一种新型抗糖尿病药物对链脲菌素诱导的AD动物模型认知障碍以及病理变化的神经保护作用研究

发布时间：2018-12-06 10:21

【摘要】：目的:探究新型GLP-1/GIP双受体激动剂DA-JC4是否能够改善STZ诱导的阿尔兹海默大鼠模型的工作以及空间学习记忆能力的伤害,是否能缓解tau蛋白的高度磷酸化,是否具有抗凋亡和抗炎症作用,并进一步探讨了DA-JC4的神经保护机制是否与脑内胰岛素信号通路相关。方法:1.大鼠被随机分为四组,1)对照组:侧脑室注射人工脑脊液+腹腔注射生理盐水;2)DA-JC4组:侧脑室注射人工脑脊液+腹腔注射DA-JC4;3)STZ组:侧脑室注射STZ+腹腔注射生理盐水;4)STZ+DA-JC4组:侧脑室注射STZ+腹腔注射DA-JC4。脑部手术第二天起,每天腹腔注射10nmol/kg DA-JC4,连续2周,随后通过Y迷宫和Morris水迷宫分别检测各组大鼠的工作工作以及空间学习记忆能力。2.采用免疫组织化学技术,观察了各组大鼠脑内的tau蛋白磷酸化水平以及GFAP阳性星型胶质细胞和IBA1阳性小胶质细胞数量变化。3.采用免疫印记法,检测了各组大鼠脑内皮层海马部位的凋亡相关分子,如BAX和Bcl-2的蛋白表达水平,以及胰岛素信号转导通路的相关分子,如IRS1和Akt的蛋白表达水平。结果:1.Y迷宫以及Morris水迷宫实验结果显示:与对照组相比,侧脑室注射STZ大鼠的自发交替行为、空间探索行为等记忆行为评分显著降低,治疗组给予DAJC4后大鼠的记忆行为评分明显得到改善。2.STZ组大鼠脑内皮层海马部位的tau蛋白磷酸化水平明显高于对照组。经DA-JC4治疗后,STZ+DA-JC4组大鼠脑内的tau蛋白磷酸化水平明显低于STZ组。3.STZ组大鼠脑内皮层海马部位的GFAP阳性星型胶质细胞和IBA1阳性小胶质细胞数量明显高于对照组。经DA-JC4治疗后,STZ+DA-JC4组大鼠脑内的GFAP阳性星型胶质细胞和IBA1阳性小胶质细胞数量明显低于STZ组。4.STZ组大鼠脑内皮层海马部位的促凋亡因子BAX与抗凋亡因子Bcl-2的蛋白表达水平比例明显高于对照组。经DA-JC4治疗后,STZ+DA-JC4组大鼠脑内的促凋亡因子BAX与抗凋亡因子Bcl-2的蛋白表达水平比例明显低于STZ组。5.STZ组大鼠脑内皮层海马部位的p-IRS1(Ser1101)蛋白表达水平明显高于对照组,p-Akt(Ser473)蛋白表达水平明显低于对照组。经DA-JC4治疗后,STZ+DA-JC4组大鼠脑内的p-IRS1(Ser1101)和p-Akt(Ser473)蛋白表达水平被部分逆转。结论:1.DA-JC4能显著改善STZ诱导的工作以及空间学习记忆能力的伤害,部分逆转认知下降。2.DA-JC4可明显降低脑内tau蛋白磷酸化水平,避免其进一步高度磷酸化,进而延缓AD病理特征-神经缠结的形成。3.DA-JC4能减少脑内GFAP阳性星型胶质细胞和IBA1阳性小胶质细胞数量,以此减轻STZ诱导的慢性炎症反应。4.DA-JC4通过调节促凋亡因子BAX与抗凋亡因子Bcl-2的蛋白表达,抑制STZ诱发的神经元凋亡。5.DA-JC4能下调p-IRS1蛋白表达水平,上调p-Akt蛋白表达水平,促使脑内胰岛素信号通路再次活化。
[Abstract]:Aim: to investigate whether DA-JC4, a novel GLP-1/GIP double receptor agonist, can improve the work of STZ induced Alzheimer rat model, damage the ability of spatial learning and memory, and alleviate the high phosphorylation of tau protein. Whether it has anti-apoptotic and anti-inflammatory effects, and further explore whether the neuroprotective mechanism of DA-JC4 is related to the insulin signaling pathway in the brain. Methods: 1. Rats were randomly divided into four groups: 1) control group: intraperitoneal injection of normal saline by intraperitoneal injection of artificial cerebrospinal fluid (CSF), 2) DA-JC4 group: intraperitoneal injection of DA-JC4; by intraperitoneal injection of artificial cerebrospinal fluid (DA-JC4;) 3) STZ group: intracerebroventricular injection of STZ intraperitoneal injection of normal saline, 4) STZ DA-JC4 group: lateral ventricle injection of STZ intraperitoneal injection of DA-JC4. From the second day of brain operation, 10nmol/kg DA-JC4, was injected intraperitoneally every day for 2 weeks, then the work and spatial learning and memory ability of each group were measured by Y maze and Morris water maze respectively. The changes of tau protein phosphorylation and the number of GFAP positive astrocytes and IBA1 positive microglias in the brain of rats were observed by immunohistochemical technique. 