miR-30c通过调节海马神经发生对APP/PS1转基因小鼠学习记忆能力的影响

目的：探讨miR-30c 通过调控海马神经发生对APP/PS1 转基因小鼠的学习记忆的影响。方法：用免疫荧光 检测Ki67 的表达和Morris 行为学检测APP/PS1 转基因小鼠海马的神经发生及学习记忆能力,并通过脑立体定位 技术显微注射miR-30c 过表达和miR-30c 敲减慢病毒载体以干扰海马齿状回区域的神经发生,检测海马神经发生 对学习记忆的影响。结果：APP/PS1 转基因小鼠的海马神经发生和学习记忆能力明显低于野生小鼠。然而,当 APP/PS1 转基因小鼠海马的miR-30c 过表达后,海马神经发生明显增加（14.2 倍）;相反,当海马miR-30c 敲减后, 海马神经发生明显降低（0.25 倍）,并且Morris 行为学结果显示,miR-30c 敲减后小鼠的学习记忆能力明显降低, 而miR-30c 过表达后小鼠的学习记忆能力明显增强。结论：miR-30c 可能通过调控海马神经发生影响学习记忆功能, 提示miR-30c 可作为干预阿尔茨海默症神经发生与学习记忆能力的潜在靶点。     

二氢杨梅素对APP/PS1转基因小鼠学习记忆和脑内自噬的影响

目的:研究二氢杨梅素(DHM)能否通过影响自噬对阿尔茨海默病(AD)模型小鼠发挥保护作用。方法:实验动物分为3组:7月龄的APP/PS1转基因痴呆模型小鼠随机分为AD组和DHM组,同月龄野生型(WT) C57BL/6J小鼠作为WT组,每组10只。DHM处理DHM组,同浓度DMSO稀释液处理AD组和WT组。采用Morris水迷宫实验检测各组学习记忆功能;免疫组织化学(IHC)、甲硫素S(ThS)染色以及ELISA法检测β-淀粉样蛋白(Aβ)的水平; Western Blot法检测自噬相关蛋白Beclin1、LC3、p62、Cathepsin D和LAMP1的表达水平;透射电镜检测脑内自噬结构微观变化。结果:与WT组相比,AD组小鼠在水迷宫检测中平均潜伏期、探索路径增长(P <0.01),穿越平台次数减少(P <0.01);脑内Aβ斑块增加(P <0.01),出现较严重的水肿,并发现了深染的晚期自噬体; LC3-Ⅱ/Ⅰ比值降低(P <0.01),p62、LAMP1和Cathepsin D表达增加(P <0.01); DHM处理后可以改善AD小鼠的学习记忆功能(...     

10月龄APP/PS1双转基因AD小鼠海马超微结构特征的观察

目的:观察APP/PS1双转基因小鼠海马内Aβ沉积的形态特征。方法:以10月龄雄性APP/PS1双转基因小鼠和C57BL/6小鼠为研究对象,采用Aβ免疫组化和透射电镜技术,观察海马部位Aβ的沉积情况及相关的病理特征。结果:在双转基因小鼠海马内存在致密性神经炎性斑块及营养障碍性突起,但脑微血管壁的Aβ沉积不甚明显;营养障碍性突起及神经细胞、微血管内皮细胞、周细胞内存在大量的自噬泡及次级溶酶体,提示该模型存在自噬溶酶体途径功能紊乱。结论:该鼠模拟了AD的老年斑及相关退行性变等病理改变,其中可能伴有细胞多种代谢的变化及血脑屏障功能的变化。从老年斑角度而言,该动物是成功的阿尔茨海默病动物模型,可用于AD的生化、病理研究及新治疗方法的探索。     

