MPTP对小鼠空间学习记忆和脑内强啡肽免疫反应的影响

为了观察1 -甲基- 4 -苯基- 1, 2, 3, 6 四氢吡啶(MPTP)对C57BL/6小鼠空间学习记忆能力和脑内强啡肽(DYN)表达强度的影响,本研究给予小鼠皮下注射MPTP20mg/kg,连续8d,制备Parkinson病(PD)模型。与对照组相比,MPTP处理小鼠爬竿(P<0. 01)和游泳(P<0. 01)分数明显降低。Morris水迷宫实验结果显示:MPTP处理小鼠寻找平台潜伏期明显延长(P<0.01),在目标及对侧象限游泳时间所占百分比分别明显降低(P<0. 01)和增加(P<0. 01)。免疫细胞化学结果表明中脑黑质致密部的酪氨酸羟化酶(TH)免疫阳性神经元数目明显减少(P<0. 01)。同时在前额叶皮层(P<0. 01 )、背侧海马的CA1区(P<0. 05)和齿状回苔状纤维通路(P<0. 05)、背侧尾核(P<0. 01)的DYN免疫反应活性明显增强。本实验结果表明:用MPTP处理诱发的PD小鼠模型伴有空间学习和记忆能力的下降;并提示前额叶皮层、背侧海马的CA1区、齿状回苔状纤维通路和背侧尾核内DYN表达的增多可能与MPTP小鼠空间学习和记忆能力下降有关。     

艾司西酞普兰对MPTP处理的PC12细胞的保护作用及机制

目的 :探讨艾司西酞普兰(ESC)对1-甲基-4-苯基-1,2,3,6-四氢吡啶(MPTP)处理的PC12细胞的保护作用及其机制。方法 :用不同浓度ESC或/和不同浓度MPTP处理PC12细胞,再用3-甲基腺嘌呤(3-MA)抑制;通过MTS检测细胞存活率,Hoechst 33258染色观察细胞凋亡,免疫印迹检测自噬标记物LC3B-Ⅱ、LC3B-Ⅰ和凋亡蛋白Bcl-2、Bax的表达。结果 :经筛选得出,10μmol/L ESC预处理PC12细胞1 h,再加入1 000μmol/L MPTP处理24 h,细胞存活率显著提高;Hoechst 33258染色显示细胞核固缩明显减少;ESC+MPTP组LC3B-Ⅱ/Ⅰ、Bcl-2/Bax表达水平较MPTP组显著上升,但ESC+MPTP+3MA组LC3B-Ⅱ/Ⅰ、Bcl-2/Bax表达水平均显著下降。结论 :ESC对MPTP处理的PC12细胞有一定的神经保护作用,可能通过PI3K/Akt途径激发自噬作用,提高了Bcl-2的表达,抑制MPTP引发的细胞凋亡,从而发挥对其保护作用。     

虫草素对MPTP诱导的帕金森病小鼠运动与认知能力的影响

目的:探索虫草素(cordycepin)对1-甲基-4-苯基-1,2,3,6-四氢吡啶(1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine,MPTP)诱导的帕金森病小鼠运动与认知能力的影响。方法:将C57BL/6小鼠每天腹腔注射MPTP (30 mg/kg),连续8 d,以建立帕金森病小鼠模型。HE染色检测虫草素对MPTP诱导小鼠黑质致密部(sub-stantia nigra pars compacta,SNpc)细胞数目的影响;Western blot检测黑质(substantia nigra,SN)内酪氨酸羟化酶(tyrosine hydroxylase,TH)表达量的变化;采用旷场实验(open-filed test,OFT)、自发交替行为(spontaneous alterna-tion behavior,SAB)及水迷宫实验(water maze test,WMT)分别检测虫草素对帕金森病小鼠自发活动、情绪变化和认知行为的影响。结果:HE染色结果表明,虫草素显著抑制由MPTP引起的小鼠SNpc细胞数目减少(P 0.05);Western blot结果表明MPTP显著降低SN内TH的表达量(P 0.05),而虫草素可以逆转这种变化(P 0.05);虫草素可提高小鼠在OFT中的平均速度(P 0.05),并增加小鼠在SAB中的总进臂次数,提高正确率(P 0.05),但对WMT潜伏期并没有明显的影响。结论:虫草素能够减轻MPTP诱导的帕金森病小鼠早期的运动障碍和探索能力降低,但对其记忆障碍并没有明显的影响。     

