感染性朊蛋白的毒性与糖磷脂酰肌醇

海绵状脑病（朊病）是一类神经退行性脑病，目前是世界上研究的热点问题之一。引起该病的病原是一种朊蛋白质，它与宿主自身的正常朊蛋白的一级结构相同，只是二级结构的构象有所不同。对海绵状脑病的致病机理至今仍不太清楚，作者针对目前研究的朊蛋白的致病性与GPI的关系作一综述。     

朊毒体蛋白的构象及其特性的研究进展

概述了朊毒体（包括构象不同的细胞型朊毒体和瘁病型朊毒体）蛋白的构象特征、朊毒体及其突变体的特性，阐明了PrP^sc是由PrP^c通过构象转变而成的，不同物种PrP的三维结构有相似的球形结构域（125-228位）和N-末端无规卷曲尾，其球形结构域均由3个螺旋（144～154、173～194和200-228位）以及1对反平行的β-折叠（128-131和161-164）组成，但其结构和表面电荷分布存在一些区别。     

感染性朊蛋白的毒性与糖磷脂酰肌醇（GPI）的相关性

海绵状脑病(朊病)是一类神经退行性脑病,目前是世界上研究的热点问题之一。引起该病的病原是一种朊蛋白质,它与宿主自身的正常朊蛋白的一级结构相同,只是二级结构的构象有所不同。对海绵状脑病的致病机理至今仍不太清楚,作者针对目前研究的朊蛋白的致病性与GPI的关系作一综述。     

朊病毒外周传播机理的研究进展

朊病是一种神经退行性疾病，能造成大脑神经细胞的广泛性损伤，最终导致死亡。一些朊病因子在感染部位向大脑入侵时先在淋巴组织中沉积，如在肠道中。对鼠模型的研究结果表明，这种沉积是必需的，它能使朊病因子有效到达大脑，如果这种在淋巴组织的沉积被阻断，就使疾病的易感性减弱。因此，鉴定在朊病毒到达大脑过程中参与的细胞和分子可以筛选出治疗本病的药物。作者就目前对朊病毒外周传播机理的理解作一综述。     

淀粉样物质结构的研究方法

由于淀粉样物质与许多疾病,如阿兹海默病、帕金森综合症和朊蛋白病相关,对其形成的基础—结构方面的研究引起了越来越多人的关注。但是其结构目前还没有被完全研究清楚,文章介绍了多种研究淀粉样物质结构的先进方法,如氢氘交换、冷冻显微技术、固态核磁共振等。人们利用这些技术能够更精确地观察和研究淀粉样物质的结构,以便进行深入研究。     

朊病毒的生物学研究进展

朊病是一种人畜共患病,对人和动物的健康影响很大,是一种致死性疾病。由于该病的临床症状和组织病理学变化等已为人们所熟知,作者仅就朊病的病因、发病机理及诊断治疗方法的最新研究进行阐述,并从病原基因遗传结构特点、致病蛋白的结构特点、诊断方法的提高、治疗的前景作一综述,为该类疾病的防治提供理论依据。     

新型克-雅氏病是人的牛海绵状脑病

本文在概述人朊病毒的基础上，重点阐述新型克－雅氏病与早先已知的各种人朊病毒病的区别和它是由牛海绵状脑病朊病毒引起的证据，并对牛海绵状脑病并非源于痒病而是原于牛散发性朊病毒病、影响新型克－雅氏病流行规模的主要因素和该病的主要预防措施等问题进行了讨论。     

朊蛋白生理功能的研究进展

朊蛋白（prion protein,PrPC）作为朊病的主要相关致病因子而受到广泛关注,虽然PrPC在疾病过程中的作用已有很多了解,但关于PrPC的正常生理功能还不是很清楚。就目前的研究而言,PrPC存在细胞核亚型和细胞质亚型两种形式,可与相应的配体及铜离子作用,并且通过细胞内吞或内陷方式被细胞膜获取;主要生理功能表现为保护神经系统,协助细胞的凋亡和分化、黏附,以及参与信号转导等。因此,文章对朊蛋白的生理功能作一总结,为朊病的治疗和朊蛋白的进一步研究提供了理论基础。     

羊痒病研究进展

痒病是一类引起绵羊和山羊的神经退行性脑病,是目前世界上研究的热点问题之一。引起该病的病原是一种朊蛋白,它与宿主自身的正常朊蛋白在一级结构上是相同的,只是二级结构的构象有所不同。论文针对目前人们研究中发现羊痒病的病毒存在不同的毒株型,以及不同毒株型的分子特性做一综述。     

美国培育出不可能感染疯牛病的牛

<正>美国农业部国家动物疾病中心采用基因工程技术,首次培育出了生物学上不可能感染疯牛病的牛。近日,美国农业部国家动物疾病中心的Richt等报告,他们采用基因工程技术,首次培育出了生物学上不可能感染疯牛病的牛。疯牛病的病原体被称为朊毒体,它由脑内、免疫系统及其他组织中一种正常的蛋白质朊蛋白发生错误折叠而产生。朊毒体会引起牛海绵样脑病(BSE),即疯牛病和人克雅病(CJD),还可引起绵羊瘙痒病及鹿和貂的致命消耗性疾病。致病朊毒体不含遗传物     

