培养晚期AD胞质杂交细胞线粒体功能的异常变化

目的探讨AD胞质杂交细胞在培养过程中的功能变化情况,为AD的线粒体发病机制提供实验依据。方法GSH检测试剂盒(比色法)检测细胞匀浆中GSH含量,COX活性检测试剂盒(分光光度计法)测定COX活性,JC-1染色后流式细胞术和激光共聚焦显微镜术检测线粒体膜电位的变化。结果培养晚期AD胞质杂交细胞的GSH含量、COX活性、膜电位水平明显低于培养早期AD胞质杂交细胞,差异有统计学意义(t=8.048,P<0.001;t=12.758,P<0.001和t=2.751,P<0.05),也显著低于培养晚期CTL胞质杂交细胞(t=23.06,P<0.001;t=26.862,P<0.001和t=3.876,P<0.01)。结论AD胞质杂交细胞随着培养时间的延长,其线粒体功能显著降低,这是由于AD患者本身线粒体基因的缺陷所至。     

线粒体DNA缺失细胞(ρ~0细胞)的构建与鉴定

目的探讨构建线粒体缺失细胞(ρ0细胞)的方法,为后继研究提供细胞模型。方法采用5μg/mL和7.5μg/mL两种浓度溴化乙锭持续作用于SH-SY5Y细胞构建线粒体缺失细胞,透射电镜观察和PCR法鉴定。结果 SHSY5Y细胞经过7.5μg/mL溴化乙锭持续作用95d后可见细胞线粒体肿胀变圆,基质消失,变成只剩外壳的"鬼影样"结构,线粒体嵴极少且变形;mtDNAPCR扩增未见目的条带。结论成功构建线粒体缺失细胞,为后继研究奠定细胞模型基础。     

线粒体表观遗传调控与阿尔茨海默病的研究进展

正线粒体表观遗传调控(mitoepigenetic regulation)是指对线粒体基因组编码基因的表观遗传学修饰调控,可引起线粒体基因组编码基因表达的改变,致使线粒体功能异常,导致多种疾病的发生。近年研究表明,线粒体表观遗传调控与阿尔茨海默病(Alzheimer disease,AD)发病机制密切相关。AD是一种中枢神经退行性疾病,典型神经病理变化是β-淀粉样蛋白(amyloid-β,Aβ)沉淀形成老年斑和神经原纤维     

线粒体功能障碍与阿尔茨海默病关系的研究进展

阿尔茨海默病是老年痴呆症中最常见的类型,其发病机制十分复杂。线粒体是动态细胞器,参与能量代谢、氧化应激,细胞凋亡等调节过程。β淀粉样蛋白、Ta u及氧化应激均能够导致线粒体功能损伤,引起线粒体功能障碍,并因此造成突触受损,神经细胞凋亡,促进阿尔茨海默病程的进展。     

淋巴瘤细胞与网织红细胞融合——胞质杂交细胞基因产物血红蛋白表达的观察

本实验选用富含有血红蛋白特征面无核的正常小鼠网织红细胞,与小鼠淋巴瘤细胞株融合,对杂交细胞基因产物血红蛋白进行联苯胺组织化学染色、荧光抗体和生化测定。结果与电镜和流式细胞光度计的实验结果一致,表明珠蛋白基因产物血红蛋白住杂交细胞中,能够表达并成为融合细胞的标志。联苯胺阳性细胞数在HAT选择性培液筛选后数量增加。BW×R胞质杂交细肥在传代过程中,仍继续出现血红蛋白。文中对杂交细胞血红蛋白表达及其在传代过程中,在一部分细胞中丢失的可能机理进行了讨论。     

β-淀粉样肽对线粒体的损伤及其在阿尔茨海默病中的作用

阿尔茨海默病(Alzheimer′s disease,AD)是以老年斑(senile plaque,SP)和神经纤维缠结(neurofibrillary tangles,NFTs)为主要病理特征的中枢神经系统退行性疾病,其病因及发病机制至今仍不明确.AD的病变产物β-淀粉样肽(amyloid-βpeptide,Aβ)在线粒体内沉积导致线粒体功能障碍,如ATP产生减少、氧化应激增强、细胞凋亡增强以及线粒体分裂/融合异常等,进而引起AD的一系列病理变化.     