APP/PS1双转基因小鼠大脑皮质老年斑的病变对其学习记忆能力的影响

β-淀粉样蛋白(Aβ)沉积形成的老年斑是Alzheimer’s disease(AD)的主要病理特点,学习记忆功能障碍是其行为学的重要变化。为探讨老年斑对学习记忆的影响,本实验用17、28、44周C57系APP/PS1双转基因小鼠各4只,同龄野生型C57小鼠做对照,用Y-型迷宫方法比较不同年龄的AD模型鼠和C57鼠学习记忆能力的差异,利用免疫组织化学方法检测小鼠大脑皮质老年斑的病理变化,并比较AD模型鼠脑内老年斑的病变与其学习记忆能力改变之间的关系。结果显示:17周和28周AD模型鼠的学习记忆能力的各测试指标与对照组相应指标间均相差不显著(P>0.05),而44周AD模型鼠的学习记忆能力明显下降,各项指标与对照组相比具有显著性(P<0.01)。17周AD模型鼠脑切片上,可见大脑皮质内有少量老年斑的形成,而44周AD鼠皮质内斑块数量明显增多,体积明显增大。本结果提示老年斑的病变在一定程度上影响了其行为学的改变,这可能与β-淀粉样蛋白沉积继发的神经元缺失密切相关。     

APP/PS1双转基因AD小鼠早期内质网应激诱导的凋亡

目的观察APP/PS1双转基因AD小鼠神经细胞凋亡,及内质网分子伴侣葡萄糖调节蛋白（GRP78）和内质网促凋亡因子半胱氨酸蛋白酶-12（Caspase-12）表达的改变,探讨APP/PS1双转基因AD小鼠早期内质网应激诱导的凋亡。方法选取5、7月龄的APP/PS1双转基因小鼠和同月龄同背景的野生型小鼠（WT）,分为5月龄WT组、5月龄APP/PS1组、7月龄WT组和7月龄APP/PS1组,每组6只,应用原位细胞凋亡检测法（TUNEL）检测凋亡细胞,免疫组织化学方法检测其脑内GRP78和Caspase-12的表达水平。结果 TUNEL检测凋亡率分别为7月龄APP/PS1鼠（35.0±6.31）%、5月龄APP/PS1鼠（9.0±2.78）%、7月龄WT鼠（4.0±1.89）%、5月龄WT鼠（4.0±1.83）%,其中7月龄APP/PS1鼠凋亡率显著升高（P〈0.05）;免疫组织化学检测GRP78阳性率分别为7月龄APP/PS1鼠（30.0±5.43）%、5月龄APP/PS1鼠（10.0±2.12）%、7月龄WT鼠（2.0±1.71）%、5月龄WT鼠（3.0±1.41）%,7月龄APP/PS1鼠GRP78表达明显升高（P〈0.05）;免疫组织化学检测Caspase-12阳性率分别为7月龄APP/PS1鼠（33.0±5.98）%、5月龄APP/PS1鼠（12.0±2.60）%、7月龄WT鼠（4.0±2.56）%、5月龄WT鼠（2.0±1.79）%,7月龄APP/PS1鼠Caspase-12表达明显升高（P〈0.05）。结论 7月龄的APP/PS1双转基因小鼠出现了内质网应激诱导的凋亡。本实验结果为临床AD早期预防和治疗提供了重要依据。     

HuD在N2aAPP细胞和APP/PS1转基因小鼠中的表达

目的研究RNA结合蛋白HuD在AD细胞和动物模型中的表达水平,探讨HuD与AD的关系;明确PI3K/AKT通路对神经细胞HuD表达的调控作用。方法免疫荧光检测HuD在C57BL/6小鼠脑组织中的表达水平;构建过表达APP的N2a APP细胞模型,Western blot检测HuD的表达情况;购买APP/PS1转基因小鼠,Western blot检测HuD在脑组织中的表达情况;应用LY294002处理3种神经细胞(N2a、SH-SY5Y和HT22),阻断PI3K/AKT通路,观察该通路对HuD的调控作用。结果 HuD在整个C57BL/6小鼠脑组织中均有表达,且在海马体的颗粒细胞尤为明显;在N2a APP细胞,HuD的表达水平明显低于对照组;APP/PS1转基因小鼠脑组织中的HuD水平也低于野生对照鼠;LY294002处理下调了HuD在神经细胞中的表达水平。结论 HuD在N2a APP细胞和APP/PS1转基因小鼠中均呈低表达;HuD在神经细胞中受PI3K/AKT通路的调控,为研究HuD与AD的关系及寻找新的AD诊断和治疗靶点提供了理论基础。     

