Formation of amyloid-like fibrils in COS cells overexpressing part of the Alzheimer amyloid protein precursor

A pathological hallmark of Alzheimer's disease is the deposition of amyloid fibrils in the brain. The principal component of the amyloid fibril is beta/A4 protein, which is derived from a large membrane-bound glycoprotein, Alzheimer amyloid protein precursor (APP). Although the deposition of amyloid is thought to result from the aberrant processing of APP, the detailed molecular mechanisms of amyloidogenesis remain unclear. A C-terminal fragment of APP which spans the beta/A4 and cytoplasmic domains has a tendency to self-aggregate. In an attempt to establish a cultured-cell model for amyloid fibril formation, we have transfected COS-1 cells with complementary DNA encoding the C-terminal 100 residues of APP. In the perinuclear regions of a small population of DNA-transfected cells, we observed inclusion-like deposits which showed a strong immunohistochemical reaction towards an anti-C-terminal APP antibody or an anti-beta/A4 amyloid core-specific antibody. Electron microscope observations of the inclusion-carrying cells revealed an accumulation of amyloid-like fibrils of 8-22 nm diameter near and on the nuclear membrane. The fibrils showed a beaded or helical structure, and reacted positively with the anti-C-terminus antibody by immunoelectron microscopy. These results suggest that the formation of amyloid fibrils is an inherent characteristic of the C-terminal peptide of APP. The present system provides a suitable model for the molecular dissection of the process of brain amyloidogenesis.     

Atomic structures of amyloid cross-beta spines reveal varied steric zippers

Amyloid fibrils formed from different proteins, each associated with a particular disease, contain a common cross-beta spine. The atomic architecture of a spine, from the fibril-forming segment GNNQQNY of the yeast prion protein Sup35, was recently revealed by X-ray microcrystallography. It is a pair of beta-sheets, with the facing side chains of the two sheets interdigitated in a dry 'steric zipper'. Here we report some 30 other segments from fibril-forming proteins that form amyloid-like fibrils, microcrystals, or usually both. These include segments from the Alzheimer's amyloid-beta and tau proteins, the PrP prion protein, insulin, islet amyloid polypeptide (IAPP), lysozyme, myoglobin, alpha-synuclein and beta(2)-microglobulin, suggesting that common structural features are shared by amyloid diseases at the molecular level. Structures of 13 of these microcrystals all reveal steric zippers, but with variations that expand the range of atomic architectures for amyloid-like fibrils and offer an atomic-level hypothesis for the basis of prion strains.     

Nanomechanics of individual amyloid fibrils using atomic force microscopy

Compression elasticity of glucagon amyloid fibrils in the transverse direction was investigated by a nanoindentation approach based on atomic force microscopy (AFM).With force-volume mapping, we obtained the correlations between radially applied force and compression of amyloid fibrils, from which the radial compressive elasticity can be deduced.The estimated elastic modulus at three typical locations of fibrils varied from (0.72±0.80) GPa to (1.26±0.62) GPa under small external forces, imply-ing the struct...     

A camelid antibody fragment inhibits the formation of amyloid fibrils by human lysozyme

Amyloid diseases are characterized by an aberrant assembly of a specific protein or protein fragment into fibrils and plaques that are deposited in various organs and tissues, often with serious pathological consequences. Non-neuropathic systemic amyloidosis is associated with single point mutations in the gene coding for human lysozyme. Here we report that a single-domain fragment of a camelid antibody raised against wild-type human lysozyme inhibits the in vitro aggregation of its amyloidogenic variant, D67H. Our structural studies reveal that the epitope includes neither the site of mutation nor most residues in the region of the protein structure that is destabilized by the mutation. Instead, the binding of the antibody fragment achieves its effect by restoring the structural cooperativity characteristic of the wild-type protein. This appears to occur at least in part through the transmission of long-range conformational effects to the interface between the two structural domains of the protein. Thus, reducing the ability of an amyloidogenic protein to form partly unfolded species can be an effective method of preventing its aggregation, suggesting approaches to the rational design of therapeutic agents directed against protein deposition diseases.     