3. The expression levels of apoptosis-related molecules, such as BAX and Bcl-2, and related molecules of insulin signal transduction pathway, such as IRS1 and Akt, were detected by immunoblotting in hippocampus of rats. Results: 1. The results of Y maze and Morris water maze test showed that compared with the control group, the scores of memory behavior, such as spontaneous alternating behavior and spatial exploration behavior, were significantly decreased in rats injected with STZ in lateral ventricle. The memory behavior scores of the rats in the treatment group were significantly improved after DAJC4, and the level of tau protein phosphorylation in the hippocampus of the cerebral cortex in the 2.STZ group was significantly higher than that in the control group. After DA-JC4 treatment, the level of tau protein phosphorylation in the brain of STZ DA-JC4 group was significantly lower than that of STZ group, and the number of GFAP positive astrocytes and IBA1 positive microglia in hippocampus of 3.STZ group was significantly higher than that of control group. After DA-JC4 treatment, The number of GFAP positive astrocytes and IBA1 positive microglia in the brain of rats in STZ DA-JC4 group was significantly lower than that in STZ group. The protein of BAX and Bcl-2 in hippocampus of cerebral cortex in 4.STZ group was significantly lower than that in STZ group. The ratio of expression level was significantly higher than that of control group. After DA-JC4 treatment, The protein expression ratio of pro-apoptotic factor BAX and anti-apoptotic factor Bcl-2 in the brain of STZ DA-JC4 group was significantly lower than that of STZ group. The expression of p-IRS1 (Ser1101) protein in hippocampus of 5.STZ group was significantly higher than that of control group. The expression of p-Akt (Ser473) protein was significantly lower than that of the control group. After DA-JC4 treatment, the expression of p-IRS1 (Ser1101) and p-Akt (Ser473) in the brain of STZ DA-JC4 rats were partially reversed. Conclusion: 1.DA-JC4 can significantly improve the work induced by STZ and the damage of spatial learning and memory ability, and partially reverse cognitive decline. 2.DA-JC4 can significantly reduce the level of tau protein phosphorylation in brain and avoid its further high phosphorylation. 3.DA-JC4 can reduce the number of GFAP positive astrocytes and IBA1 positive microglia in the brain. 4.DA-JC4 inhibits the apoptosis of neurons induced by STZ by regulating the protein expression of BAX and Bcl-2. 5.DA-JC4 can down-regulate the expression of p-IRS1 protein. The expression of p-Akt protein was up-regulated and the insulin signaling pathway was reactivated.
【学位授予单位】：山西医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R965

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