经鼻给予神经生长因子改善脑外伤大鼠预后的相关机制研究

发布时间：2018-04-30 20:56

本文选题：颅脑外伤 + 神经生长因子　；参考：《南京大学》2013年硕士论文

【摘要】：颅脑外伤(traumatic brain injury, TBI)是青年人出现神经功能障碍最常见的原因之一,TBI引起的继发性损伤较原发性损伤更为严重,包括脑水肿、阿尔茨海默病(Alzheimer's disease, AD)等,造成患者预后不良。神经生长因子(nerve growth factor, NGF)具有修复受损组织的潜力,可以改善TBI大鼠的预后,但由于血脑屏障的作用,其临床使用方面受到了限制,而经鼻给药的方式有效的解决了这一难题。本研究分别从脑水肿和AD两方面入手,旨在探讨经鼻给予NGF改善TBI大鼠预后的潜在机制。研究将成年雄性Sprague-Dawley大鼠分为TBI组、NGF+TBI组和control组,参照Feeney自由落体损伤的方法制作TBI模型,NGF+TBI组大鼠经鼻给予NGF(50ul/d),TBI组大鼠经鼻给予等量的磷酸盐缓冲液。采用干湿重评估脑组织含水量,免疫组织化学法检测脑组织内水通道蛋白-4(aquaporin-4,AQP4)、tau蛋白的表达情况,取患侧脑皮质,western blotting法测定AQP4、tau蛋白、糖原合成酶激酶-3β(glycogen synthase kinase-3β, GSK-3β)的水平,ELISA法、qRT-PCI法测定白介素-1β (interleukin-1β, IL-1β)的蛋白及基因的表达水平。EMSA法测定脑组织内核因子-κB (nuclear factor-κB, NF-κB)的DNA结合活力。qRT-PCR法测定B淋巴细胞瘤-2(B-celllymphoma-2, Bcl-2)、蛋白水解酶-3(cysteinyl aspartate specific proteinase-3, caspase-3)的基因表达情况。与control组相比,TBI组大鼠脑组织含水量明显增高,脑皮质内AQP4的表达明显增高,经NGF治疗后,大鼠的脑组织含水量及AQP4的表达明显下降；TBI后,大鼠脑组织内tau蛋白、GSK-3β的表达较control组大鼠明显增加,UNGF+TBI组大鼠tau蛋白、GSK-3β的表达较TBI组明显下降。进一步研究结果提示,TBI后大鼠IL-1β的蛋白及基因水平均明显升高,NF-κB的DNA结合活性明显增加,Bcl-2的基因水平降低,caspase-3的基因水平升高,经鼻给予NGF后IL-1β的蛋白及基因水平均明显下降,NF-κB的DNA结合活性明显降低,Bcl-2的基因水平升高,caspase-3的基因水平降低。由此可推,经鼻给予NGF可以减轻AQP4相关的脑水肿,减少GSK-3β关联的tau蛋白的磷酸化,从而改善TBI大鼠的预后,以上两个作用均与NGF减轻了TBI后Bcl-2/caspase-3/NF-κB通路相关的炎症反应有关。
[Abstract]:Traumatic brain injury, TBI) is one of the most common causes of neurologic dysfunction in young people. The secondary injury caused by TBI is more serious than that caused by primary injury, including brain edema, Alzheimer's disease and so on, resulting in poor prognosis of the patients. Nerve growth factor (NGF) nerve growth factor, NGF) has the potential to repair damaged tissue and can improve the prognosis of TBI rats. However, the clinical use of nerve growth factor growth factor, NGF) is limited due to the effect of blood-brain barrier, and the method of nasal administration can effectively solve this problem. The purpose of this study was to explore the potential mechanism of nasal administration of NGF in improving the prognosis of TBI rats from the aspects of brain edema and AD. Adult male Sprague-Dawley rats were divided into TBI group and control group. According to the method of Feeney free falling body injury, the TBI model rats in NGF TBI group were given the same amount of phosphate buffer by nasal administration. The water content of brain tissue was evaluated by wet and dry weight, the expression of aquaporin-4 aquaporin-4aqp4ttau protein in brain tissue was detected by immunohistochemical method, and the tau protein was detected by western blotting method in the affected cerebral cortex. Expression of interleukin-1 尾 -interleukin-1 尾 (IL-1 尾) protein and gene by qRT-PCI. DNA binding activity of NF- 魏 B nuclear factor- 魏 B, NF- 魏 B, NF- 魏 B in brain tissue was assayed by DNA binding activity. QRT-PCR was used to detect B-cell lymphoma-2, Bcl-2U. Gene expression of protein hydrolase-3 aspartate specific proteinase-3 (caspase-3). Compared with the control group, the water content of brain tissue and the expression of AQP4 in the cortex of the rats were significantly increased. After NGF treatment, the water content and the expression of AQP4 in the brain tissue of the rats decreased significantly. The expression of tau protein GSK-3 尾 in rat brain was significantly higher than that in control group. The expression of tau protein GSK-3 尾 in UNGF TBI group was significantly lower than that in TBI group. The results showed that the protein and gene level of IL-1 尾 increased significantly after TBI in rats. The DNA binding activity of NF- 魏 B increased significantly, and the gene level of Bcl 2 decreased the level of caspase-3. The protein and gene level of IL-1 尾 decreased significantly after nasal administration of NGF. The DNA binding activity of NF- 魏 B decreased significantly. The level of Bcl-2 gene increased and the gene level of caspase-3 decreased. Therefore, nasal administration of NGF can reduce the brain edema associated with AQP4, decrease the phosphorylation of tau protein associated with GSK-3 尾, and improve the prognosis of TBI rats. Both of these effects are related to the reduction of inflammation related to Bcl-2 / caspase-3 / NF- 魏 B pathway after TBI by NGF.
【学位授予单位】：南京大学
【学位级别】：硕士
【学位授予年份】：2013
【分类号】：R651.15

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