重组绵羊pEGFP-PRNP质粒构建及真核细胞转染

构建了携带绵羊朊蛋白基因（PRNP）的重组真核转染质粒,通过脂质体转染试剂将其转染至神经胶质瘤细胞（C6）,以期为进一步研究绵羊朊蛋白的生理功能和从细胞水平研究朊蛋白疾病的发病机制奠定基础。采用PCR扩增目的基因序列,纯化后将其克隆到带有增强型绿色荧光蛋白（EGFP）报告基因的真核表达载体pEGFP-N1中,对重组质粒pEGFP-PRNP做酶切鉴定。而后采用阳离子脂质体转染法将重组质粒转染到C6细胞,荧光显微镜观察。经鉴定,绵羊PRNP基因已克隆到真核表达载体pEGFP-N1中,成功地构建了重组pEGFP-PRNP质粒,并可稳定地在C6细胞中表达。本研究为外源朊蛋白在细胞中表达提供了平台,为进一步在细胞水平研究朊病毒疾病奠定了基础。     

Enrichment of prion protein in exosomes derived from ovine cerebral spinal fluid

Prion diseases are transmissible neurodegenerative disorders affecting humans and a wide variety of animal species including sheep and cattle. The transmissible agent, the prion, is an abnormally folded form (PrP(Sc)) of the host encoded cellular prion protein (PrP(C)). Distribution of the prion protein in the fluids of species susceptible to these diseases is of importance to human health and the iatrogenic spread of prion disease. Aside from blood which is confirmed to be a source of prion infectivity, it is currently unclear which other body fluids harbor a significant transmission risk. In the current study we examined two ovine fluids; pseudo-afferent lymph and cerebral spinal fluid (CSF), for the presence of exosomes and concurrent enrichment of the normal, cellular form of the prion protein (PrP(C)). Here we demonstrate the existence of exosomes in both pseudo-afferent lymph and CSF isolated from sheep. In the CSF derived exosomes we were able to show an enrichment of PrP(C) over unfractionated CSF. This experimental approach suggests that CSF derived exosomes could be used as a novel means of detecting abnormal forms of the prion protein and provide an in vivo link between these vesicles and prion disease pathogenesis.     

Cell models of prion infection

Due to recent renewal of interest and concerns in prion diseases, a number of cell systems permissive to prion multiplication have been generated in the last years. These include established cell lines, neuronal stem cells and primary neuronal cultures. While most of these models are permissive to experimental, mouse-adapted strains of prions, the propagation of natural field isolates from sheep scrapie and chronic wasting disease has been recently achieved. These models have improved our knowledge on the molecular and cellular events controlling the conversion of the PrP(C) protein into abnormal isoforms and on the cell-to-cell spreading of prions. Infected cultured cells will also facilitate investigations on the molecular basis of strain identity and on the mechanisms that lead to neurodegeneration. The ongoing development of new cell models with improved characteristics will certainly be useful for a number of unanswered critical issues in the prion field.     

Rodent models for prion diseases

Until today most prion strains can only be propagated and the infectivity content assayed by experimentally challenging conventional or transgenic animals. Robust cell culture systems are not available for any of the natural and only for a few of the experimental prion strains. Moreover, the pathogenesis of different transmissible spongiform encephalopathies (TSE) can be analysed systematically by using experimentally infected animals. While, in the beginning, animals belonging to the natural host species were used, more and more rodent model species have been established, mostly due to practical reasons. Nowadays, most of these experiments are performed using highly susceptible transgenic mouse lines expressing cellular prion proteins, PrP, from a variety of species like cattle, sheep, goat, cervidae, elk, hamster, mouse, mink, pig, and man. In addition, transgenic mice carrying specific mutations or polymorphisms have helped to understand the molecular pathomechanisms of prion diseases. Transgenic mouse models have been utilised to investigate the physiological role of PrP(C), molecular aspects of species barrier effects, the cell specificity of the prion propagation, the role of the PrP glycosylation, the mechanisms of the prion spread, the neuropathological roles of PrP(C) and of its abnormal isoform PrP(D) (D for disease) as well as the function of PrP Doppel. Transgenic mouse models have also been used for mapping of PrP regions involved in or required for the PrP conversion and prion replication as well as for modelling of familial forms of human prion diseases.     

