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Effect of NBP on the Expression p-ERK, Caspase-3 in Gerbil Cerebral Ischemia-reperfusion

Author: ZhaoLiZuo
Tutor: NieYingXue
School: China Medical University
Course: Neurology
Keywords: NBP p-ERK Caspase-3 cerebral ischemia and reperfusion
CLC: R743.31
Type: Master's thesis
Year: 2010
Downloads: 115
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Abstract


PrefaceIschemic stroke have a high incidence, high mortality and high disability, so prevention and treatment of ischemic stroke of social and medical attention to an important issue. The extent of ischemia-reperfusion injury of ischemic stroke is an important factor in Prognosis of ischemic stroke. In recent years, research on the pathogenesis、prevention and treatment of Cerebral ischemia-reperfusion is more and more. NBP by the Chinese Academy of Medical Sciences of drugs developed by the Institute in 2002, has been approved by the State Food and Drug Administration for the treatment of ischemic stroke in a class of drugs. Many existing research shows that NBP can improve cerebral ischemic damage from multiple links.Many studies have shown that, Extracellular signal-regulated kinase (extracellular signal-regulated kinase, ERK) play an important regulatory role of the cerebral ischemia-reperfusion injury in cell proliferation, differentiation and apoptosis. Caspase-3 directly involved in the cerebral ischemia-reperfusion injury in the occurrence of apoptosis, is an important hub for apoptosis pathway.In this experiment, we have adopted a gerbil model of cerebral ischemia-reperfusion dynamic observation of NBP on the p-ERK, Caspase-3 expression after cerebral ischemia-reperfusion injury.Materials andMethod1.GroupsAll 54 healthy male gerbils (body weight 60-70g) were randomly divided into 3 groups:Sham-operated group (CON), ischemia-reperfusion group (I/R), NBP treated group (I/R+NBP),Each group is divided into three sub-groups (1d,3d,7d). Sham-operated group, only free, but not bilateral carotid artery occlusion, Intraperitoneal injection of vegetable oil; Ischemia-reperfusion group were applied to bilateral common carotid artery clamp technique, Release the clip immediately after the intraperitoneal injection of vegetable oil after ischemia 10 minutes; NBP treatment group modeling process with the ischemia-reperfusion, Release the clip on the NBP 40mg/kg intraperitoneally immediately after ischemia 10 minutes. Each group using the Western Blot to detect p-ERK, caspase-3 expression in hippocampusin CA1 after the reperfusion 1d,3d,7d.observed hippocampal CA1 neurona pyramidal cells changes by Nissl staining.2.Preparation of gerbil cerebral ischemia-reperfusion modelUse of bilateral common carotid artery clamp technique produced gerbil model of global cerebral ischemia-reperfusion.3.Nissl stainingParaffin sections after dewaxing through ddH2O washing, and then placed in Nissl dye liquor in the dyeing, after the two sequential dehydration, the last neutral gum Fengpian. Observed hippocampal CA1 neuronal pyramidal cells changes in the microscope.4.Western blot Measuring gerbil hippocampal CA1 area p-ERK, Caspase-3Each gerbil was isolated from the left cerebral hemisphere, hippocampus, weighing, preparation of protein samples, implementation of SDS-polyacrylamide gel electrophoresis, and then transfer to a PVDF membrane. Closed separately with polyclonal anti-phospho-ERK antibodies and rabbit anti-Caspase-3 polyclonal antibody level of shaking 4℃overnight. And then at room temperature with the appropriate level of two anti-shaker 1h, TBST wash membrane 10min×3 times, luminous, analysis purpose bands with Image J.Results1. In hemicroscope, NBP treated group compared with ischemia-reperfusion group, the cells large, round, deeply stained Nissl bodies, but still shows a small amount of cytoplasm stain. From the changes in pyramidal cells on reperfusion a significant reduction in neurons, non-coloring, larger number of neurons in the NBP treatment group (p<0.01);. Vertebral body cell become least when the reperfusion time 3d.NBP group and reperfusion group Trend unanimously. 1. 1d,the NBP group than in the ischemia-reperfusion group p-ERK increased significantly(p<0.05).3 d,the expression of the two groups have decreased, but the NBP group was still significantly higher than reperfusion group (p<0.01).7 d when the reperfusion group, the expression increased again, while the NBP group decreased with the sham-operated group. Difference between the two groups was statistically significant (p<0.01).2. 1d, the NBP treatment group can be reduced in hippocampus after ischemia-reperfusion Caspase-3 protein expression (p<0.05).3d, Caspase-3 reached the peak, NBP treated group than reperfusion group expression was significantly lower (p<0.05).7d each group were down, no statistical significance.DisscussionButylphthalide (dl-3n-butylphthalide, NBP) is a specialized state-level for the treatment of cerebral thrombosis a class of drugs, a number of studies have shown the drug to the brain tissue after ischemia has a protective effect. By Nissl staining in this experiment can be observed from the morphology of the NBP treatment group compared with gerbil hippocampal CA1 cells in ischemia-reperfusion group intact, arranged more neatly, the cells large, round, deeply stained Nissl bodies, reducing the loss of pyramidal cells. The results show that NBP to relieve cerebral ischemia-reperfusion injury, consistent with past experiments. At present, some studies suggest that ERK activation and cell proliferation, can promote cell survival. This experiment studies have shown that, NBP group than in the ischemia-reperfusion group of p-ERK increased significantly (p<0.05), prompted NBP may be by regulating the expression of p-ERK play a role in protection of brain tissue. Caspase-3 is the key protease during apoptosis, the experimental studies have shown that NBP treatment can significantly inhibit Caspase-3 elevated (P<0.05), Shows that NBP can play a role in protecting brain tissue by inhibiting the activity of Caspase-3 increased.Conclusions1. NBP can improve neuronal cells damage in gerbils hippocampal CA1 after cerebral ischemia and reperfusion2. NBP can increase the gerbil hippocampal CA1 area p-ERK expression after cerebral ischemia-reperfusion. 3. NBP can inhibit the gerbil hippocampal CA1 area Caspase-3 expression after cerebral ischemia-reperfusion

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CLC: > Medicine, health > Neurology and psychiatry > Neurology > Cerebrovascular disease > Acute cerebrovascular disease ( stroke) > Transient ischemic
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