线粒体转录终止因子蛋白家族的表达以及在MPTP/MPP~+诱导下的表达变化

目的探讨线粒体转录终止因子(MTERFs)蛋白家族在组织和细胞中的表达以及在神经毒素MPTP/MPP+诱导下的表达变化。方法取正常C57BL小黑鼠的脑、肝、肾组织,以及人神经母细胞瘤细胞SH-SY5Y、人心肌细胞HCM、人肝细胞L-02,检测MTERFs的蛋白表达情况;通过MPTP/MPP+构建帕金森病(PD)动物/细胞模型,检测MTERFs的蛋白表达变化情况。结果 MTERFs在小鼠脑中和SH-SY5Y细胞中蛋白表达水平较高;MPTP/MPP+诱导的PD模型中,MTERF3蛋白表达水平明显降低。结论在小鼠脑和SH-SY5Y细胞中,MTERFs表达较高;MPTP/MPP+诱导下MTERF3蛋白表达明显降低,影响了线粒体转录水平,这可能参与了帕金森病发生的过程。     

磷酸化P38 MAPK对帕金森病MPTP模型小鼠黑质caspase-3表达的影响

目的研究磷酸化P38 MAPK在1-甲基-4-苯基-1,2,3,6四氢吡啶(MPTP)所致帕金森病(PD)模型小鼠中对黑质半胱氨酸蛋白酶-3(caspase-3)的调控作用。方法将小鼠随机分为MPTP模型组,腹腔注射MPTP(30mg/kg,生理盐水溶);抑制剂组,在注射MPTP前1h腹腔注射SB203580(10mg/kg,溶于5mg/mLDMSO)。均1次/d,连续5d;对照组,注射与模型组和抑制剂组等量生理盐水和DMSO。观察行为学、免疫组织化学和免疫蛋白印迹法观察黑质酪氨酸羟化酶(TH)、caspase-3和磷酸化P38 MAPK(p-P38MAPK)的表达。结果与对照组相比,模型小鼠出现典型的PD症状,TH阳性神经元和蛋白水平分别下降约60%和65%(P<0.01),p-P38 MAPK、caspase-3阳性细胞及蛋白水平显著增加(P<0.01);经P38 MAPK抑制剂SB203580处理后,上述变化均显著减轻(P<0.01)。结论磷酸化P38 MAPK在MPTP诱导的PD小鼠黑质caspase-3表达中可能有重要调控作用,SB203580对PD小鼠具有一定的神经保护作用。     

帕金森病MPTP食蟹猴模型嗅球细胞凋亡

目的 建立1-甲基-4-苯基-1, 2, 3, 6-四氢吡啶（MPTP）帕金森病（PD）模型,探讨模型组嗅球细胞凋亡和胶质细胞增生情况。方法 3只成年健康食蟹猴,静脉注射MPTP建立PD模型,另3只静脉注射生理盐水作为对照。取出嗅球,免疫组织化学染色检测Caspase-3、Bcl-2、离子钙接头蛋白分子1（Iba-1）和胶质纤维酸性蛋白（GFAP）在食蟹猴嗅球中的表达情况,采用Image J v1.8.0软件分析模型组和对照组之间的差异。 结果 MPTP损伤后,与对照组相比,模型组嗅球突触小球层Caspase-3阳性细胞数明显增加,而Bcl-2表达减少;与对照组相比,模型组嗅球突触小球层和外网状层的GFAP和Iba-1阳性细胞数增加。 结论 MPTP可诱导食蟹猴嗅球突触小球层细胞凋亡,并伴有星形胶质细胞增生和小胶质细胞激活,这可能与帕金森病的功能障碍有关。     

