新生大鼠缺氧缺血性脑损伤HIF-1a表达与去铁胺的作用机制

发布时间：2018-04-03 07:50

本文选题：去铁胺　切入点：低氧诱导因子-1α　出处：《山西医科大学》2012年硕士论文

【摘要】：目的：新生儿缺氧缺血性脑损伤(hypoxic-ischemic brain damage ,HIBD)是指各种围生期窒息而导致脑的缺氧缺血性损害，临床出现一系列脑病的表现,是我国伤残儿童致残重要的原因之一。本实验通过建立新生大鼠缺氧缺血性脑损伤(HIBD)模型，观察缺氧诱导因子1α(hypoxia-inducible factor 1α)表达与去铁胺（deferoxamine，DFO）可能的保护机制，为其在新生儿缺氧缺血性脑病（hypoxic-ischemic encephalopathy, HIE）的临床应用提供理论依据。方法：新生7日龄Wistar大鼠120只随机分为假手术组（8只）、HIBD模型组及去铁胺（DFO）组，后两组再根据处死时间不同又分为7个亚组：3h组、6h组、12h组、24h组、48h组、3d组及7d组，每亚组各8只。假手术组给予颈正中切口游离左侧颈总动脉，不进行缺血缺氧。模型采用阻断左侧颈总动脉后置于含8%O2的低氧环境中2小时制备而成。去铁胺组,制成HIBD动物模型前24 h以去铁胺(200mg/kg,溶于生理盐水)单次腹腔内注射。各组分别于不同时间点处死动物,肉眼观察脑组织大体形态变化，，光镜下观察各组大鼠左侧脑组织HE染色的病理改变，并采用免疫组织化学方法检测HIF-1α和Caspase3的表达。结果：(1)缺氧缺血后新生大鼠出现不同程度的行为异常。(2)缺氧缺血后各时间点可见脑组织大体有不同程度的异常改变,去铁胺干预后上述改变减轻。(3)HE染色：假手术组脑组织结构及细胞层次清晰，神经元排列整齐紧密；HIBD模型组各时间点出现不同程度的神经细胞肿胀、变性、坏死，细胞核碎裂、溶解，神经元数目减少，胶质细胞增生。去铁胺组各时间点损伤程度较HIBD组明显减轻，神经细胞存活数量多，细胞排列尚规则，仅见少量神经细胞变性坏死。(4)免疫组织化学染色：HIF-1α及Caspase-3的阳性表达均呈棕黄色细颗粒沉积，阳性细胞可表达于大脑皮层和海马，阳性着色主要位于神经细胞胞浆及突起。①HIF-1α的表达：假手术组HIF-1α呈低水平表达，各时间点无明显变化；HIBD模型组各时间点均可见HIF-1α阳性表达，同一时间点与假手术组相比，HIF-1α表达明显增加，差异均有统计学意义（P0.05）。HIF-1α在HIBD后3h即增强，12h达高峰，之后逐渐降低；而DFO组则6h达高峰，12 h和24 h仍在较高水平，各时间点HIF-1α表达水平较HIBD组明显升高,差异有统计学意义（P0.05）。②Caspase-3的表达：假手术组Caspase-3呈低水平表达，各时间点无明显变化；HIBD模型组Caspase-3阳性表达，6h明显升高, 12h和24h仍在较高水平，7d开始降低仍保持一定的表达水平，且各时间点表达高于假手术组,两组差异有统计学( P＜0.05)；DFO组，3h轻微表达，12h明显升高, 24h和48h仍保持一定的表达水平，7d开始降低,DFO组各时间点Caspase3表达低于HIBD组，两组差异有统计学意义（P＜0．05）。TUNEL阳性细胞(AI指数)表达主要分布于皮层及海马区。假手术组TUNEL阳性细胞极少，HIBD组TUNEL阳性细胞较对照组增多,且随着时间延长逐渐增多；两组比较差异有统计学意义（P0.05）。DFO组TUNEL阳性细胞比HIBD组明显降低，两组比较差异有统计学意义（P0.05）。结论：（1）新生大鼠给予去铁胺后HIBD的可减轻脑组织的病理改变，提示去铁胺对新生大鼠HIBD可能具有神经保护作用。（2）去铁胺可能通过上调HIF-1α的表达，降低HIBD后Caspase-3的表达，减轻HIBD病理损伤过程，抑制神经细胞凋亡，从而对新生大鼠缺氧缺血引起的神经细胞损伤起到一定的保护作用，进一步为临床治疗新生儿缺氧缺血性脑损伤提供新的治疗思路。
[Abstract]:Objective : To observe the protective mechanism of hypoxia - inducible factor 1伪 ( hypoxia - inducible factor 1伪 ) and deferrioxamine ( DFO ) in neonatal hypoxic - ischemic brain damage ( HIBD ) and to provide a theoretical basis for the clinical application of hypoxic - ischemic encephalopathy ( HIE ) .
Methods : 120 Wistar rats were randomly divided into sham operation group ( 8 rats ) , HIBD model group and deferrioxamine ( DFO ) group . The rats were divided into 7 subgroups : 3h group , 6h group , 12h group , 24h group , 48h group , 3d group and 7d group .
Results : ( 1 ) Different degrees of behavioral abnormalities were observed in neonatal rats after hypoxia ischemia . ( 2 ) There were some abnormal changes in brain tissue at various time points after hypoxia ischemia .
The expression of HIF - 1伪 and Caspase - 3 was significantly lower than HIBD group . The positive staining of HIF - 1伪 and Caspase - 3 was found in the cerebral cortex and hippocampus .
HIF - 1伪 positive expression was found at all time points in HIBD model group , and the expression of HIF - 1伪 was significantly increased in the same time point compared with sham operation group ( P0.05 ) . HIF - 1伪 increased at 3h after HIBD , peaked at 12h , and then decreased gradually ;
Compared with HIBD group , the expression level of HIF - 1伪 in DFO group was significantly higher than that in HIBD group ( P0.05 ) .
Caspase - 3 positive expression in HIBD model group was significantly higher than that in sham operation group at 6 h , 12 h and 24 h , and the expression level was higher than that in sham operation group ( P < 0.05 ) .
The expression levels of Caspase - 3 in DFO group and DFO group were significantly higher than those in HIBD group ( P < 0.05 ) . TUNEL positive cells ( AI index ) were mainly distributed in cortex and hippocampus . TUNEL positive cells in sham operation group were very few , TUNEL positive cells in HIBD group were higher than those of control group , and gradually increased with time .
Compared with HIBD group , TUNEL positive cells in DFO group were significantly lower than those in HIBD group ( P0.05 ) .
Conclusion : ( 1 ) In neonatal rats , HIBD can reduce the pathological changes of HIBD . It is suggested that deferrioxamine may have neuroprotection effect on neonatal rat HIBD . ( 2 ) Deferrioxamine may inhibit the expression of HIF - 1伪 , decrease the expression of Caspase - 3 after HIBD , reduce the apoptosis of HIBD , and provide a new treatment for neonatal hypoxic - ischemic brain injury .

【学位授予单位】：山西医科大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R722.1

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