Kh DIC小鼠海马神经细胞线粒体自由基水平的变化及APP17肽的影响

目的 观察Kh DIC（KM hn／IDL JIMP CAMS）小鼠海马神经细胞线粒体自由基水平的变化及APP17肽（β-amyloid precursor protein，APP，319-335肽段）的影响。方法 2．5月龄的Kh DIC小鼠随机分为模型组和APP17肽治疗组，正常对照组采用月龄和性别相匹配的正常昆明（KM）小鼠。APP17肽治疗组给予皮下注射APP17肽，每只每次0．34g-g，每周3次；模型组和正常对照给予等量的生理盐水；25W后成模。应用水迷宫试验观察小鼠学习、记忆功能的变化，应用羟胺法和硫代巴比妥酸（TBA）比色法分别检测了海马神经细胞线粒体抗氧化酶活性及自由基水平。结果 ①水迷宫结果显示，模型组小鼠存在明显的学习和记忆功能障碍，其游完全程的时间和错误反应次数均较正常对照组增多（P〈0．05）；APP17肽治疗组小鼠的行为学障碍明显轻于模型组，其上述的水迷宫检测结果与正常对照组比较无显著性差异。②与正常对照组比较，模型组小鼠海马神经细胞线粒体超氧化物歧化酶（SOD）活性明显降低（P〈0．01），丙二醛（MDA）含量显著升高（P〈0．01）；APP17肽治疗组检测结果与正常对照组接近，SOD活性高于模型组（P〈0．01），而MDA含量低于模型组（P〈0．01）。结论 ①Kh DIC小鼠存在学习和记忆功能障碍；而且其海马神经细胞线粒体氧化应激反应明显加剧。②APP17肽具有清除自由摹和抑制氧化应激反应的作用，从而提高脑组织的抗氧化能力，保护学习、记忆功能。     

促红细胞生成素对阿尔茨海默病模型大鼠记忆能力及其超微结构的影响

目的探讨促红细胞生成素(EPO)对阿尔茨海默病(AD)样大鼠记忆能力和超微结构的影响。方法雄性Wistar大鼠应用Y迷宫进行空间定向学习和记忆训练5 d后,将达到学会标准的48只大鼠随机分为4组:正常对照组、生理盐水组、模型组、EPO治疗组,每组12只。其中生理盐水组双侧海马注射生理盐水各5μl,模型组和EPO治疗组双侧海马注射凝聚态Aβ各5μl。EPO治疗组从造模手术当天按5 000 IU/kg腹腔注射EPO,隔天1次。术后第10天Y迷宫法检测各组大鼠空间定向学习和记忆能力,透射电镜观察海马线粒体和突触结构的改变,使用免疫组化技术检测各组大鼠海马组织神经生长因子(NGF)蛋白的表达。结果 1与生理盐水组相比,模型组学习记忆能力均明显下降,Y迷宫作业的错误反应次数增多,尝试次数增多,全天总反应时间显著延长(P0.05);与模型组相比,EPO治疗组Y迷宫作业的错误反应次数减少,尝试次数减少,全天总反应时间缩短(P0.05);2透射电镜结果显示:正常对照组和生理盐水组神经细胞结构完整,线粒体膜、脊完整,神经轴突和神经突触结构完整;EPO治疗组大鼠海马线粒体和神经突触基本正常;模型组大鼠海马线粒体结构破坏,线粒体肿胀,线粒体脊断裂,结构紊乱,神经突触密度降低,突触膜增厚,突触间隙不清,突触小泡数量减少。3与生理盐水组及EPO治疗组相比,模型组NGF表达降低(P0.05)。结论 EPO注射可以减轻AD样大鼠记忆能力损伤,减轻Aβ对大鼠海马超微结构的破坏。EPO可以显著改善AD样大鼠的空间记忆能力,其机制可能与提高NGF蛋白表达有关。     