外源性神经生长因子对APP/PS1转基因小鼠Wnt/β-catenin信号通路的调节

目的:探讨外源性神经生长因子(NGF)对APP/PS1转基因小鼠Wnt/β-catenin信号通路的调节。方法:鼻腔给予小鼠NGF治疗14周,应用免疫组织化学检测小鼠脑内β淀粉样蛋白(Aβ)老年斑的分布,应用免疫印迹检测小鼠脑内糖原合酶激酶3β(GSK3β)、p-GSK3β、β-连环蛋白(β-catenin)和p-β-catenin的表达。结果:Aβ免疫组织化学显色结果显示,与对照组小鼠大脑内的Aβ老年斑相比较,NGF治疗组转基因小鼠大脑内的Aβ老年斑数量减少了30.22%,沉积减少了35.85%,差异有统计学意义。免疫印迹结果显示,外源性NGF减少APP/PS1转基因小鼠脑内GSK3β的表达,增加β-catenin的表达。结论:外源性NGF可能通过抑制GSK3β的活性和增加β-catenin的活性,激活Wnt/β-catenin信号通路,对神经元起保护作用。     

链脲佐菌素对APP/PS1转基因小鼠脑内糖原合成酶-3α活性的影响

目的观察链脲佐菌素(streptozotocin,STZ)诱导的实验性糖尿病对APP/PS1转基因小鼠脑组织中糖原合成酶-3α(GSK-3α)蛋白活性的影响。方法 3月龄APP/PS1转基因小鼠腹腔内注射STZ建立实验性糖尿病模型。使用电子天平和便携式血糖仪定期检测小鼠体重和血糖的变化。应用免疫组织化学方法和western blotting方法检测AD转基因小鼠脑组织中p-GSK-3α和GSK-3α的蛋白表达。结果动物饲养20W后,与APP/PS1转基因小鼠比较,STZ组转基因小鼠体重下降(P<0.05),血糖则明显升高(P<0.01),提示糖尿病模型诱导成功。Western blotting结果显示,STZ组APP/PS1转基因小鼠脑内p-GSK-3α蛋白表达水平较APP/PS1转基因小鼠明显降低(P<0.01),而总GSK-3α蛋白没有变化(P>0.05),两者的比值p-GSK-3α/GSK-3α下降49%(P<0.01)。免疫组化染色显示,STZ组转基因小鼠大脑皮层神经元内p-GSK-3α阳性表达明显低于转基因小鼠。结论 STZ上调APP/PS1转基因小鼠脑组织内GSK-3α蛋白活性。     

丹酚酸B对APP/PS1小鼠学习记忆及氧化应激的作用及可能机制

目的 探讨丹酚酸B对APP/PS1转基因小鼠学习记忆能力、氧化应激水平及Nrf-2/HO-l信号通路的影响.方法 将7月龄的APP/PS1小鼠随机分为模型组、丹酚酸B低剂量组(30mg/kg)及丹酚酸B高剂量组(60 mg/kg),并取同龄C57BL/6小鼠作为对照组,每组10只,连续灌胃给药8周.Morris水迷宫观...     

Early temporal short-term memory deficits in double transgenic APP/PS1 mice

We tested single APP (Tg2576) transgenic, PS1 (PS1dE9) transgenic, and double APP/PS1 transgenic mice at 3 and 6 months of age on the acquisition of a hippocampal-dependent operant “differential reinforcement of low rate schedule” (DRL) paradigm. In this task mice are required to wait for at least 10 seconds (DRL-10s) between 2 consecutive nose poke responses. Our data showed that while single APP and PS1 transgene expression did not affect DRL learning and performance, mice expressing double APP/PS1 transgenes were impaired in the acquisition of DRL-10s at 6 months, but not at 3 months of age. The same impaired double transgenic mice, however, were perfectly capable of normal acquisition of signaled DRL-10s (SDRL-10s) task, a hippocampal-independent task, wherein mice were required to emit responses when the end of the 10-second delay was signaled by a lighting of the chamber. The age-dependent and early deficits of APP/PS1 mice suggest that the appetitive DRL paradigm is sensitive to the amyloid pathology present in double APP/PS1 mice, and that this mouse line represents a good model with which to study the efficacy of therapeutic strategies against Alzheimer's disease.     

