壳多糖酶-1在阿尔茨海默病模型中的表达及其作用机制的研究

发布时间：2018-06-04 22:37

本文选题：阿尔茨海默病 + 壳多糖酶-1　；参考：《重庆医科大学》2017年硕士论文

【摘要】：第一部分壳多糖酶-1在老年APP/PS1转基因鼠中的表达与认知水平相关目的:壳多糖酶-1(chitinase1,CHIT1)在阿尔茨海默病(Alzheimer’s disease,AD)中表达增加,然而CHIT1在AD中的具体作用尚不明确。本实验检测了22月龄APP/PS1双转基因鼠中CHIT1的表达水平,并结合本团队既往研究结果,探究CHIT1在AD中的意义。方法:实验组采用22月龄APP/PS1双转基因鼠,对照组选取与之相对应的同龄野生型小鼠(wild type,WT)。采用Morris水迷宫检测行为学变化,联合采用酶联免疫特异性测定法(enzyme-linked immunospecific assay,ELISA)和实时定量反转录酶-聚合酶链锁反应(quantitative real-time polymerase chain reaction,q RT-PCR)检测各组小鼠脑组织中CHIT1的表达水平。结果:与对照组相比,Morris水迷宫显示22月龄APP/PS1双转基因鼠逃避潜伏期明显延长,且在目标象限耗时百分比变短,在非目标象限耗时百分比增加。ELISA法和q RT-PCR显示22月龄APP/PS1鼠CHIT1表达水平较对照WT组明显增加。结论:随着22月龄APP/PS1双转基因鼠空间学习能力和空间记忆能力的下降,CHIT1表达水平升高。结合本团队前期关于4月龄、12月龄APP/PS1鼠的实验结果,AD早期(4月龄APP/PS1鼠)认知下降不突出,AD中晚期(12月龄、22月龄APP/PS1鼠)认知障碍明显,CHIT1在AD模型中随着认知功能下降而不断增高,提示CHIT1可作为评估AD疾病进展的标志物。第二部分壳多糖酶-1在阿尔茨海默病非转基因大鼠模型中的作用机制目的:文献报道壳多糖酶-1的活性在AD患者中明显增加,然而CHIT1在AD中的作用机制尚不明确。本实验旨在探究CHIT1对于AD病理改变以及小胶质细胞活性的影响。方法:本研究采用D-半乳糖和Al Cl3持续注入大鼠腹腔诱导的痴呆模型,并予以外源性的CHIT1和CHIT1抑制剂(chitinase-IN-2),分别检测CHIT1对于炎症因子(TNFα,IL-1β,Arg-1,MRC1/CD206)和Aβ寡聚体的影响,Morris水迷宫检测大鼠行为学改变。此外,进一步采用Aβ处理的N9小胶质细胞株以验证CHIT1对于炎症因子的影响是通过调节小胶质细胞活性实现的。结果:无论是D-galactose和Al Cl3诱导的AD动物模型还是Aβ处理的N9小胶质细胞株均可检测到CHIT1活性明显增高。经CHIT1处理的AD大鼠模型中,认知得到改善,同时,还可检测到抗炎因子(Arg-1,MRC1/CD206)表达水平增高、促炎因子(TNFa,IL-1β)表达水平降低。与之相对应地,在加入CHIT1的Aβ诱导的N9小胶质细胞株中,小胶质细胞M2抗炎状态的标志物表达增多,M1促炎状态标志物表达水平下降,表明CHIT1可以调节小胶质细胞活性,使其向M2抗炎状态转换。此外,我们还发现,CHIT1能够减少Aβ寡聚体在AD大鼠脑组织内的沉积。结论:CHIT1可以减少Aβ寡聚体沉积、促使小胶质细胞向M2抗炎形态转化,从而在AD中发挥保护作用。
[Abstract]:Part I the expression of chitosanase 1 in senile APP/PS1 transgenic mice and its cognitive level objective: the expression of chitinase 1 (CHIT1) in Alzheimer's disease (AD) is increased, but the specific role of CHIT1 in AD is not clear. This study examined the expression of CHIT1 in 22-month-old APP/PS1 transgenic mice and explored the significance of CHIT1 in AD. Methods: the 22 month old APP/PS1 double transgenic mice were used in the experimental group, and the corresponding wild type mice of the same age in the control group were selected. The behavioral changes were detected by Morris water maze, and the expression of CHIT1 in brain tissue of mice in each group was detected by enzyme linked immunospecific assay (Elisa) and real-time quantitative reverse transcriptase polymerase chain reaction quantitative real-time polymerase chain reactionQ (RT-PCRQ). Results: compared with the control group, Morris water maze showed that the escape latency of 22-month-old APP/PS1 transgenic mice was significantly prolonged, and the percentage of time spent in the target quadrant was shortened. The percentage of time consuming in non-target quadrant was increased. Elisa and Q RT-PCR showed that the expression of CHIT1 in 22-month-old APP/PS1 mice was significantly higher than that in control WT group. Conclusion: with the decrease of spatial learning ability and spatial memory ability of 22-month-old APP/PS1 transgenic mice, the expression level of CHIT1 increased. According to the results of our team's previous study on 4-month and 12-month-old APP/PS1 mice, cognitive decline was not significant in AD early and 4-month-old APP/PS1 rats.) Cognitive impairment was significantly increased with cognitive function decline in AD model. The results suggest that CHIT1 can be used as a marker to evaluate the progression of AD. The second part of the mechanism of chitosan enzyme-1 in Alzheimer's disease non-transgenic rats objective: it is reported that the activity of chitosan enzyme-1 is significantly increased in AD patients, but the mechanism of CHIT1 in AD is not clear. The aim of this study was to investigate the effects of CHIT1 on AD pathological changes and microglial activity. Methods: the dementia model was induced by continuous injection of D-galactose and Al Cl3 into the abdominal cavity of rats. The effects of CHIT1 on the inflammatory factor TNF- 伪, IL-1 尾, Arg-1MRC 1 / CD206) and A 尾 oligomer were measured by Morris water maze in rats. In addition, A 尾 -treated N9 microglia cell lines were further used to verify the effect of CHIT1 on inflammatory cytokines by regulating microglial activity. Results: both the AD model induced by D-galactose and Al Cl3 and the N9 microglia cell line treated with A 尾 could significantly increase the activity of CHIT1. In AD rat model treated with CHIT1, the cognition was improved, and the expression of anti-inflammatory factor Arg-1 / MRC1 / CD206 was also detected. In contrast, in A 尾 -induced N9 microglia cell line added with CHIT1, the expression of anti-inflammatory markers of M2 in microglia increased and the expression level of M1 pro-inflammatory markers decreased, suggesting that CHIT1 could regulate microglia activity. Make it to M2 anti-inflammatory state transition. In addition, we also found that CHIT1 can reduce the deposition of A 尾 oligomer in brain tissue of AD rats. ConclusionChIT1 can reduce A 尾 -oligomer deposition, promote microglia to M2 anti-inflammatory morphologic transformation, and thus play a protective role in AD.
【学位授予单位】：重庆医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R749.16;R-332

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