The Potential of Mesenchymal Stem Cell in Prion Research

Scrapie and bovine spongiform encephalopathy are fatal neurodegenerative diseases caused by the accumulation of a misfolded protein (PrP^res), the pathological form of the cellular prion protein (PrP^C). For the last decades, prion research has greatly progressed, but many questions need to be solved about prion replication mechanisms, cell toxicity, differences in genetic susceptibility, species barrier or the nature of prion strains. These studies can be developed in murine models of transmissible spongiform encephalopathies, although development of cell models for prion replication and sample titration could reduce economic and timing costs and also serve for basic research and treatment testing. Some murine cell lines can replicate scrapie strains previously adapted in mice and very few show the toxic effects of prion accumulation. Brain cell primary cultures can be more accurate models but are difficult to develop in naturally susceptible species like humans or domestic ruminants. Stem cells can be differentiated into neuron‐like cells and be infected by prions. However, the use of embryo stem cells causes ethical problems in humans. Mesenchymal stem cells (MSCs) can be isolated from many adult tissues, including bone marrow, adipose tissue or even peripheral blood. These cells differentiate into neuronal cells, express PrP^C and can be infected by prions in vitro. In addition, in the last years, these cells are being used to develop therapies for many diseases, including neurodegenerative diseases. We review here the use of cell models in prion research with a special interest in the potential use of MSCs.     

Healthy goats naturally devoid of prion protein

Prion diseases such as scrapie in small ruminants, bovine spongiform encephalopathy (BSE) in cattle and Creutzfeldt-Jakob disease (CJD) in man, are fatal neurodegenerative disorders. These diseases result from the accumulation of misfolded conformers of the host-encoded prion protein (PrP) in the central nervous system. To date naturally-occurring PrP free animals have not been reported. Here we describe healthy non-transgenic animals, Norwegian Dairy Goats, lacking prion protein due to a nonsense mutation early in the gene. These animals are predicted to be resistant to prion disease and will be valuable for research and for production of prion-free products.     

Recent developments in prion disease research: diagnostic tools and in vitro cell culture models

After prion infection, an abnormal isoform of prion protein (PrP(Sc)) converts the cellular isoform of prion protein (PrP(C)) into PrP(Sc). PrP(C)-to-PrP(Sc) conversion leads to PrP(Sc) accumulation and PrP(C) deficiency, contributing etiologically to induction of prion diseases. Presently, most of the diagnostic methods for prion diseases are dependent on PrP(Sc) detection. Highly sensitive/accurate specific detection of PrP(Sc) in many different samples is a prerequisite for attempts to develop reliable detection methods. Towards this goal, several methods have recently been developed to facilitate sensitive and precise detection of PrP(Sc), namely, protein misfolding cyclic amplification, conformation-dependent immunoassay, dissociation-enhanced lanthanide fluorescent immunoassay, capillary gel electrophoresis, fluorescence correlation spectroscopy, flow microbead immunoassay, etc. Additionally, functionally relevant prion-susceptible cell culture models that recognize the complexity of the mechanisms of prion infection have also been pursued, not only in relation to diagnosis, but also in relation to prion biology. Prion protein (PrP) gene-deficient neuronal cell lines that can clearly elucidate PrP(C) functions would contribute to understanding of the prion infection mechanism. In this review, we describe the trend in recent development of diagnostic methods and cell culture models for prion diseases and their potential applications in prion biology.     

Elevated manganese levels in blood and central nervous system occur before onset of clinical signs in scrapie and bovine spongiform encephalopathy

Prion diseases, or transmissible spongiform encephalopathies, are neurodegenerative diseases that can only be accurately diagnosed by analysis of central nervous system tissue for the presence of an abnormal isoform of the prion protein known as PrP(Sc). Furthermore, these diseases have long incubation periods during which there are no clear symptoms but where the infectious agent could still be present in the tissues. Therefore, the development of diagnostic assays to detect a surrogate marker for the presence of prion disease is essential. Previous studies on mice experimentally infected with scrapie, an ovine spongiform encephalopathy, suggested that changes in the levels of Mn occur in the blood and brain before the onset of symptoms of the disease. To assess whether these findings have relevance to the animal diseases scrapie and bovine spongiform encephalopathy, tissues from bovine spongiform encephalopathy- and scrapie-infected cattle and sheep were analyzed for their metal content and compared with values for noninfected animals. In field cases and experimentally infected animals, elevated Mn was associated with prion infection. Although some central nervous system regions showed elevated Mn, other regions did not. The most consistent finding was an elevation of Mn in blood. This change was present in experimentally infected animals before the onset of symptoms. In scrapie-infected sheep, elevated Mn levels occurred regardless of the genotype of the sheep and were even detected in scrapie-resistant sheep in which no symptoms of disease were detected. These findings suggest that elevated blood Mn could be a potential diagnostic marker for prion infection even in the absence of apparent clinical disease.     

朊病毒检测方法综述

传染性海绵状脑病(transmissible spongiform encephalopathies,TSEs)是由朊病毒引起的人和多种哺乳动物以神经退行性变化为主要特征的一种慢性消耗性传染病,也称作朊病毒病;其是由体内正常细胞表面的PrP^C转变成PrP^Sc蛋白所导致。但PrP^Sc主要在脑内表达,在其他组织表达量很低,因此快捷准确的诊断方法对于该病有重要意义。作者重点介绍以朊病毒的致病特点为依据建立的检测方法,便于对该疾病的早期诊断、预防以及食品安全检测提供帮助,保障畜牧业的有序发展以及人类的健康。