姜黄素对阿尔茨海默病小鼠的治疗作用

目的通过测定阿尔茨海默病(AD)小鼠脑组织中β-淀粉样(Aβ)和磷酸化tau(Ptau)蛋白的阳性表达强度、mRNA水平及脑组织细胞凋亡程度,探讨姜黄素对AD小鼠的治疗机制。方法选取野生C57BL/6和转基因APP/V717雌性小鼠作为研究对象,实验分为对照组(C57BL/6野生雌性小鼠,n=10)、AD模型组(APP/V717转基因雌性小鼠,n=10)和姜黄素治疗组(APP/V717转基因雌性小鼠,n=10),三组均给予正常饮食。对照组和模型组每天灌胃一次生理盐水0.1 ml,姜黄素治疗组每天灌胃一次姜黄素50 mg/kg,共治疗8 w。Morris水迷宫检测各组小鼠记忆能力;苏木素-伊红(HE)染色和Tunnel染色普通光学及荧光显微镜下观察各组小鼠脑组织神经细胞损伤和凋亡程度;免疫组织化学染色和实时荧光定量PCR法检测各组小鼠脑组织中Aβ和Ptau蛋白的阳性表达强度及mRNA水平。结果 Morris水迷宫检测姜黄素治疗组小鼠记忆能力较模型组明显提高(P0.05)。HE染色可见姜黄素治疗组小鼠脑组织神经细胞固缩坏死较模型组明显减轻(P0.05)。Tunnel染色可见姜黄素治疗组小鼠脑组织神经细胞凋亡数量较模型组明显减少(P0.05)。免疫组织化学染色和实时荧光定量PCR可见姜黄素治疗组小鼠脑组织中Aβ和Ptau蛋白的阳性表达强度及mRNA水平均较模型组明显降低(P0.05)。结论姜黄素治疗能够降低AD小鼠脑组织中Aβ和Ptau蛋白的阳性表达强度及mRNA水平,提高AD小鼠学习记忆能力并抑制脑组织细胞凋亡。     

莱菔硫烷对阿尔茨海默病模型小鼠认知功能的干预作用

目的探讨莱菔硫烷(SFN)对阿尔茨海默病(AD)模型小鼠认知功能的影响。方法健康成年C57BL/6小鼠,按体重随机分为3组,每组10只,雌雄各半。模型组与干预组小鼠连续13 w喂饲含铝水(铝浓度0.4 g/100 ml),并隔日1次皮下注射200 mg/kg D-半乳糖;同时干预组小鼠每日1次灌胃给予25 mg/kg SFN,模型组小鼠灌胃等量蒸馏水,并设立溶媒对照组。通过Morris水迷宫和旷场实验检测小鼠的神经行为学改变,采用尼氏染色方法观察小鼠海马CA1区细胞形态及数量的变化,运用透射电镜观察小鼠海马CA1区细胞超微结构的改变。结果水迷宫实验结果显示,模型组小鼠逃避潜伏期与对照组相比明显延长,穿越圆台所在位置的次数明显少于对照组(P0.05);干预组小鼠与模型组相比,逃避潜伏期缩短,穿越圆台所在位置的次数增多(P0.05)。旷场实验结果表明,模型组小鼠与对照组相比,跨格数减少,中央格停留时间明显延长(P0.05);而干预组小鼠与模型组相比,跨格次数明显增加(P0.05)。尼氏染色结果发现模型组小鼠海马CA1区神经细胞数量明显降低,细胞排列散乱,而干预组小鼠海马CA1区神经细胞同模型组相比数量较多,并且细胞排列较为整齐紧密。电镜结果显示模型组小鼠CA1区神经细胞核膜、线粒体及突触结构等受损,而干预组小鼠海马CA1区神经细胞超微结构未见明显异常。结论 SFN可明显改善AD模型小鼠的认知功能,对海马CA1区神经细胞具有保护作用。     

大蒜素对阿尔茨海默病小鼠学习记忆能力和脑组织抗氧化能力的影响

目的 观察大蒜素对阿尔茨海默病(AD)小鼠学习记忆能力和脑组织抗氧化能力的影响.方法　采用 D-半乳糖建立AD小鼠模型,大蒜素治疗组在造模的同时皮下注射大蒜素.通过行为学方法　,观察大蒜素对AD小鼠记忆能力的影响;测定各组脑组织中丙二醛(MDA)含量和超氧化物歧化酶(SOD)的活性;常规HE和镀银改良法染色观察小鼠海马及皮质的形态学改变.结果 模型组与对照组、大蒜素治疗组比较,小鼠记忆功能明显减退,神经细胞破坏明显,海马和皮质出现β-淀粉样蛋白沉积及老年斑;大蒜素治疗组和对照组的丙二醛(MDA)含量较模型组显著降低(P<0.05),SOD活性明显高于模型组(P<0.05).结论 大蒜素在清除自由基和抗氧化损伤方面有显著功效,可以有效改善AD小鼠空间学习记忆能力损害.     