阿尔茨海默病转基因小鼠海马结构NIX的变化

目的观察阿尔茨海默病转基因小鼠海马结构NIX的变化。方法以Morris水迷宫检测野生型和突变型转基因小鼠学习记忆能力,免疫组织化学和共聚焦激光扫描显微技术观察转基因小鼠海马结构促凋亡蛋白NIX的变化结果野生型和突变型小鼠逃避潜伏期中位数分别为29.00 s和38.00 s,差异无统计学意义,P>0.05;野生型和突变型小鼠搜索策略相比,突变型较野生型使用的搜索策略减少,差异有统计学意义,P<0.05;野生型和突变型小鼠NIX免疫反应阳性物灰度值中位数分别为103.83和128.85,差异有统计学意义,P<0.05;野生型和突变型小鼠海马结构NIX平均荧光强度分别为92.18±7.81和103.07±14.94,差异有统计学意义,P<0.05;野生型和突变型小鼠海马结构NIX与线粒体共定位的数目分别为240.94±169.48和544.18±336.44,差异有统计学意义,P<0.05。结论阿尔茨海默病转基因小鼠出现学习记忆障碍,海马结构促凋亡蛋白NIX的量增多,且NIX与线粒体共定位的量增多,提示NIX在阿尔茨海默病病理改变过程中可能起到一定的作用。     

喜树碱诱导Jurkat细胞线粒体膜电势和线粒体质量变化的研究

目的： 研究喜树碱（camptothecin，CPT）诱导Jurkat细胞凋亡过程中线粒体膜电势和线粒体质量的变化。 方法： 用喜树碱处理Jurkat细胞，利用Annexin V-FITC/PI双染流式细胞术研究细胞早期凋亡, PI染色流式细胞术测细胞周期, Annexin V-PE/DiOC6(3) 双染流式细胞术检测线粒体膜电势（△ψm），NAO染色流式细胞术检测线粒体质量。 结果： 在10 μmol·L-1 CPT诱导下，6 h 时Jurkat 细胞早期凋亡的细胞比率（22.59±1.04）%显著高于对照组（3.93±0.73）%（P<0.01）。CPT组坏死比率（2.48±0.53）%与对照组（2.78±0.63）%无显著差异（P>0.05）；并可使细胞出现明显的凋亡峰。晚期凋亡的细胞比率为（13.58±0.97）%显著高于对照组（3.18±0.51）%（P<0.01），CPT组G0/G1期细胞比率（48.14±0.96）% ，明显高于对照组（44.09±0.43）%（P<0.01）。CPT组线粒体发生明显去极化现象，AnnexinV+DiOC6(3)-的细胞比率为（19.47±0.69）%，而对照组比率为（4.21±0.40）%，差异显著（P<0.01）。同时，CPT组线粒体质量显著低于对照组：CPT组NAO+细胞比率为（74.77±1.66）%，对照组为（92.24±1.41）%（P<0.01）。 结论： CPT诱导Jurkat细胞凋亡过程中线粒体去极化作用增强并且线粒体质量下降，表明该凋亡过程与线粒体途径密切相关。     

线粒体与细胞凋亡

细胞凋亡是一种由基因控制的细胞自杀性死亡过程 ,近年来成为生物学、医学领域的一个热点。通过对凋亡机制深入的研究 ,人们发现线粒体的改变已不仅是凋亡的特征 ,还是凋亡调控的中心环节。     