p38MAPK抑制剂对MPTP模型小鼠黑质多巴胺能神经元的保护作用

目的:研究SB239063在1-甲基-4-苯基-1,2,3,6四氢吡啶(MPTP)所致帕金森病(PD)模型小鼠中抑制p38丝裂原激活蛋白激酶(mitogen-activated protein kinase,MAPK)活化对多巴胺(DA)能神经元的保护作用。方法:雄性C57BL/6N小鼠随机分为MPTP(30 mg/kg)模型组;SB239063(5 mg/kg)抑制剂组;SB239063(15 mg/kg)抑制剂组;SB239063(25 mg/kg)抑制剂组。抑制剂组于每次注射MPTP前3 h腹腔注射SB239063;对照组注射与模型组和抑制剂组等量生理盐水和DMSO。采用免疫组织化学和免疫蛋白印迹法观察各组小鼠黑质酪氨酸羟化酶(TH)和磷酸化p38蛋白激酶(p-p38 protein kinase,p-p38MAPK)之间表达变化的关系。结果:模型组小鼠黒质区p38MAPK显著活化,同时伴有TH阳性神经元明显丢失;SB239063 15 mg/kg与25 mg/kg组均可明显减少TH神经元丢失,而5 mg/kg组无显著影响;免疫荧光双标记结果显示p38MAPK与TH阳性神经元存在共表达。结论:p38MAPK对PD模型小鼠中脑黑质多巴胺能神经元丢失可能有重要调控作用,SB239063对多巴胺神经元具有一定的神经保护作用。     

MPTP诱导的帕金森病模型小鼠味觉偏好变化

目的:探讨1-甲基-4-苯基-1,2,3,6-四氢吡啶(MPTP)对帕金森病(PD)模型小鼠味觉的影响。方法:将雄性C57BL/6J小鼠分成对照组(control)和MPTP组,通过转棒、爬杆、嗅觉和悬尾实验检测小鼠行为,组织免疫荧光染色检测小鼠黑质致密部(SNpc)多巴胺(DA)能神经元。采用双瓶偏好实验检测小鼠对蔗糖、糖精、氯化钠及奎宁溶液的偏好率。利用real time RT-PCR检测小鼠舌上皮TAS1R1、TAS1R2和TAS1R3的mRNA表达的变化。结果:与control组相比,MPTP组小鼠转棒时间显著性降低,爬杆时间、找到隐藏食物的时间以及悬尾实验中静止时间均显著增加,免疫荧光结果显示MPTP处理可显著减少SNpc的DA能神经元数量。通过双瓶偏好实验发现MPTP可诱导小鼠对蔗糖、糖精、氯化钠的偏好率增加;但对苦味剂奎宁的偏好率没有影响。qRT-PCR分析发现MPTP抑制了小鼠舌上皮味觉受体TAS1R1和TAS1R2 mRNA的表达,而对TAS1R3 mRNA的表达没有影响。结论:MPTP诱导的PD模型小鼠对蔗糖、糖精、氯化钠的饮用偏好率增加,而对奎宁的偏好率没有影响,...     

多聚ADP-核糖聚合酶抑制剂对帕金森病小鼠相关脑区病理变化的影响

目的研究帕金森病(Park inson’s d isease,PD)小鼠黑质和纹状体多巴胺(dopam inergic,DA)能神经元数量和超微结构的变化及多聚ADP-核糖聚合酶(poly(ADP-ribose)polym erase,PARP)抑制剂PJ34的干预作用。方法采用1-甲基-4-苯-1,2,3,6-四氢吡啶(1-m ethyl-4-phenyl-1,2,3,6-tetrahydropyrid ine,MPTP)制备PD小鼠模型,并用PJ34进行干预,2h、24h、72h进行酪氨酸羟化酶(tyrosine hydroxylase,TH)免疫组化染色观察DA能神经元数量,透射电镜观察超微结构改变。结果与正常对照小鼠比较,PD小鼠黑质TH阳性神经元进行性减少,核膜皱缩,染色质凝聚成块并有边聚现象;纹状体TH阳性神经纤维稀疏,突触数量减少。与PD小鼠比较,PJ34干预组黑质TH阳性神经元明显增多,纹状体TH阳性神经纤维密度增加(P<0.01),细胞形态比模型组明显改善。结论PARP的活性改变在PD的发病过程中发挥重要作用,PARP抑制剂对DA能神经元有保护作用。     