远志皂苷对APP/PS1双转基因小鼠海马神经细胞线粒体的保护作用

目的观察远志皂苷对APP/PS1双转基因小鼠海马神经细胞线粒体功能的调控,探讨远志皂苷的神经保护作用机制。方法将APP/PS1双转基因小鼠分为模型组,远志皂苷低、中和高剂量组(18.5、37.0和74.0 mg·kg~(-1)·d~(-1)),选同月龄C57BL/6小鼠为空白对照组,采用免疫荧光检测海马神经细胞线粒体膜电位,流式细胞术检测细胞凋亡率,比色法等检测超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-PX)活力和丙二醛(MDA)含量。结果与空白对照组相比,模型组线粒体膜电位降低,神经细胞凋亡率增高,SOD,过氧化氢酶(CAT)和(GSH-PX)活性明显降低,MDA含量明显增高;与模型组相比,远志皂苷各剂量组的线粒体膜电位升高,神经细胞凋亡率下降,SOD,CAT和GSH-PX活性明显升高,MDA含量明显降低(P<0.05)。结论远志皂苷可能通过调控线粒体功能,发挥保护海马神经元的作用。     

APP17肽对早衰小鼠学习记忆功能和海马神经细胞线粒体膜电位的影响

目的观察 APP17肽(β-Amyloid precursop protem,APP;319～335肽段)对早衰小鼠 Kh DIC 学习、记忆功能及海马神经细胞线粒体膜电位的影响。方法 2.5月龄的 Kh DIC 小鼠随机分为模型组和 APP17肽治疗组,对照组采用月龄和性别相匹配的正常昆明(KM)小鼠。APP17肽治疗组给予皮下注射 APP17肽,每只每次0.34μg,每周3次;模型组和正常对照给予等量的生理盐水;25周后开始实验。应用水迷宫试验观察小鼠学习、记忆功能的变化.应用流式细胞技术分析海马神经细胞凋亡率和线粒体膜电位的变化。结果 (1)水迷宫结果显示,模型组小鼠存在明显的学习和记忆功能障碍,其第3d、4d、5d 游完全程的时间(98.06±27.53、93.18±30.31、81.65±28.35)s、和错误反应次数(6.11±1.76、5.71±2.05、5.23±2.13)均较对照组(76.00±19.69、69.71±24.12、52.93±21.23)和(4.59±2.09、3.41±2.29、2.35±1.11)增多(P<0.05):APP17肽治疗组小鼠的行为学障碍明显轻于模型组,其上述的水迷宫检测结果与对照组比较差异无统计学意义。(2)与对照组小鼠海马神经细胞线粒体膜电位(189.5±15.6)和凋亡发生率(2.8±0.9)比较,模型组线粒体膜电位(106.0±12.7)明显降低(P<0.01),凋亡发生率(7.6±1.5)显著增加(P<0.01);APP17肽治疗组检测结果与对照组接近,海马神经细胞线粒体膜电位(160.3±18.6)高于模型组(P<0.01),而凋亡发生率(5.5±1.7)低于模型组(P<0.05)。结论 Kh DIC 小鼠存在学习和记忆功能障碍;而且其海马神经细胞凋亡率增加、线粒体膜电位降低。提示海马神经细胞发生凋亡和线粒体膜电位下降可能参与脑老化的发生和发展;APP17肽具有部分改善这一病理过程的作用。     