Associative and motor learning in 12-month-old transgenic APP+PS1 mice

Doubly transgenic 12-month-old amyloid precursor protein and presenilins 1 (APP+PS1) mice (n=14) and littermate control mice (n=17) were tested on eyeblink classical conditioning-a task impaired in humans with Alzheimer's disease (AD). Mice were also tested on a motor learning task (rotorod) and on sensory tasks (prepulse inhibition [PPI] and acoustic startle). Transgenic mice had impaired motor performance on rotorod. Overall, APP+PS1 mice performed similarly to controls on both 500ms delay and 500ms trace eyeblink conditioning as well as on prepulse inhibition (PPI) and acoustic startle. However, within the transgenic group, cortical amyloid burden correlated significantly with decreased trace eyeblink conditioning. Moreover, cortical amyloid burden and hippocampal microglia activation correlated significantly with decreased PPI. These data suggest that only those transgenic mice with the most severe amyloid pathology exhibited deficits in hippocampus-dependent tasks. Transgenic mouse models of amyloid deposition differ from Alzheimer patients not only by the absence of major neuronal loss, but also by the general absence of severe impairments in eyeblink conditioning, except for mice with the greatest amyloid pathology.     

鹿茸多肽对APP/PS1双转基因小鼠Rho/ROCK通路的影响

目的 探讨鹿茸多肽(VAP)对APP/PS1双转基因小鼠Rho/ROCK通路的影响.方法 APP/PS1双转基因小鼠随机分为模型组和鹿茸多肽组,每组20只,另以同窝同性别阴性小鼠20只设立为对照组.鹿茸多肽组小鼠鹿茸多肽100 mg/kg灌胃给药,每天1次,连续28 d.治疗后水迷宫实验检测并记录小鼠逃避潜伏期和穿越平...     

Beneficial effect of ovocystatin on the cognitive decline in APP/PS1 transgenic mice

PurposeCystatin C plays an important role in the course of neurodegenerative diseases and has a beneficial effect through inhibiting cysteine proteases and amyloid-β aggregation. It also induces proliferation and autophagy. Cystatin isolated from chicken egg white, called ovocystatin, has been widely used in the medical and pharmaceutical research due to its structural and biological similarities to human cystatin C. The aim of this study was to assess the effect of administering ovocystatin on the development of dementia-specific cognitive deficits in APP/PS1 transgenic mice.Materials/methodsThe study was conducted on transgenic B6C3-Tg(APPswe,PSEN1dE9)85Dbo/Mmjax mice. Ovocystatin was administered to four-month-old transgenic (AD) and wild type (NCAR) mice in drinking water for 24 weeks (at a dose of 40 and 4 μg/ mouse). The locomotor activity and cognitive functions were determined using an actimeter and the Morris water maze test, respectively.ResultsThe results of the study indicate that ovocystatin has a beneficial effect on the cognitive functions in APP/PS1 transgenic mice. The strongest effects of ovocystatin were found in the group of AD mice, where ovocystatin was administered in drinking water at a dose of 40 μg/mouse (p < 0.05). Mice from the AD group swam statistically significantly further in the target zone during the trial in the Morris water maze compared to the AD (vehiculum) group (p < 0.05).ConclusionsThe obtained results encourage further research into the protective effect, which may be used as an adjuvant in the treatment of deteriorating cognitive functions.     

理气化痰降浊法经验方改善AD转基因小鼠学习记忆能力

目的:通过两种不同中药方剂温脾补肾法(方剂1)和理气化痰降浊法(方剂2)经验方对阿尔茨海默病(AD)APP/PS1转基因小鼠学习记忆能力影响的对照研究探讨两首中药方剂治疗AD的可能性。方法:24只AD小鼠随机分为对照组、方剂1组、方剂2组。中药方剂(生理盐水)灌胃前后使用Morris水迷宫分别行定位航行和空间搜索试验。结果:随着时间延长,三组实验小鼠的潜伏期均呈缩短趋势,但是仅有方剂2组用药前后的潜伏期时长具有显著性差异,并比对照组、方剂1组明显缩短。用药后,方剂2组小鼠游泳总距离缩短、平均速度变慢、在站台所在象限停留时间延长、穿越站台所在位置次数明显增多,均具有统计学意义。结论:方剂2理气化痰降浊法经验方具有改善AD转基因小鼠学习记忆能力的作用。     

Locus coeruleus degeneration exacerbates olfactory deficits in APP/PS1 transgenic mice

