疯牛病研究进展

近年来,英国疯牛病成为国内外新闻媒体关注的焦点,它不仅给英国养牛业造成了巨大的经济损失,同时也引起了世界各国政府及农业主管部门的高度重视,疯牛病是对牛海绵状脑病（Bovine Spongiform Encephalopathy BSE)的俗称,它是一种慢性,具有传染性的致死性中枢神经系统疾病,该病临床和组织学病理学特征是病畜神经失常,共济失调,感觉过敏和中枢神经系统灰质空泡化,疯牛病被认为与朊病毒（Prion)有关,因此与克-雅氏病,羊痒病等一起被统称为朊病毒病,朊病毒病是由正常朊病毒蛋白（PrPC)发生蛋白折叠错误变成异常朊病毒蛋白（PrPSC)引起的,BSE主要发生于以英国为中心的欧洲各国家和地区,各国采取了一系列的措施,防制BSE危害人类健康,中国至今尚无疯牛病发生。     

朊病毒与血液制品安全性

疯牛病是对牛海绵伏脑病（Bovine Spongiform Encephalopathy BSE）的简称,是一种慢性、具有传染性的致死性中枢神经系统疾病,英国于1985年4月首先发现该病,并于1986年11月定名为BSE,此病的组织病理学变化和临床症状与人的库鲁病（Kuru）、克雅氏病（Creutzfeld-Jakob,Disease CJD）和羊痒病（Scrapie）相似,因此,被认为与朊病毒（Prion）有关,亦称为朊病毒病,其传播途径与潜伏CJD的献血者的血液和血浆是否关联,是人们极为关注的问题,本文就朊病毒的理化特性与生物学特性、传播方式、危害和对血液制品的安全性可能带来的影响作一综述。     

朊病毒疾病的诊断技术

朊病毒疾病即海绵状脑病,是人的动物中的一类致死性中央神经系统疾病,介绍了朊病毒疾病的诊断技术,病理诊断,基因诊断,免疫诊断和血液检测。     

哺乳动物中蛋白质折叠异常与朊病毒病

在过去的两年中 ,有关朊病毒 (ｐｒｉｏｎ)蛋白结构的最新ＮＭＲ数据不断扩展 ,这主要是在仓鼠和人体蛋白质中取得的。另外 ,两种朊病毒蛋白的折叠动力学机制亦被阐明。目前已经得到几种朊病毒的样品 ,它们可以在溶液中采取不同的构象。近来所做的有关朊病毒蛋白链分子基础的研究工作进一步巩固了蛋白质独自致病的假说。通过对最小蛋白质片断的辨认 ,疾病与结构的相互关系的研究也取得了重要进展。1 .引言在 2 5种被确证为由动物组织中变性蛋白质沉积而引起的病症中 ,只有传染性海绵状脑病 (ＴＳＥｓ)会产生传染性物质。可能朊病毒疾病最独…     

朊病毒病

朊病毒是一类可传递的蛋白质感染颗粒,似乎仅仅由一种修饰后的蛋白ＰｒＰ＾ｓｃ构成。ＰｒＰ＾ｓｃ由政党细胞蛋白ＰｒＰ＾ｃ通过翻译后加工、发生构象变化而形成。牛海绵状脑病和人Ｃｒｅｕｔｚｆｅｌｄｔ－Ｊａｃｏｂ病（ＣＪＤ）是朊病毒引起的最名的中枢神经系统变性疾病。ＤＪＤ有散发性、家族性、传染性、医原性等,有语气表明牛朊病毒已传递至人类导致新的变异性ＣＪＤ。针对朊病毒独特的致病机制可得出若干防治策略。     

疯牛病病原体的研究进展

疯牛病病原体,即朊病毒蛋白（PrionPrP^Sc),是一种不具有核酸的全新致病物质,在动物和人类引起致命性海绵状脑病（Prions疾病）,本文阐述了Prion的含义,Prion蛋白的种类,疾病生物化学以及与Prions疾病的关系。     

朊病毒研究的现状

朊病毒疾病是人和动物中的一种传染性,散发性和遗传性的神经退行性脑病,到目前为止,关于此病的致病机制并不十分清楚,但有研究表明该病是由一种正常蛋白PrP的不正常折叠形式在大脑中积聚所致,许多哺乳动物和鸟类的朊病毒基因和氨基酸序列都已被分析,而且传染源的部分性质已经被阐明,朊病毒疾病的传染和潜伏期在人和动物中有很大差异,一些变异与疾病的自发性,易感性和抵抗性有关,为得到关于现毒更多的信息,已对其基因两端进行了大范围的DNA测序分析,至于正常细胞蛋白PrP的生理功能,现在也并不确定。     