Amyloid-like fibrils of ribonuclease A with three-dimensional domain-swapped and native-like structure

Amyloid or amyloid-like fibrils are elongated, insoluble protein aggregates, formed in vivo in association with neurodegenerative diseases or in vitro from soluble native proteins, respectively. The underlying structure of the fibrillar or 'cross-beta' state has presented long-standing, fundamental puzzles of protein structure. These include whether fibril-forming proteins have two structurally distinct stable states, native and fibrillar, and whether all or only part of the native protein refolds as it converts to the fibrillar state. Here we show that a designed amyloid-like fibril of the well-characterized enzyme RNase A contains native-like molecules capable of enzymatic activity. In addition, these functional molecular units are formed from a core RNase A domain and a swapped complementary domain. These findings are consistent with the zipper-spine model in which a cross-beta spine is decorated with three-dimensional domain-swapped functional units, retaining native-like structure.     

Long-term dendritic spine stability in the adult cortex

The structural dynamics of synapses probably has a crucial role in the development and plasticity of the nervous system. In the mammalian brain, the vast majority of excitatory axo-dendritic synapses occur on dendritic specializations called 'spines'. However, little is known about their long-term changes in the intact developing or adult animal. To address this question we developed a transcranial two-photon imaging technique to follow identified spines of layer-5 pyramidal neurons in the primary visual cortex of living transgenic mice expressing yellow fluorescent protein. Here we show that filopodia-like dendritic protrusions, extending and retracting over hours, are abundant in young animals but virtually absent from the adult. In young mice, within the 'critical period' for visual cortex development, approximately 73% of spines remain stable over a one-month interval; most changes are associated with spine elimination. In contrast, in adult mice, the overwhelming majority of spines (approximately 96%) remain stable over the same interval with a half-life greater than 13 months. These results indicate that spines, initially plastic during development, become remarkably stable in the adult, providing a potential structural basis for long-term information storage.     

Experience-dependent and cell-type-specific spine growth in the neocortex

Functional circuits in the adult neocortex adjust to novel sensory experience, but the underlying synaptic mechanisms remain unknown. Growth and retraction of dendritic spines with synapse formation and elimination could change brain circuits. In the apical tufts of layer 5B (L5B) pyramidal neurons in the mouse barrel cortex, a subset of dendritic spines appear and disappear over days, whereas most spines are persistent for months. Under baseline conditions, new spines are mostly transient and rarely survive for more than a week. Transient spines tend to be small, whereas persistent spines are usually large. Because most excitatory synapses in the cortex occur on spines, and because synapse size and the number of alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptors are proportional to spine volume, the excitation of pyramidal neurons is probably driven through synapses on persistent spines. Here we test whether the generation and loss of persistent spines are enhanced by novel sensory experience. We repeatedly imaged dendritic spines for one month after trimming alternate whiskers, a paradigm that induces adaptive functional changes in neocortical circuits. Whisker trimming stabilized new spines and destabilized previously persistent spines. New-persistent spines always formed synapses. They were preferentially added on L5B neurons with complex apical tufts rather than simple tufts. Our data indicate that novel sensory experience drives the stabilization of new spines on subclasses of cortical neurons. These synaptic changes probably underlie experience-dependent remodelling of specific neocortical circuits.     

人体完整腰椎骨三维运动的测量研究

为了进行人体完整腰椎骨运动的研究，设计并制造了平行光三维运动测量和分析系统．通过平行光测量系统，将每个椎体上三个标志点的运动投影到光屏上，以摄象机动态记录并输入计算机图象处理系统进行数据处理，同时运用刚体运动学原理进行计算，获得人体完整腰椎骨三维运动的运动特征．     

基于X线片的脊柱三维重建及分析

 以青少年脊柱侧凸诊断上的应用为背景,研究了一种在X线片上通过横向和纵向推理提取每块脊椎偏移和旋转信息的方法,然后结合标准三维脊柱模型重构出患者病变脊柱的空间形态,以便医生能直观、形象地从多个视角对患者的脊柱形态进行全方位的分析,最后对提出的一种Cobb角计算机辅助测量方法进行可行性分析.对文中提出的Marching Cubes与三维信息相结合的三维重建算法进行验证,实验证明该算法具有计算简单、可实现性好等优点,能精确地重构出患者的脊椎空间模型.     