紫外线对角质细胞线粒体功能及凋亡的影响研究

目的： 分析紫外线（UV）照射对HaCaT角质细胞系线粒体功能及凋亡的影响。 方法： 以紫外线低剂量（UVA2J/cm2，UVB10mJ/cm2）和高剂量（UVA6J/cm2，UVB30mJ/cm2）照射HaCaT细胞，继续培养15 h，用流式细胞仪检测线粒体膜电位（△ψm）、线粒体质量（mitochondrial mass）；使用碘化丙啶（PI）进行DNA染色并用流式细胞仪检测亚二倍体分析凋亡，以及进行Annexin V-FITC与PI共染色，用激光共聚焦显微镜观察细胞凋亡情况。 结果： HaCaT细胞经紫外线照射后，△ψm下降的细胞比例随着照射剂量的增加而增加，对照组、低剂量组及高剂量组分别为7.94%±1.02%、25.87%±4.55%、39.27%±5.32%；线粒体质量降低的细胞比例同样随着照射剂量的增加而增加，对照组、低剂量组以及高剂量组分别为15.19%±1.58%、40.36%±4.41%、68.79%±5.46%。用PI检测DNA含量所产生的亚二倍体峰观测凋亡率，对照组、低剂量组以及高剂量组凋亡率分别为1.82%±0.51%、30.16%±5.47%、58.49%±5.98%。用Annexin V-FITC/PI染色分析细胞凋亡程度，其结果与PI-DNA染色所得结果一致，对照组仅有少量细胞死亡，低剂量组凋亡细胞较对照组为多（Annexin V-FITC单阳性细胞），高剂量组以坏死细胞为多（Annexin V-FITC/PI双阳性）。 结论： HaCaT细胞经紫外照射后，线粒体去极化作用增强，同时线粒体质量降低，这些变化与细胞凋亡相关。     

线粒体与阿尔茨海默病

随着人类社会老龄化步伐的加快 ,老年人口所占比例逐渐增加 ,老年性痴呆的发病人数随年龄的增高而增多。阿尔茨海默病 (Ａｌｚｈｅｉｍｅｒ’ｓｄｉｓｅａｓｅ ,ＡＤ)作为老年性痴呆的一种重要类型 ,是中枢神经系统的一种渐进性退行性疾病 ,临床上以认知能力下降、记忆损害和痴呆为主要特征 ,神经病理上以脑细胞内神经纤维缠结 (ＮＦＴｓ)和细胞外老年斑 (ＳＰｓ)为主要特征 ,其中ＮＦＴ的主要成分为高度磷酸化的ｔａｕ蛋白 ,ＳＰｓ的主要成分为淀粉样蛋白前体 (ＡＰＰ)产生的β淀粉样肽 (Ａβ)。目前 ,ＡＤ的病因研究较多 ,其中线粒体因在…     

Alzheimer基因研究现状

阿尔茨海默病(AD)目前临床无客观性生化检测手段,仅靠精神量表,且常在临床症状出现后才能诊断。近年来,一系列新的基因研究有助于阿尔茨海默病的早期识别。这些基因的改变往往先于阿尔茨海默病的病理变化和临床表现,对其早期诊断有重要意义。本文就这些基因的研究现状作一综述。     

Alzheimer's disease: diverse aspects of mitochondrial malfunctioning

Alzheimer''s disease is a progressive neurodegenerative disorder, either assuming a sporadic, age-associated, late-onset form, or a familial form, with early onset, in a smaller fraction of the cases. Whereas in the familial cases several mutations have been identified in genes encoding proteins related with the pathogenesis of the disease, for the sporadic form several causes have been proposed and are currently under debate. Mitochondrial dysfunction has surfaced as one of the most discussed hypotheses acting as a trigger for the pathogenesis of Alzheimer''s disease. Mitochondria assume central functions in the cell, including ATP production, calcium homeostasis, reactive oxygen species generation, and apoptotic signaling. Although their role as the cause of the disease may be controversial, there is no doubt that mitochondrial dysfunction, abnormal mitochondrial dynamics and degradation by mitophagy occur during the disease process, contributing to its onset and progression.     

Age-related changes of mitochondrial structure and function in Caenorhabditis elegans

A number of observations have been made to examine the role that mitochrondrial energetics and superoxide anion production play in the aging of wild-type Caenorhabditis elegans. Ultrastructural analyses reveal the presence of swollen mitochondria, presumably produced by fusion events. Two key mitochondrial functions - the activity of two electron transport chain complexes and oxygen consumption - decreased as animals aged. Carbonylated proteins, one byproduct of oxidative stress, accumulated in mitochondria much more than in the cytoplasm. This is consistent with the notion that mitochondria are the primary source of endogenous reactive oxygen species. However, the level of mitochondrially generated superoxide anion did not change significantly during aging, suggesting that the accumulation of oxidative damage is not due to excessive production of superoxide anion in geriatric animals. In concert, these data support the notion that the mitochondrial function is an important aging determinant in wild-type C. elegans.     

