意向性运动对局灶性脑缺血大鼠神经功能的影响及其机制研究

发布时间：2018-03-11 15:39

本文选题：意向性运动　切入点：局灶性脑缺血　出处：《中南大学》2014年硕士论文　论文类型：学位论文

【摘要】：目的：通过观察意向性运动对局灶性脑缺血大鼠神经功能及缺血半暗带ERK-CREB通路表达及GluR2与PICK1共定位表达比率的影响,初步探讨意向性运动对局灶性脑缺血后神经功能的影响及其可能的作用机制。方法：雄性Sprague-Dawley (SD)大鼠144只,随机均分为大脑中动脉栓塞模型(MCAO)组、游泳运动(SE)组、饲养环境改变(EM)组和意向性运动(WM)组,各组又分7d、15d、30d三个亚组。采用线栓法制备MCAO大鼠模型,动态观察各组大鼠行为学变化,TTC染色观察不同运动方式对15d组大鼠脑梗死体积的影响,RT-PCR及免疫荧光单标染色动态检测缺血半暗带脑组织ERK、CREB、GluR2mRNA及pERK、pCREB蛋白的表达水平,免疫荧光双标染色检测15d组GluR2与PICK1的共定位表达比率。结果： 1.各组大鼠在脑缺血再灌注2h时的神经功能评分最高,1d后略下降,第3d略有回升,以后随时间推移各组评分均呈下降趋势,各组评分差别无统计学意义(P0.05)。SE组评分在再灌7d较MCAO组升高(P0.05),而再灌15、30d下降(P0.05)；EM组及WM组评分在再灌7、15、30d,均较MCAO组下降(P0.05),以WM组下降更明显。EM组及WM组大鼠的爬梯频率均呈逐渐增加趋势,且WM组较EM组显著增加(P0.05)。 2.TTC染色结果显示：脑缺血再灌注15d时,与MCAO组比较,SE组梗死体积差异无统计学意义(P0.05)；EM组及WM组脑梗死体积减小(P0.05),以WM组更明显(P0.05)。 3. RT-PCR结果显示：与MCAO组比较,SE组脑缺血再灌注7d时ERK、CREB mRNA的表达减少(P0.05),15、30d后的表达明显增加(P0.05),而GluR2mRNA的表达在7d、15d、30d时均明显增加(P0.05)；EM组及WM组各实验点ERK、CREB、GluR2mRNA的表达较MCAO组均增加(P0.05),以WM组更明显(P0.05)。 4.免疫荧光单标染色结果显示：pERK在胞核、胞浆、胞膜均有表达,pCREB主要表达于胞核。MCAO组在脑缺血再灌注15d时,pERK.pCREB表达较7d时上调,随后基本保持恒定；SE组、EM组、WM组在再灌注7、15、30d,两蛋白的表达随时间呈上升趋势；但SE组与MCAO组比较,两蛋白表达在再灌7d时减少(P0.05),15、30d明显增加(P0.05)；EM组在各时间点两蛋白的表达均较MCAO组增多(P0.05)；WM组两蛋白的表达在各时间点均较其他三组明显增多(P0.05)。 5.免疫荧光双标结果显示：GluR2与PICK1蛋白均表达于胞浆和胞膜,且两者的分布情况及变化趋势存在高度一致性。脑缺血再灌注15d,SE组、EM组及WM组GluR2、PICK1的阳性细胞数均较MCAO组明显增加,而WM组双标阳性细胞数较其他三组明显增加(P0.05) 结论：意向性运动能改善局灶性脑缺血大鼠的神经功能,减小脑梗死体积,且较游泳运动有更明显的优势,可能与其上调并激活缺血半暗带脑组织ERK-CREB通路,并增强GluR2与PICK1的相互作用有关。
[Abstract]:Objective: to observe the effects of intentionality exercise on the expression of ERK-CREB pathway in ischemic penumbra and the ratio of co-localization of GluR2 and PICK1 in focal cerebral ischemia rats. To explore the effect of intentionality exercise on neural function after focal cerebral ischemia and its possible mechanism. Methods: a total of 144 male Sprague-Dawley SD rats were randomly divided into three groups: middle cerebral artery embolization (MCAO) group, swimming exercise group (Sprague-Dawley) group, feeding environment change group and intentionality exercise group. Each group was divided into three subgroups: 7 d, 15 d and 30 d, respectively. The effects of different exercise modes on cerebral infarct volume in rats of 15 d group were observed dynamically by TTC staining. RT-PCR and immunofluorescence staining were used to dynamically detect the expression of ERKN CREBN GluR2 mRNA and pCREB protein in ischemic penumbra. The co-localization ratio of GluR2 and PICK1 in 15 d group was detected by double immunofluorescence staining. Results:. 1. After 2 hours of cerebral ischemia and reperfusion, the neurological function scores of the rats in each group decreased slightly after 1 day, then increased slightly on the third day, and then decreased with the passage of time. There was no significant difference in the scores of each group. The scores of group E were significantly higher than those of group MCAO on the 7th day after reperfusion, while the scores of group EM and group WM were lower than those of group MCAO on the 7th day after reperfusion. The climbing frequency of rats in the WM group was more obvious than that of the rats in the group of WM and the group of EM and WM, and the scores of the EM group and the WM group were lower than those of the group of MCAO on the 7th day after reperfusion, and the scores of the EM group and the WM group were lower than those of the group of MCAO on the 7th day after reperfusion. All showed a trend of gradual increase, Compared with EM group, WM group significantly increased P0.05. 