蒺藜皂苷对老年痴呆小鼠脑组织中H2O2、CAT、GSH-Px含量及海马超微结构的影响

目的 研究蒺藜皂苷对凝聚态β-淀粉样肽(Aβ25～35)致衰老小鼠脑组织中过氧化氢(H2O2)、过氧化氢酶(CAT)、谷胱甘肽过氧化氢酶(GSH-Px)含量及海马超微结构的影响.方法 采用一次性小鼠右侧脑室注射Aβ25～35 3 μl造成AD小鼠模型,并于侧脑室注射后14 d测定各组小鼠脑组织中H2O2的含量、CAT、GSH-Px的活力并于光镜、电镜下观察小鼠大脑皮质及海马组织超微结构.结果 侧脑室注射Aβ25～35 3 μl后可引起小鼠脑组织内H2O2含量增加,同时使小鼠脑组织内CAT和GSH-Px活力显著降低,而连续蒺藜皂苷14 d治疗后,与模型组相比蒺藜皂苷150 mg/kg治疗组可显著降低H2O2含量、提高CAT和GSH-Px活性(P<0.05),蒺藜皂苷450 mg/kg治疗组可显著降低H2O2含量、提高CAT和GSH-Px的活性(P<0.01);光镜下观察结果显示正常对照组海马CA1区神经元密集、神经元结构正常;模型组神经元间隙加大,说明有轻度水肿,部分神经元核、质界限模糊,细胞嗜酸性增强;电镜下观察结果显示正常对照组神经元、神经纤维超微结构正常;模型组神经元轻度水肿,脂褐素样包含物增多,线粒体减少或消失;蒺藜皂苷各治疗组可不同程度的减轻上述病理学改变.结论 侧脑室注射Aβ25～35可引起小鼠大脑自由基增加,海马超微结构改变,蒺藜皂苷能明显改善衰老小鼠大脑组织的超微结构并呈剂量效应关系.     

海风藤提取物对D-半乳糖脑老化模型小鼠行为学、海马神经细胞线粒体膜电位及凋亡的影响

目的研究海风藤提取物对D-半乳糖(D-gal)脑老化模型小鼠行为学、海马神经细胞线粒体膜电位及凋亡的影响。方法用D-gal复制小鼠脑老化模型,随机分为模型组和海风藤治疗组,正常对照组采用健康昆明种小鼠。海风藤组每天给予腹腔注射海风藤注射液1次,每次1ml/100g体重;模型组和正常对照组给予等量2.5%二甲基亚砜;10w后成模。应用水迷宫试验观察小鼠学习记忆功能的变化,应用流式细胞术分析海马神经细胞凋亡率和线粒体膜电位的变化。结果①水迷宫结果显示,模型组小鼠第3、4、5天游完全程的时间和错误反应次数均较正常对照组增多(P<0.05);海风藤组小鼠的行为学障碍明显轻于模型组(P<0.05),与正常对照组比较差异无统计学意义。②模型组小鼠海马神经细胞凋亡发生率显著增加(P<0.01)、线粒体膜电位明显降低(P<0.01);海风藤组海马神经细胞凋亡发生率低于模型组(P<0.01)、线粒体膜电位高于模型组(P<0.01);与正常对照组接近。结论D-gal脑老化模型小鼠存在学习和记忆功能障碍,且其海马神经细胞凋亡率增加、线粒体膜电位降低;海风藤能够明显改善该小鼠的学习和记忆能力,其作用机制可能是通过稳定线粒体膜电位、抑制凋亡的发生而实现的。     

饮高氟水小鼠脑海马回组织超微结构观察

目的 研究氟对中枢神经系统的毒性作用。方法 对饮含氟量为１００ｍｇ／Ｌ的高氟水６个月的小鼠脑海马回组织超微结构进行观察。结果 高氟组小鼠海马回神经细胞、神经突触、神经纤维及血脑屏障的超微结构均出现显著的病理变化。结论 氯对中枢神经系统具有直接的毒性作用。     

阿尔茨海默病转基因鼠模型的建立

目的 建立阿尔茨海默病(AD)双转基因鼠,为进一步研究AD发病机制提供较为理想的实验动物.方法 将人载脂蛋白E4转基因鼠和突变APP转基因鼠杂交.结果 经PCR初筛,对阳性小鼠基因组DNA作进一步的Southern杂交鉴定,获得2只双转基因小鼠,之后传代建系.结论 为进一步研究多个基因对AD的致病作用提供较为理想的实验动物.     