内固定器对人体颈椎运动生物力学性能的影响

在失稳颈椎内安装固定器后,其稳定性的生物力学评价是医学研究的一个重要课题。该文提出了一种双目三维光学精密运动测量方法对其进行评价。采用该方法分别对保存良好的人体颈椎标本在失稳、内固定以及疲劳3种状态下的三维运动情况(包括前屈、后伸、左侧弯、右侧弯4种运动)进行了测量,并以每种运动情况下的颈椎C 1-C 2节相互旋转角度衡量其内固定的效果。实验结果表明,内固定后颈椎的旋转角度明显小于失稳时的旋转角度;内固定后模拟远期效果的疲劳加载后测试结果证实内固定的效果仍然是可靠的。     

颈椎融合器的力学测试及有限元验证

目的:设计一款聚乳酸材质融合器,完成力学测试并判定融合器的可行性.方法:分别对健康的和植入融合器的C5-C6节段羊颈椎进行压缩、前屈、后伸、侧弯实验,得出载荷-位移曲线,比较融合器植入前后颈椎的力学性能;再用万能电子试验机检验融合器的疲劳强度;最后对融合器进行有限元仿真观察应力分布.结果:加入融合器后颈椎的稳定性有所提高;融合器的疲劳强度合格,应力分布合理,未产生应力集中.结论:聚乳酸材质融合器有足够的力学强度,满足力学要求.     

钢筋混凝土双重网格楼盖结构构造特点的研究

钢筋混凝土双重网格结构在钢筋混凝土空腹夹层板基础上发展而来,但它的网格组成形式、结构构造发生了改变,对其构造特点进行研究分析,说明该结构结构型式合理,建筑造型新颖美观,适用于跨度和空间较大的公共建筑.     

辛三代数的结构群和结构代数

在文献[1]中,FAULKNER J R和FERRAR J C引入了辛三代数的定义,建立了它与李三系、李代数的联系,并且讨论了它的半单性、迹型和可解性.在文献[2]中,MEYBERG K定义了Jordan三系的结构群和结构代数.本文给出了辛三代数的结构群和结构代数的定义,并得到了几个重要结果:1)辛三代数y的结构群和与y相关联的李三系的自同构群的一个子群同构;2)辛三代数y的结构代数的一个子代数和与y相关联的李三系的导子代数的一个子代数同构;3)辛三代数y的结构代数的一个σ-不动点集与y的导子代数同构;4)辛三代数y的结构群对其内结构代数的一个作用是稳定的.     

椎间盘与韧带对腰椎变形影响的生物力学研究

利用CT扫描图像和三维重建软件,重建羊腰椎L1-L4的三维实体模型,建立包括椎体、椎间盘和韧带的有限元模型,数值计算得到椎体,椎间盘和韧带的应力分布规律.数值计算结果表明:施加单一牵引载荷和侧向弯曲载荷时,考虑间盘与韧带时腰椎的位移场与未考虑间盘与韧带时腰椎的位移场有明显的区别,对椎体的等效应力影响较小.文中结果表明,椎间盘与韧带明显影响腰椎变形的,研究腰椎变形需要考虑椎间盘与韧带.     

光子的夸克-胶子结构与结构函数分析

给出光子的夸克uF＋2/3（或dF＋1/3）和它的超对称性伴子ye,B^-2/3（或ye,B^＋1/3）由胶子弦粘合在一起的夸克-胶子结构,分析光子的结构函数．所得光子结构与实验观测事实相符;光子结构函数在动量分数x=2．5×10^-3时没有出现象质子结构函数那样的上升迹象,其原因可能是uF^＋2/3的超对称性伴子ye,B^-2/3质量标度很高所致．