亚低温干预下新生大鼠HIBD脑组织Bax与HIF-1α基因表达的研究

发布时间：2018-05-28 03:09

本文选题：亚低温(mildhypothemia) + 缺氧缺血性脑损伤(HIBD)　；参考：《山西医科大学》2014年硕士论文

【摘要】：目的：新生儿缺氧缺血性脑病（hypoxic-ischemic encephalopathy,HIE）是由于围产期窒息缺氧所引起的神经系统损伤，是新生儿时期最常见的疾病之一，为婴幼儿永久性神经系统功能障碍甚至是导致新生儿死亡的重要原因。现阶段临床上的药物疗法疗效甚微。本文的目的是在成功建立新生大鼠缺氧缺血性脑损伤（hypoxic-ischemic brain damage,HIBD）动物模型后，立刻给予全身性亚低温（mild hypothemia）治疗，在不同的时间点检测不同组别新生大鼠患侧脑组织Bax及HIF-1α基因的表达并探究两者之间的关系。试图阐明亚低温干预在治疗新生大鼠HIBD时可能的神经保护机制，为现阶段临床上治疗新生儿HIE提供更进一步的理论依据。方法：新生7日龄SD大鼠120只，雌雄不拘，随机分成三组：假手术组（A组），常温组（B组），亚低温组（C组），每组各40只。B组和C组均需先制备HIBD动物模型。三组均根据实验要求即处死时间点的不同分为2h组、6h组、12h组、24h组、48h组5个亚组，每亚组各8只。B组和C组新生大鼠均采用阻断左侧颈总动脉，后放置于含8%O2+92%NO2的环境中缺氧2h制备动物模型，其过程中进行行为学观察；C组新生大鼠HIBD模型制备成功后立刻予全身性亚低温处理。各组根据实验要求采用断头法提取脑组织。观察脑组织大体形态变化，采用实时荧光定量PCR（real-timePCRRT-PCR）法分别检测不同组别新生大鼠各时间点患侧脑组织Bax及HIF-1α基因表达变化，记录统计结果，行统计学分析。结果：（1）行HIBD造模的新生大鼠前后均出现了异常的行为活动。（2）肉眼观察B组患侧脑组织各时间点均出现明显水肿，，而C组脑组织各时间点无水肿。（3）BaxmRNA的表达：A组新生大鼠脑组织各时间点BaxmRNA的表达甚微。B组新生大鼠脑组织各时间点BaxmRNA表达2h上调且逐渐增加，12h达高峰，12h-48h维持在高峰。C组新生大鼠脑组织各时间点BaxmRNA的表达2h出现下降趋势并于6h达到最低水平，后期稍升高，即12h~48h，BaxmRNA的表达出现微升高。（4）HIF-1αmRNA表达：A组新生大鼠脑组织各时间点HIF-1αmRNA的表达偏少，B组新生大鼠脑组织各时间点HIF-1αmRNA表达2h出现升高且逐渐增加，48h达到高峰；C组新生大鼠脑组织各时间点HIF-1αmRNA表达2h稍升高，后期呈逐渐降低趋势。结论：（1）新生大鼠HIBD后予亚低温干预，可使HIBD后脑组织的水肿程度减轻，以提示亚低温干预对新生大鼠HIBD后的神经系统可能发挥保护性作用。（2）亚低温可能抑制促凋亡基因BaxmRNA表达，减轻HIBD后的病理损伤过程，从而抑制神经细胞凋亡。（3）亚低温干预后期HIF-1αmRNA的表达使下游促凋亡基因BaxmRNA表达较前略有增加，从而清除HIBD后脑组织已凋亡细胞，对新生大鼠HIBD后的神经系统起到一定的保护作用，从而为新生儿HIE提供新的临床诊疗思路。
[Abstract]:Objective: Hypoxic-ischemic encephalopathy (HIEE) is a nervous system injury caused by perinatal asphyxia and is one of the most common diseases in neonatal period. Permanent neurological dysfunction is an important cause of neonatal death. At this stage, the clinical efficacy of drug therapy is minimal. The aim of this study was to treat neonatal rats with hypoxic-ischemic brain damage (HIBD) with mild hypothermia immediately after the establishment of HIBD model. The expression of Bax and HIF-1 伪 genes in brain tissue of different groups of newborn rats were detected at different time points and the relationship between them was explored. This paper attempts to elucidate the possible neuroprotective mechanism of mild hypothermia intervention in the treatment of neonatal HIBD in order to provide a further theoretical basis for the clinical treatment of neonatal HIE at this stage. Methods: 120 male and female Sprague-Dawley rats of 7 days old were randomly divided into three groups: sham operation group (group A), normal temperature group (group B) and mild hypothermia group (group C). According to the requirements of the experiment, the three groups were divided into 5 subgroups, including 2 h group, 6 h group, 12 h group, 24 h group, 48 h group, 8 rats in each subgroup, 8 rats in group B and group C, all of which were treated with occlusion of left common carotid artery. The animal model was established by hypoxia for 2 hours in the environment containing 8%O2 92%NO2. During the course of behavioral observation, the HIBD model of neonatal rats in group C was treated with systemic mild hypothermia immediately after the establishment of HIBD model. According to the requirements of the experiment, the brain tissue was extracted by head-cut method in each group. The changes of Bax and HIF-1 伪 gene expression in brain tissue of different groups of neonatal rats were detected by real-time PCR RT-PCRmethod. The statistical results were recorded and analyzed statistically. Results (1) abnormal behavioral activity was observed before and after HIBD modeling in newborn rats. (1) the brain tissue of the affected side of group B showed obvious edema at every time point after observation with naked eyes. The expression of BaxmRNA in brain tissue of neonatal rats in group C was minimal at each time point. The expression of BaxmRNA in brain tissue of group B was up-regulated for 2 h and gradually increased at 12 h to reach the peak at 12 h and maintained at the peak at 48 h. In group C, the expression of BaxmRNA in brain tissue of neonatal rats decreased at 2 h and reached the lowest level at 6 h. Later, a little higher, The expression of HIF-1 伪 mRNA in brain tissue of neonatal rats in group A was slightly increased at 12 h and 48 h. The expression of HIF-1 伪 mRNA in brain tissue of neonatal rats in group A was slightly lower than that in group B and the expression of HIF-1 伪 mRNA in brain tissue of neonatal rats in group B increased for 2 h and gradually increased to the peak at 48 h. The expression of HIF-1 伪 mRNA in the brain of neonatal rats in group C reached the peak at 48 h. The expression of HIF-1 伪 mRNA increased slightly at different time points. In the later stage, the trend was decreasing gradually. Conclusion (1) mild hypothermia intervention after HIBD in neonatal rats can reduce the edema of brain tissue after HIBD, which suggests that mild hypothermia intervention may play a protective role in the nervous system after HIBD in neonatal rats. 2) mild hypothermia may inhibit the expression of pro-apoptotic gene BaxmRNA. After mild hypothermia intervention, the expression of HIF-1 伪 mRNA increased slightly, and the BaxmRNA expression of the downstream apoptotic gene increased slightly, thus eliminating the apoptotic cells in brain tissue after HIBD. It can protect the nervous system of newborn rats after HIBD and provide a new way of clinical diagnosis and treatment for neonatal HIE.
【学位授予单位】：山西医科大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R742

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