Melatonin alleviates behavioral deficits associated with apoptosis and cholinergic system dysfunction in the APP 695 transgenic mouse model of Alzheimer's disease

Melatonin is an endogenous antioxidant and free radical scavenger. A transgenic (Tg) mouse model for Alzheimer's disease mimics the accumulation of senile plaques, neuronal apoptosis and memory impairment. Previous studies indicated that melatonin reduced beta-amyloid (Abeta)-induced neurotoxicity. In this study, after giving melatonin at 10 mg/kg to APP 695 transgenic (APP 695 Tg) mice for 4 months, we evaluated the long-term influence of melatonin on behavior, biochemical and neuropathologic changes in APP 695 Tg mice. Step-down and step-through passive avoidance tests suggested that 8-month-old APP 695 Tg mice showed decreases in step-down latency and step-through latency and increases in count of error throughout the entire learning trial and memory session, which suggested learning and memory impairment. However, melatonin alleviated learning and memory deficits. Additionally, choline acetyltransferase (ChAT) activity also decreased in the frontal cortex and hippocampus of APP 695 Tg mice compared with non-Tg littermates. Melatonin supplementation increased ChAT activity in the frontal cortex and hippocampus. DNA fragmentation was present in the frontal cortex of the APP 695 Tg mice; melatonin reduced the number of apoptotic neurons. Congo Red staining and Bielschowsky silver impregnation both showed the apparent extracellular Abeta deposition in frontal cortex of APP 695 Tg mice. However, melatonin decreased the Abeta deposits. Our results indicate that neuroprotection by melatonin is partly related to modulation of apoptosis and protection of the cholinergic system. Early rational melatonin interventions may be one of the most promising strategies in the development of approaches to retard or prevent Abeta-mediated disease progression.     

突变型人APP基因与增强型绿色荧光蛋白基因融合载体的构建

目的 构建携带突变型人淀粉样前体蛋白基因(APP695)与增强型绿色荧光蛋白(EGFP)基因融合载体.方法 在NCBI上查找APP695的序列,设计引物.以pCB6质粒携带的野生型APP695序列为模板,以突变体引物扩增突变型APP695. 将突变型APP695与pIRES2-EGFP连接并测序证实.结果 经过酶切鉴定、PCR和DNA测序等方法,证实构建的pIRES2-EGFP/APP695突变型质粒载体序列正确.结论 成功构建APP695-EGFP融合基因载体,为研究AD发病的分子机制和治疗时的药物筛选奠定基础.     

Brain estrogen deficiency accelerates Abeta plaque formation in an Alzheimer's disease animal model

Much evidence indicates that women have a higher risk of developing Alzheimer's disease (AD) than do men. The reason for this gender difference is unclear. We hypothesize that estrogen deficiency in the brains of women with AD may be a key risk factor. In rapidly acquired postmortem brains from women with AD, we found greatly reduced estrogen levels compared with those from age- and gender-matched normal control subjects; AD and control subjects had comparably low levels of serum estrogen. We examined the onset and severity of AD pathology associated with estrogen depletion by using a gene-based approach, by crossing the estrogen-synthesizing enzyme aromatase gene knockout mice with APP23 transgenic mice, a mouse model of AD, to produce estrogen-deficient APP23 mice. Compared with APP23 transgenic control mice, estrogen-deficient APP23 mice exhibited greatly reduced brain estrogen and early-onset and increased beta amyloid peptide (Abeta) deposition. These mice also exhibited increased Abeta production, and microglia cultures prepared from the brains of these mice were impaired in Abeta clearance/degradation. In contrast, ovariectomized APP23 mice exhibited plaque pathology similar to that observed in the APP23 transgenic control mice. Our results indicate that estrogen depletion in the brain may be a significant risk factor for developing AD neuropathology.     

早老性痴呆转基因小鼠的研究

目的 建立早老性痴呆的转基因小鼠,为进一步的发病机理研究及药物筛选提供动物模型。方法 显微注射方法制备转基因小鼠,通过ＰＣＲ、Ｓｏｕｔｈｅｒｎ杂交鉴定。结果 质粒ｐＰｄＡＰ７５１的Ｔｔｈ１１１Ⅰ＋ＸｂａⅠ双酶切片段长度为４．３ｋｂ,经回收、纯化后,注射到小鼠受精卵的雄性原核,经ＰＣＲ,Ｓｏｕｔｈｅｒｎ杂交鉴定,确定首建鼠一只。首建鼠现已传代建系。结论 通过显微注射的方法,建立了早老性痴呆的转基因动     

