肾上腺髓质素对大鼠脑缺血再灌注神经元的作用及机制
发布时间：2018-11-20 09:43
【摘要】：目的 肾上腺髓质素(ADM)是由52个氨基酸组成的血管活性多肽,属于降钙素基因相关肽(CGRP)家族,与多种组织的缺血再灌注损伤有关。有学者证实在中枢神经系统有ADM的表达,并与脑缺血再灌注损伤有一定关系。也有持相反意见的研究报道。所以我们选用大鼠局灶性脑缺血再灌注模型,应用免疫组织化学方法、原位杂交组织化学法、逆转录—聚合酶链式(RT-PCR)分析方法等,验证正常大鼠脑组织是否有ADM及ADM mRNA的表达及具体表达的部位;观察ADM及ADM mRNA在局灶性缺血再灌注大鼠脑组织中的表达变化规律;探讨ADM对局灶性缺血再灌注大鼠神经元凋亡、梗死体积及Egr-1 mRNA的影响,进一步研究ADM在局灶性缺血再灌注脑损伤中的作用。 方法 健康雄性SD大鼠186只,体重200-250g,由中国医科大学实验动物中心提供。随机分为正常组(n=12),假手术组(n=12),局灶性脑缺血2h再灌注2h组(n=18)、4h组(n=18)、22h组(n=54)、46h组(n=18)、70h组(n=18)、118h组(n=18)、166h组(n=18)。其中再灌注22h组又分为:股静脉注射ADM组(n=12),颈内动脉注射ADM组(n=12),侧脑室注射ADM组(n=12)。采用线栓法制成大鼠大脑中动脉缺血再灌注(MCAO)模型,阻断血流2h进行再灌注。缺血再灌注各组大鼠在缺血2h再灌注4h时,参考Bederson等的5分制评分标准对其进行神经功能缺损评分,评分后分别制备石蜡切片和冰冻切片,TTC染色测定梗死体积,HE染色光镜下观察大鼠大脑皮质、海马的组织学变化,TUNEL法检测神经元凋亡,免疫组织化学法(SABC法)检测ADM阳性细胞表达,原位杂交法检测ADM mRNA
[Abstract]:Objective adrenomedullin (ADM) is a vasoactive polypeptide consisting of 52 amino acids. It belongs to the calcitonin gene-related peptide (CGRP) family and is related to ischemia-reperfusion injury in various tissues. Some scholars have confirmed the expression of ADM in the central nervous system, which is related to cerebral ischemia reperfusion injury. There are also reports of contrary views. So we chose the rat model of focal cerebral ischemia-reperfusion, using immunohistochemistry, in situ hybridization histochemistry, reverse transcription-polymerase chain reaction (RT-PCR), and so on. To verify the expression of ADM and ADM mRNA in the brain of normal rats. To observe the expression of ADM and ADM mRNA in the brain of rats with focal ischemia-reperfusion. To investigate the effects of ADM on neuronal apoptosis, infarct volume and Egr-1 mRNA in focal ischemia-reperfusion rats, and to further study the role of ADM in focal ischemia-reperfusion brain injury. Methods 186 healthy male SD rats, 200-250 g, were provided by Experimental Animal Center of China Medical University. They were randomly divided into 4 groups: normal group (nnm12), sham-operation group (NN12), focal cerebral ischemia 2h reperfusion group (NN18), focal cerebral ischemia 2h reperfusion 2 h group (NN18), 4h group (nong18), 22h group (nnm54), 46h group (nm18), 70h group (nm18), 118h group (N18), 166h group (NN18). Among them, 22 h reperfusion group was divided into three groups: femoral vein injection group (ADM 12), internal carotid artery injection group (ADM group 12), lateral ventricle injection group (ADM group 12). The middle cerebral artery (MCA) ischemia reperfusion (MCAO) model was established by the method of thread embolization, and the blood flow was blocked for 2 h for reperfusion. After ischemia 2 h and reperfusion 4 h after ischemia, the rats were evaluated for neurological impairment by referring to the 5-point scoring standard of Bederson et al. After scoring, paraffin sections and frozen sections were prepared, and the infarct volume was measured by TTC staining. Histological changes of cerebral cortex and hippocampus of rats were observed under HE staining light microscope, neuronal apoptosis was detected by TUNEL method, expression of ADM positive cells was detected by immunohistochemistry (SABC method), ADM mRNA was detected by in situ hybridization.
【学位授予单位】：中国医科大学
【学位级别】：博士
【学位授予年份】：2005
【分类号】：R363
本文编号：2344564
[Abstract]:Objective adrenomedullin (ADM) is a vasoactive polypeptide consisting of 52 amino acids. It belongs to the calcitonin gene-related peptide (CGRP) family and is related to ischemia-reperfusion injury in various tissues. Some scholars have confirmed the expression of ADM in the central nervous system, which is related to cerebral ischemia reperfusion injury. There are also reports of contrary views. So we chose the rat model of focal cerebral ischemia-reperfusion, using immunohistochemistry, in situ hybridization histochemistry, reverse transcription-polymerase chain reaction (RT-PCR), and so on. To verify the expression of ADM and ADM mRNA in the brain of normal rats. To observe the expression of ADM and ADM mRNA in the brain of rats with focal ischemia-reperfusion. To investigate the effects of ADM on neuronal apoptosis, infarct volume and Egr-1 mRNA in focal ischemia-reperfusion rats, and to further study the role of ADM in focal ischemia-reperfusion brain injury. Methods 186 healthy male SD rats, 200-250 g, were provided by Experimental Animal Center of China Medical University. They were randomly divided into 4 groups: normal group (nnm12), sham-operation group (NN12), focal cerebral ischemia 2h reperfusion group (NN18), focal cerebral ischemia 2h reperfusion 2 h group (NN18), 4h group (nong18), 22h group (nnm54), 46h group (nm18), 70h group (nm18), 118h group (N18), 166h group (NN18). Among them, 22 h reperfusion group was divided into three groups: femoral vein injection group (ADM 12), internal carotid artery injection group (ADM group 12), lateral ventricle injection group (ADM group 12). The middle cerebral artery (MCA) ischemia reperfusion (MCAO) model was established by the method of thread embolization, and the blood flow was blocked for 2 h for reperfusion. After ischemia 2 h and reperfusion 4 h after ischemia, the rats were evaluated for neurological impairment by referring to the 5-point scoring standard of Bederson et al. After scoring, paraffin sections and frozen sections were prepared, and the infarct volume was measured by TTC staining. Histological changes of cerebral cortex and hippocampus of rats were observed under HE staining light microscope, neuronal apoptosis was detected by TUNEL method, expression of ADM positive cells was detected by immunohistochemistry (SABC method), ADM mRNA was detected by in situ hybridization.
【学位授予单位】：中国医科大学
【学位级别】：博士
【学位授予年份】：2005
【分类号】：R363
【共引文献】
相关期刊论文 前1条
1 毕国荣;张辉;周慧杰;张贺敏;海虹;方秀斌;;大鼠局灶性缺血再灌注后大脑皮质ADM及其MRNA的表达(英文)[J];Neuroscience Bulletin;2005年06期
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