Chromosome 12 and late-onset Alzheimer's disease

Alzheimer's disease is a complex neurodegenerative disorder, characterized by cognitive decline and distinctive neuropathology. Using large extended families with multiple affected, we found that three markers on chromosome 12 were linked with late-onset Alzheimer's disease. These markers were downstream from the gene for alpha-2 macroglobulin. It is likely that multiple genes will be identified either as risk factors or as causative agents for late-onset Alzheimer's disease.     

干细胞移植治疗阿尔茨海默病的研究进展

阿尔茨海默病是一种脑退行性疾病,目前药物对其治疗无明显效果,干细胞移植提供了一个新手段.     

Alzheimer's disease and Down's syndrome

The neuropathology of Down's syndrome at middle age is compared with that of Alzheimer's disease at that age, through a review of the published literature and from the author's personal observations. The pathological changes of Down's syndrome at middle age, i.e. the form and distribution of senile plaques and neurofibrillary tangles, and the pattern of involvement (atrophy) of neuronal systems are qualitatively the same as those of Alzheimer's disease at that age. Quantitative differences do occur and these may relate to biological or sociological variations inherent to the two parent populations. It is concluded that, in pathological terms, patients with Down's syndrome at middle age do indeed have Alzheimer's disease. Some ways in which a study of patients with Down's syndrome can give insight into the nature and development of the pathological changes of Alzheimer's disease are put forward and discussed.     

Cleavage and conformational changes of tau protein follow phosphorylation during Alzheimer's disease

Phosphorylation, cleavage and conformational changes in tau protein all play pivotal roles during Alzheimer's disease (AD). In an effort to determine the chronological sequence of these changes, in this study, using confocal microscopy, we compared phosphorylation at several sites (Ser(199/202/396/404/422)-Thr(205) and the second repeat domain), cleavage of tau (D(421)) and the canonical conformational Alz-50 epitope. While all of these posttranslational modifications are found in neurofibrillary tangles (NFTs) at all stages of the disease, we found significantly higher numbers of phospho-tau positive NFTs when compared with cleaved tau (P = 0.006 in Braak III; P = 0.002 in Braak IV; P = 0.012 in Braak V) or compared with the Alz-50 epitope (P < 0.05). Consistent with these findings, in a double transgenic mice model (Tet/GSK-3beta/VLW) overexpressing the enzyme glycogen synthase kinase-3beta (GSK-3beta) and tau with a triple FTDP-17 mutation (VLW) with AD-like neurodegeneration, phosphorylation at sites Ser(199/202)-Thr(205) was greater than truncated tau. Taken together, these data strongly support the notion that the conformational changes and truncation of tau occur after the phosphorylation of tau. We propose two probable pathways for the pathological processing of tau protein during AD, either phosphorylation and cleavage of tau followed by the Alz-50 conformational change or phosphorylation followed by the conformational change and cleavage as the last step.     

Chromosome fragility in Alzheimer's disease

We present cytogenetic findings for 12 patients with Alzheimer's Disease (AD) mean age 75.8 ± 6.01 years and 35 normal age and sex matched controls (mean age 74.8 ± 4.04 years). The study, undertaken due to reports of increased fragments and chromosome breakage in individuals with AD, was performed blind on coded peripheral blood specimens and the allocation of AD or control was not known to the cytogenetic staff until the end of the study. Both the AD group and the controls have been very carefully selected and both underwent the same clinical assessment and screening procedures which included CT scanning.
Chromosomes were analysed after 72 h cultures, using deprived medium TC199 which is known to enhance the appearance of fragile sites. A minimum of 50 cells was examined in each case and any rearrangement found was classified as ctg, csg, ctb, csb and the chromosome in which it occurred was recorded. Analysis of results showed that there was no statistically significant difference between the AD group and the controls for either the total occurrence of breaks, the type of aberration or the chromosome(s) involved. In both groups the commonest break was in 3p.