前咽缺陷蛋白-1在APP/PS1转基因阿尔茨海默病小鼠模型中枢神经系统的分布及表达

目的 研究y-分泌酶组件蛋白APH-1(anterior pharynx defective 1,APH-1)在成年APP/PS1双转基因痴呆小鼠中枢神经系统(central nervous system,CNS)中的分布及表达,以及在阿尔茨海默病(Alzheimer's disease,AD)模型鼠和野生型小鼠脑中的表达差异,以进一步明确APH-1与AD的关系.方法 对APP/PS1双转基因AD模型种鼠交配后产下的子代进行基因分型,用免疫组化方法检测APH-1在成年APP/PS1双转基因痴呆模型小鼠CNS内的分布及表达,以及比较APH-1在出生后1d、7d、21 d、120 d的痴呆小鼠及同窝野生型小鼠脑内的表达和分布情况.结果 APH-1广泛分布于成年痴呆小鼠CNS的各区域,包括大脑皮质、海马、嗅脑、丘脑、纹状体、尾状核、中缝大核、小脑、脑干和脊髓等;出生1 dAD模型鼠与同窝野生型小鼠大脑皮质内APH-1均有高水平表达,而出生7d、21d表达水平较低,且成年小鼠脑中其表达量有所回升;各时间点痴呆小鼠脑内APH-1免疫阳性反应都比同窝野生型小鼠显著增强(P＜0.05).结论 APH-1在痴呆小鼠的CNS中广泛分布,但不同脑区分布和表达存在差异;且痴呆小鼠和正常野生型小鼠脑内APH-1分布和表达有区别,这些差异可能与AD的发生相关.     

PKCepsilon increases endothelin converting enzyme activity and reduces amyloid plaque pathology in transgenic mice

Deposition of plaques containing amyloid beta (Abeta) peptides is a neuropathological hallmark of Alzheimer's disease (AD). Here we demonstrate that neuronal overexpression of the epsilon isozyme of PKC decreases Abeta levels, plaque burden, and plaque-associated neuritic dystrophy and reactive astrocytosis in transgenic mice expressing familial AD-mutant forms of the human amyloid precursor protein (APP). Compared with APP singly transgenic mice, APP/PKCepsilon doubly transgenic mice had decreased Abeta levels but showed no evidence for altered cleavage of APP. Instead, PKCepsilon overexpression selectively increased the activity of endothelin-converting enzyme, which degrades Abeta. The activities of other Abeta-degrading enzymes, insulin degrading enzyme and neprilysin, were unchanged. These results indicate that increased neuronal PKCepsilon activity can promote Abeta clearance and reduce AD neuropathology through increased endothelin-converting enzyme activity.     

Study of Alzheimer's disease using transgenic mice: The mechanism of formation of brain Aβ amyloid and neurofibrillary tangles

Background:   The purpose of the present paper was to clarify the role of brain amyloid β protein (Aβ) in Alzheimer's disease (AD) with the eventual aim of developing new treatments.
Methods:   Three kinds of transgenic mice (Tg) were examined: those overexpressing βAPP695ΔNL (APPsw mice), those expressing the R406W mutant form of human 4-repeat tau (tauR406W mice), and those with both βAPP695ΔNL and R406W mutant tau (APPsw × tau R406 double Tg).
Results:   First, cored plaques, diffuse plaques and amyloid angiopathy were detected in the brains of APPsw mice. Loss of neurons and synapses was seen in the cored plaques, and accumulation of phosphorylated-tau in dystrophic neurites around the cored plaques. However, Neurofibrillary tangles were not seen. These mice displayed memory impairment and decreased levels of acetylcholine in the cortex and hippocampus. Second, tau R406W mice were produced. These mice developed motor disturbances and memory impairment, accompanied by gliosis and marked tau accumulation in neurons of the frontotemporal cortex, hippocampus and amygdala. No Aβ was detected. Third, APPsw × tauR406W double Tg were produced. The pattern of Aβ deposition was similar in these double Tg and the APPsw mice. Gallyas silver-staining showed greater levels of tauopathy in the hippocampus in these double Tg compared to the tauR406W mice.
Conclusion:   These findings suggest that deposition of brain Aβ is the initial event leading to subsequent disease such as accumulation of phosphorylated tau, neuronal loss and memory disturbance, and that the treatment of brain Aβ should be the first priority in finding a cure for AD.     