Neuronal loss in the locus coeruleus (LC) is 1 of the early pathological events in Alzheimer's disease (AD). Projections of noradrenergic neurons of the LC innervate the olfactory bulb (OB). Because olfactory deficits have been reported in early AD, we investigated the effect of induced LC degeneration on olfactory memory and discrimination in an AD mouse model. LC degeneration was induced by treating APP/PS1 mice with N-(2-chloroethyl)-N-ethyl-bromo-benzylamine (DSP4) repeatedly between 3 and 12 months of age. Short term odor retention, ability for spontaneous habituation to an odor, and spontaneous odor discrimination were assessed by behavioral tests. DSP4 treatment in APP/PS1 mice resulted in an exacerbation of short term olfactory memory deficits and more discrete weakening of olfactory discrimination abilities, suggesting that LC degeneration contributes to olfactory deficits observed in AD. Importantly, DSP4 treatment also increased amyloid β (Aβ) deposition in the olfactory bulb of APP/PS1 mice, which correlated with olfactory memory, not with discrimination deficits.     

Xanthoceraside modulates neurogenesis to ameliorate cognitive impairment in APP/PS1 transgenic mice

Neuronal loss is reported to be an important pathological process in Alzheimer’s disease (AD). Neurogenesis is a process of generation of new neurons to fill the neuronal loss. Xanthoceraside has been shown to attenuate the cognitive deficits in several AD animal models. However, little is known about the effect of xanthoceraside on neurogenesis in APP/PS1 transgenic mice. Thus, in this study, we investigated whether xanthoceraside can ameliorate learning and memory impairment by promoting NSCs proliferation and neuronal differentiation. The results suggested that xanthoceraside significantly ameliorated the cognitive impairment and induced NSCs proliferation and neuronal differentiation in APP/PS1 transgenic mice. Meanwhile, in vitro study revealed that xanthoceraside increased the size of NSCs and induced NSCs differentiation into neurons compared with amyloid beta-peptide (25–35) (Aβ_25–35) treatment. Furthermore, we found that xanthoceraside significantly increased the expression of Wnt3a and p-GSK3β, decreased the expression of p-β-catenin, and induced nuclear translocation of β-catenin in APP/PS1 transgenic mice. Furthermore, in vitro study found that the effect of xanthoceraside on inducing NSCs proliferation and neuronal differentiation were inhibited by Wnt pathway inhibitor Dickkopf-1 (Dkk-1). Our data demonstrated that xanthoceraside may promote the proliferation and differentiation of NSCs into neurons by up-regulating the Wnt/β-catenin pathway to fill the neuronal loss, thereby improving learning and memory impairment in APP/PS1 transgenic mice.     

骨髓基质细胞移植改善APP/PS1转基因小鼠的认知功能

目的探究骨髓基质细胞(Bone marrow stromal cells,BMSCs)移植治疗对APP/PS转基因的阿尔兹海默病(Alzheimer's disease,AD)模型小鼠认知功能障碍的改善作用。方法分离培养C57BL/6小鼠的BMSCs,诱导其分化为神经细胞,通过相差显微镜技术和免疫细胞化学荧光染色技术鉴定其表型与分化。通过显微注射技术,将绿色荧光蛋白(Enhanced green fluorescent protein,eGFP)转基因小鼠(生后3个月)来源的BMSCs注入APP/PS转基因小鼠(生后12个月)侧脑室内后,以荧光显微镜技术检测移植细胞的存活和整合情况,HE染色检测AD模型小鼠海马区萎缩的恢复程度并以Morris水迷宫试验与旷场试验检测移植治疗后小鼠认知功能障碍的改善情况。结果分离自C57BL/6小鼠的BMSCs在增殖培养基中贴壁生长,形态呈纺锤形,免疫表型CD271+/CD45-++,经神经分化诱导培养可形成β-tubulin-III神经元和GFAP星形胶质细胞。GFP转基因小鼠来源的BMSCs移植入APP/PS1转基因小鼠脑内可存活并与宿主海马区神经组织有效整合。移植治疗3周后宿主小鼠海马区面积显著增加(<0.001),水迷宫实验逃生潜伏期明显缩短(<0.01),旷场试验评分显著提高(<0.05)。结论骨髓基质细胞移植治疗可有效改善APP/PS1转基因的AD模型小鼠的认知功能障碍。