miR-29C通过靶基因Bak1在预电刺激小脑顶核诱导的大鼠脑缺血再灌注损伤中的作用

发布时间：2018-07-09 12:55

本文选题：电刺激 + 靶基因预测　；参考：《广西医科大学》2014年硕士论文

【摘要】：目的：研究预电刺激小脑顶核诱导大鼠脑缺血再灌注损伤过程中miR-29C对Bak1的调控作用，，并探讨预电刺激小脑顶核的保护机制。方法：60只体重280-300g的SD成年雄性大鼠，随机分成3组：(1)预电刺激小脑顶核+大脑中动脉阻塞模型（预电刺激组）；（2）大脑中动脉阻塞模型组（单纯造模组）；(3)假手术组。预电刺激组的处理为电刺激左侧小脑顶核后制作大脑中动脉阻塞模型，缺血2小时后再灌注，再灌注24小时后处死。通过TTC染色测定梗死体积；动物行为学评分评估神经功能缺损情况；TUNEL组织化学染色检测凋亡指数；芯片分析预电刺激组和单纯造模组间差异表达的miRNA并利用定量PCR验证；并通过生物信息学软件预测该的miRNA靶基因；最后利用双荧光报告验证其与靶基因的关系。结果：预电刺激组和单纯造模组相比，脑梗死体积明显减少（37.0±8.9%vs62.0±10%）（P0.01）；TUNEL染色检测的凋亡指数下降（23.4±2.4%vs58.9±4.0%）(P0.01)；动物行为学评分降低（1.5±0.52vs2.4±0.51）(P0.01)；芯片分析发现miR-29C在两组间表达差异最大； PCR验证miR-29C在预电刺激组较单纯造模组表达下调(P0.01)；生物信息学软件预测是Bak1是miR-29C的靶基因；双荧光报告体外验证Bak1-wt/rnomiR-29C组的相对荧光强度在Bak1-wt/NC、Bak1-wt/rno-miR-29C、Bak1-Mut/NC和Bak1-Mut/rno-miR-29C4组中最小(P0.01)，并且其余三组间无明显差异(P0.05) 结论：预电刺激大鼠小脑顶核可以减轻脑缺血再灌注损伤；预电刺激小脑顶核可以下调miR-29C的表达；Bak1是miR-29C的靶基因；miR-29C是否通过调控Bak1在预电刺激诱导的缺血脑保护机制中起作用还需要进一步体内验证。
[Abstract]:Aim: to investigate the regulatory effect of miR-29C on Bak1 during cerebellar fastigial nucleus stimulation in rats with cerebral ischemia-reperfusion injury, and to explore the protective mechanism of preelectric stimulation of cerebellar parietal nucleus. Methods 60 adult Sprague-Dawley male rats weighing 280-300g were randomly divided into three groups: (1) the model of middle cerebral artery occlusion in cerebellar parietal nucleus (); (_ 2) and the sham-operation group (); (_ 3) in prestimulation group. The model of middle cerebral artery occlusion was made after electrical stimulation of left cerebellar fastigial nucleus in preelectric stimulation group. After 2 hours of ischemia and reperfusion, the rats were killed after 24 hours of reperfusion. The infarct volume was measured by TTC staining, the neurological impairment was evaluated by animal behavior score and apoptosis index was detected by Tunel histochemical staining, miRNA differentially expressed between preelectric stimulation group and simple model group was analyzed by microarray and verified by quantitative PCR. The miRNA target gene was predicted by bioinformatics software, and the relationship between miRNA target gene and the target gene was verified by double fluorescence report. Results: compared with the control group, the volume of cerebral infarction was significantly decreased (37.0 卤8.9%vs62.0 卤10%) (P0.01), the apoptotic index was decreased (23.4 卤2.4%vs58.9 卤4.0%), the animal behavior score was decreased (1.5 卤0.52vs2.4 卤0.51) (P0.01), and the expression of miR-29C was found to be the most significant difference between the two groups by microarray analysis. The expression of miR-29C was down-regulated in preelectric stimulation group (P0.01), and the bioinformatics software predicted that Bak1 was the target gene of miR-29C. The double-fluorescence report confirmed in vitro that the relative fluorescence intensity of Bak1-wt / rnomiR-29C group was the smallest in Bak1-wtrno-miR-29C group (P0.01), and the other three groups had no significant difference (P0.05). Conclusion: preelectric stimulation of cerebellar fastigial nucleus can reduce cerebral ischemia-reperfusion injury in rats. The expression of miR-29C can be down-regulated by prestimulation of cerebellar fastigial nucleus. Whether miR-29C, the target gene of miR-29C, may play a role in the protective mechanism of ischemic brain induced by preelectric stimulation needs to be further verified in vivo.
【学位授予单位】：广西医科大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R743.3

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