Melatonin treatment restores mitochondrial function in Alzheimer's mice: a mitochondrial protective role of melatonin membrane receptor signaling

Mitochondrial dysfunction is a hallmark of Alzheimer's disease (AD) and is observed in mutant amyloid precursor protein (APP) transgenic mouse models of familial AD. Melatonin is a potent antioxidant, can prevent toxic aggregation of Alzheimer's beta-amyloid (Aβ) peptide and, when taken long term, can protect against cognitive deficits in APP transgenic mice. To study the effects of melatonin on brain mitochondrial function in an AD model, APP/PS1 transgenic mice were treated for 1 month with melatonin. Analysis of isolated brain mitochondria from mice indicated that melatonin treatment decreased mitochondrial Aβ levels by two- to fourfold in different brain regions. This was accompanied by a near complete restoration of mitochondrial respiratory rates, membrane potential, and ATP levels in isolated mitochondria from the hippocampus, cortex, or striatum. When isolated mitochondria from untreated young mice were given melatonin, a slight increase in respiratory rate was observed. No such effect was observed in mitochondria from aged mice. In APP-expressing neuroblastoma cells in culture, mitochondrial function was restored by melatonin or by the structurally related compounds indole-3-propionic acid or N(1)-acetyl-N(2)-formyl-5-methoxykynuramine. This restoration was partially blocked by melatonin receptor antagonists indicating melatonin receptor signaling is required for the full effect. Therefore, treatments that stimulate melatonin receptor signaling may be beneficial for restoring mitochondrial function in AD, and preservation of mitochondrial function may an important mechanism by which long term melatonin treatment delays cognitive dysfunction in AD mice.     

Modeling Alzheimer's disease in transgenic mice: effect of age and of presenilin1 on amyloid biochemistry and pathology in APP/London mice

In transgenic mice that overexpress mutant Amyloid Precursor Protein [V717I], or APP/London (APP/Lo) (1999a. Early phenotypic changes in transgenic mice that overexpress different mutants of Amyloid Precursor Protein in brain. J. Biol. Chem. 274, 6483-6492; 1999b. Premature death in transgenic mice that overexpress mutant Amyloid precursor protein is preceded by severe neurodegeneration and apoptosis. Neuroscience 91, 819-830) the AD related phenotype of plaque and vascular amyloid pathology is late (12-15 months). This typical and diagnostic pathology is thereby dissociated in time from early symptoms (3-9 months) that include disturbed behavior, neophobia, aggression, glutamate excitotoxicity, defective cognition and decreased LTP. The APP/Lo transgenic mice are therefore a very interesting model to study early as well as late pathology, including the effect of age. In ageing APP*Lo mice, brain soluble and especially "insoluble" amyloid peptides dramatically increased, while normalized levels of secreted APPsalpha and APPsbeta, as well as cell-bound beta-C-stubs, remained remarkably constant, indicating normal alpha- and beta-secretase processing of APP. In double transgenic mice, i.e. APP/LoxPS1, clinical mutant PS1[A246E] but not wild-type human PS1 increased Abeta, and plaques and vascular amyloid developed at age 6-9 months. The PS1 mutant caused increasing Abeta42 production, while ageing did not. Amyloid deposits are thus formed, not by overproduction of Abeta, but by lack of clearance and/or degradation in the brain of ageing APP/Lo transgenic mice. The clearance pathways of the cerebral amyloid peptides are therefore valuable targets for fundamental research and for therapeutic potential. Although hyper-phosphorylated protein tau was evident in swollen neurites around the amyloid plaques, neurofibrillary pathology is not observed and the "tangle" aspect of AD pathology is therefore still missing from all current transgenic "amyloid" models. Also the "ApoE4" risk for late onset AD remains a problem for modeling in transgenic mice. We have generated transgenic mice that overexpress human ApoE4 (2000. Expression of Human Apolipoprotein E4 in neurons causes hyperphosphorylation of Protein tau in the brains of transgenic mice. Am. J. Pathol. 156 (3) 951-964) or human protein tau (1999. Prominent axonopathy in the brain and spinal cord of transgenic mice overexpressing four-repeat human tau protein. Am. J. Pathol. 155, 2153-2165) in their neurons. Both develop a similar although not identical axonopathy, with progressive degeneration of nerves and with muscle wasting resulting in motoric problems. Remarkably, ApoE4 transgenic mice are, like the tau transgenic mice, characterized by progressive hyper-phosphorylation of protein tau also in motor neurons which explains the motoric defects. Further crossing with the APP/Lo transgenic mice is ongoing to yield "multiple" transgenic mouse strains to study new aspects of amyloid and tau pathology.