硝酸酯药物后处理对大鼠心肌缺血再灌注损伤保护作用的研究

发布时间：2018-04-04 13:02

本文选题：缺血后处理　切入点：药物后处理　出处：《青岛大学》2015年博士论文

【摘要】：目的:目前众多研究显示缺血后处理具有减轻心肌缺血再灌注损伤(MIRI),保护心肌作用,但其操作的有创性限制了临床应用,因此药物后处理的研究引起更多关注。既往研究显示一氧化氮(NO)参与了MIRI的发生发展,增加内源性NO生成及给予外源性NO均能减轻MIRI。本研究通过在大鼠MIRI模型中给予外源性NO供体硝酸酯药物后处理和缺血后处理对比,观察其是否对MIRI具有保护作用;另外通过检测特异性内质网应激损伤相关蛋白Grp78和Caspase 12的表达及其基因水平的变化,以期探讨内质网应激途径是否参与缺血后处理和硝酸酯药物后处理对MIRI的影响。方法:32只Wistar大鼠随机分为Sham组(假手术组)、I/R组(缺血再灌注组)、IPostC组(缺血后处理组)及PPost C组(硝酸酯后处理组)4组,每组8只。分别测定各组的相关指标:采用Evan’s blue与TTC双染法检测及使用Image Pro Ilus 6.0图像分析软件分析得出缺血和梗死心肌面积、ELISA方法检测血c Tn I水平,TUNEL法检测心肌细胞凋亡;荧光定量RT-PCR检测心肌组织Grp78、Caspase12基因表达;Western-blot检测心肌组织Grp78、Caspase12蛋白表达。结果:1.与I/R组比较IPostC组、PPostC组心肌缺血和梗死面积均明显减少(P0.01,P0.01)、血c Tn I水平降低(P0.01,P0.01),心肌细胞凋亡指数明显改善(P0.01,P0.01)。与IPost C组比较,PPost C组心肌缺血面积明显减少(P0.05),但两组间心肌梗死面积、血c Tn I水平和心肌细胞凋亡指数无明显改变(P值均0.05)。2.(1)心肌组织Grp78 m RNA表达及蛋白表达结果:与假手术组相比,I/R组、IPost C组和PPost C组均明显升高,P值均0.05;与I/R组相比,IPost C组和PPost C组均明显升高,P值均0.05;而IPost C组与PPost C组相比P0.05,差异无显著意义。(2)心肌组织Caspase12蛋白表达及m RNA表达结果:与假手术组相比,I/R组、IPost C组和PPost C组均明显升高,P值均0.05;与I/R组相比,IPost C组和PPost C组的均明显降低,P值均0.05;而IPost C组与PPost C组相比P0.05,差异无显著意义。结论:1.硝酸酯药物后处理与缺血后处理对心肌缺血再灌注损伤具有同样的保护作用,能够减少心肌梗死面积,降低血清c TNI水平,减轻细胞凋亡,并且较缺血后处理心肌缺血面积减少更显著。内质网应激途径导致的细胞凋亡参与了心肌缺血再灌注损伤的过程,缺血后处理和硝酸酯后处理均能通过调节内质网应激反应减少心肌细胞凋亡。
[Abstract]:Objective: at present, many studies have shown that post-ischemic treatment can reduce myocardial ischemia-reperfusion injury and protect myocardium, but its invasive operation limits the clinical application, so the study of drug post-treatment has attracted more attention.Previous studies have shown that nitric oxide (no) is involved in the pathogenesis and development of MIRI. Increasing endogenous no production and administration of exogenous no can reduce MIRI.In this study, we compared the effects of exogenous nitric oxide (no) donor nitrate after treatment with ischemic posttreatment on MIRI model in rats, and observed whether it had protective effect on MIRI.In addition, the expression of specific endoplasmic reticulum stress-related proteins (Grp78 and Caspase 12) and the changes of their gene levels were detected in order to investigate whether endoplasmic reticulum stress pathway is involved in ischemic post-treatment and nitrate post-treatment on MIRI.Methods Thirty-two Wistar rats were randomly divided into Sham group (sham-operation group), I / R group (ischemia reperfusion group) and PPost C group (nitrate-treated group, n = 8 in each group).The relative indexes of each group were detected by Evan's blue and TTC double staining and analyzed by Image Pro Ilus 6.0 image analysis software. The myocardial area of ischemia and infarction was detected by Elisa and Tunel method was used to detect apoptosis of cardiac myocytes.The expression of Caspase12 gene in myocardial tissue was detected by fluorescence quantitative RT-PCR and the expression of Caspase12 protein in myocardial tissue was detected by Western-blot.The result is 1: 1.Compared with the I / R group, the myocardial ischemia and infarct area in IPostC group were significantly decreased, while the level of serum c TnI was decreased in P0.01P0.01and P0.01P 0.01, and the myocardial apoptosis index was significantly improved.Compared with IPost C group, myocardial ischemia area decreased significantly in PPost C group, but myocardial infarction area between two groups was significantly decreased.Results of Grp78 m RNA expression and protein expression in myocardial tissue: compared with sham operation group, the expression of Grp78 m RNA and PPost C group were significantly higher than those in sham operation group, and were significantly higher than those in I / R group and PPost C group, respectively, and were significantly higher than those in I / R group and I / R group, respectively.The expression of Caspase12 protein and m RNA in myocardial tissue of IPost C group and PPost C group were significantly higher than those of PPost C group and PPost C group, and the expression of Caspase12 protein and m RNA in myocardial tissue of IPost C group were not significantly different from those of PPost C group. Compared with sham operation group, the expression of Caspase12 protein and m RNA in IPost C group were significantly higher than those in sham operation group, and that in PPost C group and I / R group were higher than those in sham operation group.Compared with I / R group, the P value of PPost C group and PPost C group decreased significantly (P 0.05), but there was no significant difference between IPost C group and PPost C group (P 0.05).Conclusion 1.After treatment with nitrate ester has the same protective effect on myocardial ischemia-reperfusion injury, which can reduce myocardial infarction size, reduce serum c TNI level and reduce apoptosis.The area of myocardial ischemia decreased more significantly than that of post-ischemic treatment.Apoptosis induced by endoplasmic reticulum stress pathway is involved in the process of myocardial ischemia-reperfusion injury. Both post-ischemic treatment and nitrate post-treatment can reduce myocardial apoptosis by regulating endoplasmic reticulum stress.
【学位授予单位】：青岛大学
【学位级别】：博士
【学位授予年份】：2015
【分类号】：R542.2

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