琥珀酸美托洛尔早期干预对心肌梗死兔心脏的保护作用及机制研究

发布时间：2018-05-16 02:18

本文选题：心肌梗死 + 美托洛尔　；参考：《南方医科大学》2017年硕士论文

【摘要】：背景:心血管病死亡占我国居民总死亡原因的首位,心肌梗死(MI)后并发的室颤等恶性心律失常是引起心脏性猝死(SCD)的主要原因。目的:探讨琥珀酸美托洛尔早期干预对兔心肌梗死40天后心脏组织缝隙连接重构以及细胞凋亡的影响。方法:将24只成年雄性新西兰大白兔随机分成假手术组(SH组,n=6)、心肌梗死组(MI组,n=6)、心梗后早期干预组(EBMI组,n=6)和心梗后常规干预组(RBMI组,n=6),通过结扎冠状动脉前降支建立心肌梗死模型,假手术组不结扎血管,余操作均相同。EBMI组和RBMI组分别于麻醉清醒后和心梗24小时后以琥珀酸美托洛尔(12.5mg/kg,溶于2ml蒸馏水)灌胃,SH组和MI组以等量蒸馏水灌胃。术后6小时,以生物化学法检测血清心肌坏死标志物(血清乳酸脱氢酶和肌酸激酶)水平。术后40天开胸暴露心脏,通过双极起搏电极测定心室颤动阈值;以免疫荧光法观察缝隙连接蛋白43(Cx43)及其磷酸化(p-Cx43)和缝隙连接蛋白45(Cx45)在心室肌组织的分布情况,TUNEL法测定心肌细胞凋亡率,Western blot法检测心室肌组织Cx43、p-Cx43、Cx45和Bcl-2、Bax、Caspase-3蛋白表达水平;实时荧光定量PCR法测定心肌组织中Cx43及Cx45的mRNA表达水平。结果:1、心肌梗死组的血清乳酸脱氢酶和肌酸激酶水平均高于假手术组(P均0.01);2、MI组的心室颤动阈值低于SH组(P0.05),EBMI组和RBMI组心室颤动阈值均高于MI组(P均0.05)。3、缝隙连接蛋白在心室肌的分布情况:SH组、EBMI组和RBMI组可见CX43主要位于闰盘内,与细胞长轴垂直,呈线状排列,而侧向分布的Cx43相对较少,MI组则相反;p-Cx43和Cx45的分布表现与Cx43相似。4、心肌组织中缝隙连接蛋白的表达:(1)蛋白表达水平:MI组p-Cx43和Cx45表达水平以及p-Cx43/Cx43比例均低于SH组(P均0.05);EBMI组和RBMI组p-Cx43和Cx45表达水平以及p-Cx43/Cx43比例均高于MI组(P均0.05),且EBMI组高于RBMI组(P0.05);各组CX43总蛋白的灰度分析无统计学差异(P0.05)。(2)mRNA表达水平:MI组Cx43和Cx45的mRNA表达水平低于SH组(P均0.05);EBMI组和RBMI组Cx43和Cx45的mRNA表达水平均高于MI组(P均0.05),且EBMI组高于RBMI组(P0.05)。5、凋亡相关检测:(1)TUNEL法:MI组心肌细胞凋亡率明显高于SH组(P0.05),EBMI组和RBMI组的心肌细胞凋亡率低于MI组(P均0.05),且EBMI组低于RBMI组(P0.05);(2)凋亡相关蛋白表达水平:MI组的Bcl-2水平及Bcl-2/Bax比值低于SH组(P均0.05),EBMI组和RBMI组的Bcl-2水平及Bcl-2/Bax比值高于MI组(P均0.05),且EBMI组高于RBMI组(P0.05);MI 组的 Bax 和 Caspase-3 水平高于 SH 组(P 均0.05),而 EBMI组和RBMI组的Bax和Caspase-3水平低于MI组(P均0.05),且EBMI组低于 RBMI 组(P0.05)。结论:琥珀酸美托洛尔通过改善心梗后心室肌缝隙连接蛋白的空间分布,减少缝隙连接蛋白的降解及去磷酸化,降低心梗后恶性心律失常的易感性,抑制心肌细胞凋亡,从而对心肌梗死兔心脏起保护作用,并且早期给药获益更明显。
[Abstract]:Background: cardiovascular disease is the leading cause of death in China. Malignant arrhythmias such as ventricular fibrillation after myocardial infarction (MI) are the main causes of sudden cardiac death (SCD). Aim: to investigate the effects of early intervention of metoprolol succinate on gap junction remodeling and apoptosis of heart tissue after 40 days of myocardial infarction in rabbits. Methods: Twenty-four adult male New Zealand white rabbits were randomly divided into sham operation group (SH group), myocardial infarction group (MI group), early intervention group (EBMI group) and post myocardial infarction group (RBMI group). Myocardial infarction model, The rest of the procedures were the same in sham operation group. EBMI group and RBMI group were perfused intragastrically with metoprolol succinate 12.5 mg / kg, dissolved in 2ml distilled water, respectively after awake anesthesia and 24 hours after myocardial infarction. The serum levels of lactate dehydrogenase and creatine kinase were measured by biochemical method at 6 hours after operation. The ventricular fibrillation threshold was measured by bipolar pacing electrode 40 days after operation. The distribution of gap junction protein 43 (Cx43) and its phosphorylated protein (p-Cx43) and gap junction protein 45 (Cx45) in ventricular myocytes were observed by immunofluorescence method. Tunel method was used to determine the apoptosis rate of cardiac myocytes. Western blot was used to detect the expression of Cx43 + Cx43 and