2. The results of TTC staining showed that at 15 days after cerebral ischemia and reperfusion, there was no significant difference in infarct volume between SE group and MCAO group. 3. RT-PCR results showed that compared with MCAO group, the expression of ERKO CREB mRNA in SE group decreased significantly after 15 days of cerebral ischemia and reperfusion compared with that in MCAO group, and the expression of GluR2mRNA increased significantly at the end of the 15th day after reperfusion, while the expression of GluR2mRNA increased significantly in EM group and WM group on the 7th day, 15 days after reperfusion and 30 days after reperfusion, and the expression of ERKB CREB GluR2 mRNA was significantly higher than that in MCAO group. The addition of P0.05 was more obvious in WM group. 4. The results of immunofluorescence single label staining showed that the expression of pCREB in nucleus, cytoplasm and cell membrane was mainly up-regulated at 15 days after cerebral ischemia and reperfusion in MCAO group, and the expression of pERK.pCREB was up-regulated at 7 days after cerebral ischemia and reperfusion. After 30 days of reperfusion, the expression of two proteins increased in SE group, EM group and MCAO group, but compared with MCAO group, the expression of two proteins in SE group was higher than that in MCAO group. The expression of two proteins decreased on the 7th day after reperfusion. The expression of two proteins in the P0.05A EM group was significantly higher than that in the MCAO group at each time point, and the expression of the two proteins in the P0.05G group was significantly higher than that in the other three groups at each time point compared with the other three groups. 5. The results of immunofluorescence double labeling showed that both the protein of GluR2 and PICK1 were expressed in cytoplasm and membrane, and the distribution and change trend of them were highly consistent. The number of GluR2PICK1 positive cells in EM group and WM group were significantly higher than that in MCAO group on the 15th day after cerebral ischemia reperfusion, and the expression of GluR2PICK1 protein in both groups was significantly higher than that in MCAO group. The number of double-labeled positive cells in WM group was significantly higher than that in the other three groups (P0.05). Conclusion:. Intention exercise can improve the neural function and reduce the volume of cerebral infarction in rats with focal cerebral ischemia. It may be related to the up-regulation and activation of ERK-CREB pathway in ischemic penumbra. The interaction between GluR2 and PICK1 was enhanced.
【学位授予单位】：中南大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R743.3

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