朊病毒,应改名

Prion是感染性蛋白质,能引发疯牛病等海绵状脑病。朊病毒是其被广泛沿用的中文译名．暗示prion是某种病毒。实际上,prion是蛋白质,不是病毒。朊病毒这个名字与实际不副,可能误导人们对prion的认识．也会误导对海绵状脑病的防治。因此,朊病毒不宜继续沿用下去,而应另起名实相副的新名。     

朊病毒的分子遗传学研究

院病毒（prion）是一种能引起哺乳动物中枢神经系统退行性疾病,即一般称为传染性海绵状脑病（BE）的病原因子。这些疾病包括人类克雅氏病忙reutrfeltJocob）、致死性家族失眠症、羊瘙痒症和疯牛病等[1]．1982年美国科学家Prusiner从感染了瘙痒症的叙利亚老鼠脑中分离出具有传染性的蛋白因子,并首先提出朊病毒这一名词,以与病毒、类病毒相区别。朊病毒最显著的特征是不含核酸。现已知道朊病毒（prionprotein,PrP是由宿主染色体基因PrP编码的一种正常细胞蛋白,在非患病个体中以低水平存在。这种蛋白在细胞内质网上合成、高尔基小体中…     

利用RNAi逆转录病毒载体系统对人朊病毒蛋白表达的稳定抑制

朊病毒是引起可传染的致死性海绵状脑病的致病因子,细胞中正常的朊病毒蛋白(PrPC)在该疾病病程发展中起着必不可少的作用。同时,PrPC已被证明在胃癌、乳腺癌等癌症中发挥着保护癌细胞的作用。根据人源PrPC(HuPrPC)cDNA序列,本研究设计了4种19nt的siRNA,将其构建成RNAi逆转录病毒载体系统,进行了其对HuPrPC表达的抑制效应的分析,从中获得了能高效稳定抑制HuPrPC表达的3种靶向序列,其中si626(5′-GGTTGAGCAGATGTGTATC-3′)的抑制效果最为明显,其抑制效率可达85%以上。随后,利用筛选出的si292和si626的稳定干扰细胞系进行了细胞浸润性实验,结果发现,PrPC干扰细胞系细胞浸润能力显著下降。这为进一步研究朊病毒疾病的基因治疗、以PrPC为靶标进行PrPC相关癌症的辅助治疗研究奠定了一定的基础。     

Characterization and enzymatic degradation of Sup35NM, a yeast prion-like protein

Transmissible spongiform encephalopathies (TSEs) are believed to be caused by an unconventional infectious agent, the prion protein. The pathogenic and infectious form of prion protein, PrPSc, is able to aggregate and form amyloid fibrils, very stable and resistant to most disinfecting processes and common proteases. Under specific conditions, PrPSc in bovine spongiform encephalopathy (BSE) brain tissue was found degradable by a bacterial keratinase and some other proteases. Since this disease-causing prion is infectious and dangerous to work with, a model or surrogate protein that is safe is needed for the in vitro degradation study. Here a nonpathogenic yeast prion-like protein, Sup35NM, cloned and overexpressed in E. coli, was purified and characterized for this purpose. Aggregation and deaggregation of Sup35NM were examined by electron microscopy, gel electrophoresis, Congo red binding, fluorescence, and Western blotting. The degradation of Sup35NM aggregates by keratinase and proteinase K under various conditions was studied and compared. These results will be of value in understanding the mechanism and optimization of the degradation process.     

Biochemical and strain properties of CJD prions: complexity versus simplicity

Prions, the agents responsible for transmissible spongiform encephalopathies, are infectious proteins consisting primarily of scrapie prion protein (PrP(Sc)), a misfolded, β-sheet enriched and aggregated form of the host-encoded cellular prion protein (PrP(C)). Their propagation is based on an autocatalytic PrP conversion process. Despite the lack of a nucleic acid genome, different prion strains have been isolated from animal diseases. Increasing evidence supports the view that strain-specific properties may be enciphered within conformational variations of PrP(Sc). In humans, sporadic Creutzfeldt-Jakob disease (sCJD) is the most frequent form of prion diseases and has demonstrated a wide phenotypic and molecular spectrum. In contrast, variant Creutzfeldt-Jakob disease (vCJD), which results from oral exposure to the agent of bovine spongiform encephalopathy, is a highly stereotyped disease, that, until now, has only occurred in patients who are methionine homozygous at codon 129 of the PrP gene. Recent research has provided consistent evidence of strain diversity in sCJD and also, unexpectedly enough, in vCJD. Here, we discuss the puzzling biochemical/pathological diversity of human prion disorders and the relationship of that diversity to the biological properties of the agent as demonstrated by strain typing in experimental models.     

