24 hours, the changes of myocardial (LDH, Tn T-HSST) were detected, the area of myocardial infarction was observed by TTC staining, and the myocardial cells were observed by Tunel staining. Apoptosis, and Western Blotting method was used to detect the phosphorylation of apoptotic protein Caspase-3, Akt, ERK1/2 and STAT3 protein, and the role of survival pathway in.2. second was preliminarily understood. The protective effect of gastrin on I/R injury in isolated heart was studied and the mechanism of its action was further explored. Ngendorff established an isolated myocardial ischemia reperfusion injury model (cardiac global ischemia 40min, reperfusion 60min, drug intervention for 20min before ischemia). In this model, the neurohumoral factors and the effect of gastrin on the coronary flow were removed. The left ventricular balloon was continuously recorded by pressure sensor and Bridge magnifying instrument. Changes of myocardial enzymes (LDH, Tn T-HSST) were detected before and before ischemia and reperfusion for 60min. TTC staining was used to detect myocardial infarction area, TUNEL staining and cardiomyocyte apoptosis. Western Blotting method was used to detect Caspase-3, Akt, ERK1/2, and STAT3 phosphorylation. The effects of gastrin preconditioning on cardiac ischemia reperfusion injury were observed respectively. Four. Results 1. in body ischemia reperfusion injury, the release of LDH (Tn T-HSST) was inhibited after 24 hours of gastrin preconditioning, the area of myocardial infarction was reduced, and the apoptosis of myocardial cells decreased. These phenomena could be antagonized by its receptor. CI 988 was blocked. Western Blotting method detected that gastrin pretreatment promoted Akt, ERK1/2, and STAT3 protein phosphorylation of.2. in ischemic reperfusion injury of isolated heart. Gastrin pretreatment could promote the recovery of cardiac function after ischemia (LVDP, LVEDP, +dp/ dtmax), decrease the area of myocardial infarction and inhibit myocardial enzyme SST) release and reduce the apoptosis of cardiac myocytes, which can be blocked by its receptor antagonist CI 988. In addition, the Western Blotting assay showed that gastrin pretreatment promoted the phosphorylation of Akt, ERK1/2 and STAT3 protein, and the gastric secreting was detected by Akt, ERK1/2 and STAT3 antagonists. The protective effect of peptide on cardiac I/R injury is inhibited, which shows an increase in infarct size and increased myocardial enzyme release. Five. Conclusion gastrin preconditioning has protective effect on cardiac I/R damage. This effect can be achieved through the activation of gastrin receptor (PI3K/Akt and ERK1/2) and SAFE signaling pathway (STAT3) through the activation of gastrin receptor.

【学位授予单位】：第三军医大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R541

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