布洛芬作用于酸敏感离子通道发挥脑保护作用的机制研究

发布时间：2018-03-24 21:26

  本文选题：缺血再灌注损伤　切入点：布洛芬　出处：《中国人民解放军医学院》2017年硕士论文

【摘要】：研究背景和目的:脑卒中是一种急性的脑血管疾病。因其具有高发病率、高复发率、高致残率和死亡率,严重影响着患者的生活质量。其中,缺血性脑卒中较出血性脑卒中具有更高的发病率,占脑卒中总数的60%-70%。缺血性脑卒中的发病机制和病理生理过程极其复杂,因此给临床治疗带来了极大的困难。酸中毒是组织缺血缺氧后的一个重要病理生理变化。近年来研究发现,胞外pH值下降能激活一种特殊的配体门控通道,酸敏感离子通道(Ac id-sensing ion channels—AS I Cs),这极大地改变了人们对缺血性脑损伤机制的认识[1]。目前,已经克隆出六种ASIC亚基(1a、1 b、2a、2b、3和4) [2],其中,H+敏感性高且对Ca2+通透ASIC1 a同聚体通道和ASIC1 a/2b通道在参与脑缺血和再灌注损伤过程中发挥着重要作用[3-8]。布洛芬是临床上常用的抗炎镇痛药,早在上世纪九十年代就有文章报道了其脑保护作用[9,10]。但是,对于其神经保护作用的机制目前还没有统一的定论。自从ASICs被发现以来,已有多篇文献[11-13]报道了布洛芬对ASICs的作用。本研究旨在①观察大脑中动脉阻塞(MCAO)后大鼠皮层缺血半暗带ASIC1a和ASIC2a表达变化情况并探讨其意义;②观察布洛芬的神经保护作用及其对ASIC1a和ASIC2a表达和上膜的影响,探索其以ASIC2a为靶点的神经保护作用的机制。方法:①20只SD大鼠随机分为假手术组(sham) 10只和MCAO模型组(MCAO) 10只,于术后24h取脑,提取脑组织蛋白和分离膜蛋白,用Western blot检测ASIC1a和ASIC2a的总蛋白和膜蛋白情况;②48只SD大鼠随机分为假手术+溶媒组(S+R n=12)、假手术+布洛芬组(S+B n=12)、MCAO模型+溶媒组(C+R n=12)、MCAO模型+布洛芬组(C+B n=12),于术后24h通过神经行为学评分、TTC染色等方法观察布洛芬的脑保护作用和用Western blot观察布洛芬对皮层缺血半暗带ASIC1a和ASIC2a蛋白含量的影响。结果:①脑缺血再灌注24h后,与假手术组相比,MCAO模型组皮层缺血半暗带的ASIC1a的表达没有明显变化(P=0.7004),其膜蛋白含量增加(P 0.001),ASIC2a的表达明显上调(P 0.001);②布洛芬能提高MCAO模型大鼠的神经行为学评分(P0.05),减少脑梗死容积(P0.01),并且能降低皮层缺血半暗带ASIC1a膜蛋白的含量(P 0.05)和下调ASIC2a的表达(P0.01)。结论:布洛芬能够明显改善大鼠脑缺血再灌注损伤后的神经行为学障碍,缩小脑梗死的范围,具有神经保护作用;布洛芬可能通过下调皮层缺血半暗带ASIC2a的表达,减少ASIC1a的膜上分布,发挥神经保护作用。
[Abstract]:Background and objective: stroke is an acute cerebrovascular disease. Because of its high incidence, high recurrence rate, high disability rate and mortality, it seriously affects the quality of life of patients. The incidence of ischemic stroke is higher than that of hemorrhagic stroke, accounting for 60-70% of the total stroke. The pathogenesis and pathophysiological process of ischemic stroke are extremely complex. Acidosis is an important pathophysiological change after ischemia and hypoxia. In recent years, it has been found that the decrease of extracellular pH can activate a special ligand gated channel. The acid-sensitive ion channel, ac id-sensing ion channels-AS I CSN, has greatly changed the understanding of the mechanism of ischemic brain injury [1]. Six subunits of ASIC have been cloned, I. e., 1 ASIC subunit, 1 BX, 2a, 2bt3 and 4) [2], of which H sensitive and transparent to ASIC1 a homopolymer channel and ASIC1 a / 2b channel play an important role in the process of cerebral ischemia and reperfusion injury [3-8]. Ibuprofen is a common clinical practice. Used as an anti-inflammatory analgesics, As early as the nineties of the last century, it was reported that its brain protective effect [9]. However, there is no uniform conclusion on the mechanism of its neuroprotective effect. Since the discovery of ASICs, The effect of ibuprofen on ASICs has been reported in several papers [11-13]. The purpose of this study was to observe the changes of ASIC1a and ASIC2a expression in ischemic penumbra of rat cortex after middle cerebral artery occlusion (MCAO) and to explore its significance. Protective effects and their effects on the expression of ASIC1a and ASIC2a, and on the membrane, Methods one hundred and twenty Sprague-Dawley rats were randomly divided into sham group (n = 10) and MCAO model group (n = 10). Brain tissue proteins and membrane proteins were extracted 24 hours after operation. The total protein and membrane protein of ASIC1a and ASIC2a were detected by Western blot. 248 SD rats were randomly divided into sham-operated medium group and sham-operated ibuprofen group, and the sham-operated ibuprofen group and the sham-operated ibuprofen group were divided into two groups: the sham-operated ibuprofen group and the sham-operated ibuprofen group. The protective effect of ibuprofen on brain and the effect of ibuprofen on the contents of ASIC1a and ASIC2a protein in ischemic penumbra of cortex were observed by neurobehavioral score and TTC staining in 24 hours. Results after 24 hours of cerebral ischemia and reperfusion, the effects of ibuprofen on the contents of ASIC1a and ASIC2a in cerebral ischemia penumbra were observed. Compared with the sham-operated group, the expression of ASIC1a in the ischemic penumbra of the MCAO model group did not change significantly. The increase of membrane protein content in the MCAO model group increased the expression of ASIC2a and increased the neurobehavioral score (P0.05) of the MCAO model rats. Conclusion: ibuprofen can significantly improve neurobehavioral disorders after cerebral ischemia reperfusion injury in rats, and can decrease the content of ASIC1a membrane protein in ischemic penumbra of cortex and down-regulate the expression of ASIC2a. Ibuprofen may play a neuroprotective role by down-regulating the expression of ASIC2a in the ischemic penumbra of cortex and reducing the distribution of ASIC1a on the membrane.
【学位授予单位】：中国人民解放军医学院
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R743.3

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