MPTP损伤侧猴中脑黑质组织的基因表达

目的：采用mRNA差异显示技术（mRNA differential display,mRNA DD)寻找MPTP诱导和未诱导的猴中脑黑质组织差异表达的cDNA。方法：对诱导侧及未诱导侧的黑质mRNA用3′端3个锚定引物分别与5′端30种随机引物进行逆转录－聚合酶链反应（RT－PCR）扩增产物经聚丙烯酰胺凝胶电泳及硝酸银染色显示目的的条带。结果：获得一批表达差异的cDNA片段,对其克隆后作DNA序列测定,同源性比较分析发现,其中2个克隆为未知序列,已被GenBank接收,接收号为AF159161、AF159162。结论：MPTP诱导后黑质可能有效新基因的表达。     

Driving GDNF expression: the green and the red traffic lights

Glial cell line-derived neurotrophic factor (GDNF) is widely recognized as a potent survival factor for dopaminergic neurons of the nigrostriatal pathway that degenerate in Parkinson's disease (PD). In animal models of PD, GDNF delivery to the striatum or the substantia nigra protects dopaminergic neurons against subsequent toxin-induced injury and rescues previously damaged neurons, promoting recovery of the motor function. Thus, GDNF was proposed as a potential therapy to PD aimed at slowing down, halting or reversing neurodegeneration, an issue addressed in previous reviews. However, the use of GDNF as a therapeutic agent for PD is hampered by the difficulty in delivering it to the brain. Another potential strategy is to stimulate the endogenous expression of GDNF, but in order to do that we need to understand how GDNF expression is regulated. The aim of this review is to do a comprehensive analysis of the state of the art on the control of endogenous GDNF expression in the nervous system, focusing mainly on the nigrostriatal pathway. We address the control of GDNF expression during development, in the adult brain and after injury, and how damaged neurons signal glial cells to up-regulate GDNF. Pharmacological agents or natural molecules that increase GDNF expression and show neuroprotective activity in animal models of PD are reviewed. We also provide an integrated overview of the signalling pathways linking receptors for these molecules to the induction of GDNF gene, which might also become targets for neuroprotective therapies in PD.     

Apoptosis-inducing factor: a matter of neuron life and death

The mitochondrial flavoprotein apoptosis-inducing factor (AIF) is the main mediator of caspase-independent apoptosis-like programmed cell death. Upon pathological permeabilization of the outer mitochondrial membrane, AIF is translocated to the nucleus, where it participates in chromatin condensation and is associated to large-scale DNA fragmentation. Heavy down-regulation of AIF expression in mutant mice or reduced AIF expression achieved with small interfering RNA (siRNA) provides neuroprotection against acute neurodegenerative insults. Paradoxically, in addition to its pro-apoptotic function, AIF likely plays an anti-apoptotic role by regulating the production of reactive oxygen species (ROS) via its putative oxidoreductase and peroxide scavenging activities. In this review, we discuss accumulating evidence linking AIF to both acute and chronic neurodegenerative processes by emphasising mechanisms underlying the dual roles apparently played by AIF in these processes.     

Brain renin-angiotensin--a new look at an old system

The classic renin-angiotensin system (RAS) is described as a circulating hormone system focused on cardiovascular and body water regulation, with angiotensin II as its major effector. Detlef Ganten's discovery some years ago of an independent local brain RAS composed of the necessary functional components (angiotensinogen, peptidases, angiotensins and specific receptor proteins) significantly expanded the possible physiological and pharmacological functions of this system. This review first describes the enzymatic pathways resulting in active angiotensin ligands and their interaction with AT(1), AT(2) and AT(4) receptor proteins. We discuss the characterization and distribution of the AT(1) and AT(2) receptor subtypes and the current controversy over the identity of the AT(4) receptor subtype. Research findings favoring the candidates insulin-regulated aminopeptidase (IRAP) and the type 1 tyrosine kinase receptor c-Met, are presented. Next, we summarize current research efforts directed at the use of angiotensin analogues in the treatment of clinical disorders such as memory dysfunction, cerebral blood flow and cerebroprotection, stress, depression, alcohol consumption, seizure, Alzheimer's and Parkinson's diseases, and diabetes. The use of ACE inhibitors, and AT(1) and/or AT(2) receptor blockers, has shown promise in the treatment of several of these pathologies. The development of blood-brain barrier penetrant AT(4) receptor agonists and antagonists is of major importance regarding the continuing evaluation of the efficacy of new treatment approaches.