Bcl-2BaxCaspase-3 in ventricular myocytes. The expression of Cx43 and Cx45 in myocardial tissue was measured by real-time fluorescence quantitative PCR. Results the serum lactate dehydrogenase and creatine kinase levels in the myocardial infarction group were higher than those in the sham operation group (P < 0.01). The ventricular fibrillation threshold in the myocardial infarction group was lower than that in the SH group (P 0.05) and the ventricular fibrillation threshold in the RBMI group was higher than that in the MI group (P < 0.05). In the distribution of ventricular muscle, CX43 was mainly located in intercalated disc in RBMI and EBMI group. Perpendicular to the long axis of the cell. On the contrary, the distribution of p-Cx43 and Cx45 was similar to that of Cx43 in the laterally distributed Cx43 group. The expression level of gap junction protein: 1) in myocardial tissue was lower than that in SH group (P < 0.01). The expression level of p-Cx43 and Cx45 and p-Cx43/Cx43 ratio were lower than those in SH group (P The expression level of p-Cx43 and Cx45 and the ratio of p-Cx43/Cx43 in both groups were higher than those in MI group (P 0.05) and EBMI group (P < 0.05), and the expression of CX43 total protein in EBMI group was higher than that in RBMI group (P < 0.05), and there was no significant difference in the gray level of CX43 total protein expression between the two groups. The mRNA expression levels of Cx43 and Cx45 in the group of RBMI were lower than those in the group of SH-SH. The mRNA expression levels of Cx43 and Cx45 in both groups were higher than those in MI group and RBMI group, and that in EBMI group was higher than that in RBMI group (P 0.05. 5). The apoptosis-related assays showed that the apoptotic rate was significantly higher in group 1: 1 Tunel than that in group SH (P 0.05) and group RBMI (P < 0.05). The apoptotic rate was significantly lower in EBMI group than that in SH group (P 0.05) and RBMI group (P < 0.05). Apoptosis-related protein expression level in EBMI group was lower than that in RBMI group (P 0.05) Bcl-2 level and Bcl-2/Bax ratio were lower in EBMI group than in SH group (P < 0.05). Bcl-2 level and Bcl-2/Bax ratio in RBMI group and RBMI group were higher than those in MI group (P 0.05), and EBMI group was higher than that in RBMI group (P 0.05). The levels of Bax and Caspase-3 were higher than those of SH group (P 0.05), while the levels of Bax and Caspase-3 in EBMI group and RBMI group were lower than those in MI group (P 0.05), and EBMI group was lower than RBMI group (P 0.05). Conclusion: metoprolol succinate can reduce the degradation and dephosphorylation of gap junction protein, decrease the susceptibility to malignant arrhythmia and inhibit cardiomyocyte apoptosis by improving the spatial distribution of gap junction protein in ventricular muscle after myocardial infarction. It can protect the heart of myocardial infarction rabbits, and the benefit of early administration is more obvious.
【学位授予单位】：南方医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R542.22

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