QQ客服 官方微博 反馈留言

题名	Cdk5介导的自噬参与MPTP的神经毒性作用
作者	吕莉
学位类别	硕士
答辩日期	2015-05
授予单位	中国科学院研究生院
授予地点	北京
导师	姚永刚
关键词	帕金森病 MPTP Cdk5 自噬 多巴胺神经
其他题名	The Cdk5-mediated autophagy is involved in MPTP-induced neurotoxicity
中文摘要	帕金森病（Parkinson’s disease）是第二大常见的神经退行性疾病，其病理特征是黑质致密区多巴胺神经元丢失和路易氏小体（Lewy body）的形成。目前，全球有六百多万人正遭受帕金森病的折磨。帕金森病患者主要表现出运动障碍，包括颤抖，运动迟缓、姿势反射丧失等。 1-甲基-4-苯基-1,2,3,6-四氢吡啶（1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine，MPTP）能够通过线粒体途径选择性的损伤黑质致密区多巴胺神经元，所以已在猴子、猫、狗和小鼠等哺乳动物中被广泛用于构建PD模型。有研究表明，神经毒素MPTP的代谢产物MPP+通过多巴胺转运蛋白进入多巴胺神经元，在线粒体中积累并抑制线粒体复合物I活性，造成活性氧升高（reactive oxygen species， ROS），线粒体膜电位崩溃，三磷酸腺苷（adenosine triphosphate，ATP）消耗，最终导致多巴胺神经元损伤。然而，MPTP导致多巴胺损伤的具体分子机制和信号途径还需要深入研究。 自噬（Autophagy）是一种细胞自身降解细胞器和错误折叠的蛋白的过程。研究表明，自噬与帕金森病相关，然而自噬在帕金森病中的具体作用机制还不清楚。为了研究自噬在帕金森病中的分子作用机制，我们首先检测MPTP处理的C6细胞中的ROS水平和线粒体复合物I的活性。结果发现，MPTP通过线粒体功能异常（包括ROS水平升高，线粒体复合物I活性降低），进而激活自噬并导致Cdk5水平升高。为了进一步验证MPTP通过激活自噬引起神经元损伤，我们用MPTP构建了帕金森病小鼠模型。我们观察到了帕金森小鼠中脑黑质致密区酪氨酸羟化酶（tyrosine hydroxylase，TH）阳性细胞数量减少和多巴胺神经元树突密度降低。与细胞水平上的结果一致，我们发现伴随TH降低，Cdk5表达增强，自噬活性异常升高。在行为上，MPTP构建的帕金森病小鼠行为出现运动迟缓，表明MPTP主要损伤了黑质致密区多巴胺神经元，进而造成小鼠出现类似帕金森病患者的运动迟缓等行为。进一步为了研究MPTP诱导的自噬是否由Cdk5介导，我们分别在细胞水平和动物水平敲低Cdk5，观察到MPTP无法诱导自噬活性升高，并且能够拯救小鼠中脑黑质致密区多巴胺神经元损伤和运动迟缓。此外，我们还发现MPTP诱导的Cdk5介导的自噬可能是Atg7依赖的。 综上所述，本研究从细胞水平出发，深入探索MPTP是否通过Cdk5介导自噬发挥毒性作用，揭示自噬在帕金森病中的作用机制，最终回到整体动物水平初步探讨抑制Cdk5介导的自噬参与MPTP导致的神经毒性。因此，我们的研究有望为阐释自噬在帕金森病中的作用机制以及为临床治疗提供新的理论依据。
英文摘要	Parkinson’s disease (PD) is the second most common neurodegenerative disease. The pathologic features of PD are the loss of dopaminergic (Daergic) neurons in the substantia nigra pars compacta (SNc) and formation of Lewy bodies. More than six million people around the world are affeceted by PD. Patients are characterized by motor symptoms including tremor, bradykinesia and postural instability. 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) produces parkinsonain syndromes in several mammalian species, including monkeys, cats, dogs and mice. MPTP leads to selective degeneration of the Daergic neurons in the SNc through mitochondrial pathway. MPTP is converted into MPP+ which is taken up via the DA transporter (DAT) and accumulates in mitochondria of dopaminergic neurons. MPP+ inhibits Complex I of the electron transport chain, thereby causing the increased reactive oxygen species (ROS) and reducing ATP generation, inducing loss of dopaminergic neurons. Further studies are essential to elucidate the mechanisms of progressive dopamine neuron loss induced by MPTP. Autophagy, a self-degradative process, remove damaged organelles and misfolded proteins. Autophagy is involved in the pathogenesis of PD. However, little is known about the regulation of autophagy in PD. To explore the molecular mechanism of autophagy in PD, we examined ROS level and mitochondrial Complex I activity in C6 cells with MPTP treatment. The results showed that MPTP was able to induce mitochondrial dysfunction (including increased ROS level and decreased mitochondrial Complex I). We found an excessive activation of autophagy and increased level of Cdk5 in C6 cells induced by MPTP. A mouse PD model was used to discern whether MPTP induces autophagy to damage dopaminergic neurons in vivo. We found that MPTP treatment significantly reduced the number of TH-positive cells and dendrites of dopaminergic neurons in the SNc area of mice. In accordance with the results observed in vitro, we found decreased protein level of tyrosine hydroxylase (TH) and increased levels of Cdk5 and autophagy. The induced dyskinesia by chronic MPTP treatment suggested that MPTP results in selective degeneration dopaminergic neuronal loss. To further identify the potential roles of Cdk5 in regulation of MPTP-induced autophagy, we used Cdk5 siRNA and overexpression of Cdk5. In addition, we evaluated the effect of Cdk5 inhibition by using intracerebroventricular injection of siRNA oligonucleotide that targeted the Cdk5 gene in vivo. We found that knockdown of Cdk5 reduced MPTP-induced autophagy. Cdk5 inhibition in mice significantly attenuated the dopaminergic deficit and rescued dyskinesia. Furthermore, we found that Cdk5-mediated autophagy by MPTP might be Atg7-dependent. In summary, we found that an excessive activation of autophagy induced by MPTP, which is mediated by Cdk5 in both cellular and animal models. MPTP-induced autophagic activity could be salvaged by Cdk5 knockdown, which finally protected neuron function and prevented the behavioral abnormality.
语种	中文
内容类型	学位论文
源URL	[http://159.226.149.26:8080/handle/152453/10091]
专题	昆明动物研究所_重大疾病机理的遗传学
推荐引用方式 GB/T 7714	吕莉. Cdk5介导的自噬参与MPTP的神经毒性作用[D]. 北京. 中国科学院研究生院. 2015.

个性服务

查看访问统计

相关权益政策

暂无数据

收藏/分享

除非特别说明，本系统中所有内容都受版权保护，并保留所有权利。