Comparative analysis of Japanese and foreign L-type BSE prions

L-type bovine spongiform encephalopathy (BSE) is an atypical form of BSE. To characterize the Japanese L-type BSE prion, we conducted a comparative study of the Japanese and foreign L-type BSE isolates. The L-type BSE isolates of Japan, Germany, France and Canada were intracerebrally inoculated into bovinized prion protein-overexpressing transgenic mice (TgBoPrP). All the examined L-type BSE isolates were transmitted to TgBoPrP mice, and no clear differences were observed in their biological and biochemical properties. Here, we present evidence that the Japanese and Canadian L-type BSE prions are identical to those from the European cases.     

Novel Aspects of Prions,Their Receptor Molecules,and Innovative Approaches for TSE Therapy

1. Prion diseases are a group of rare, fatal neurodegenerative diseases, also known as transmissible spongiform encephalopathies (TSEs), that affect both animals and humans and include bovine spongiform encephalopathy (BSE) in cattle, scrapie in sheep, chronic wasting disease (CWD) in deer and elk, and Creutzfeldt–Jakob disease (CJD) in humans. TSEs are usually rapidly progressive and clinical symptoms comprise dementia and loss of movement coordination due to the accumulation of an abnormal isoform (PrP^Sc) of the host-encoded prion protein (PrP^c). 2. This article reviews the current knowledge on PrP^c and PrP^Sc, prion replication mechanisms, interaction partners of prions, and their cell surface receptors. Several strategies, summarized in this article, have been investigated for an effective antiprion treatment including development of a vaccination therapy and screening for potent chemical compounds. Currently, no effective treatment for prion diseases is available. 3. The identification of the 37 kDa/67 kDa laminin receptor (LRP/LR) and heparan sulfate as cell surface receptors for prions, however, opens new avenues for the development of alternative TSE therapies.     

Conversion of the BASE prion strain into the BSE strain: the origin of BSE?

Atypical neuropathological and molecular phenotypes of bovine spongiform encephalopathy (BSE) have recently been identified in different countries. One of these phenotypes, named bovine "amyloidotic" spongiform encephalopathy (BASE), differs from classical BSE for the occurrence of a distinct type of the disease-associated prion protein (PrP), termed PrP(Sc), and the presence of PrP amyloid plaques. Here, we show that the agents responsible for BSE and BASE possess different biological properties upon transmission to transgenic mice expressing bovine PrP and inbred lines of nontransgenic mice. Strikingly, serial passages of the BASE strain to nontransgenic mice induced a neuropathological and molecular disease phenotype indistinguishable from that of BSE-infected mice. The existence of more than one agent associated with prion disease in cattle and the ability of the BASE strain to convert into the BSE strain may have important implications with respect to the origin of BSE and spongiform encephalopathies in other species, including humans.     

Spongiform encephalopathies and prions: An overview of pathology and disease mechanisms

Abstract The etiology of spongiform encephalopathies has been sharply contested for decades. At the heart of the issue is the question of disease origin: Are prion diseases representative of primary neurodegenerative genetic disorders, or are they bona fide infectious diseases? This article provides a brief outline of the progress made in the elucidation of prion disease mechanisms in the context of pathological support of the 'protein only' hypothesis. The answer to the above question appears to be that spongiform encephalopathies are uniquely both infectious and genetic neurodegenerative diseases.     

Thermodynamic characterization for the denatured state of bovine prion protein and the BSE Associated variant E211K

ABSTRACT

Propagation of transmissible spongiform encephalopathies involves the conversion of cellular prion protein, PrP^C, into a misfolded oligomeric form, PrP^Sc. The most common hereditary prion disease is a genetic form of Creutzfeldt-Jakob disease in humans, in which a mutation in the prion gene results in a glutamic acid to lysine substitution at position 200 (E200K) in PrP. In cattle, the analogous amino acid substitution is found at residue 211 (E211K) and has been associated with a case of bovine spongiform encephalopathy. Here, we have compared the secondary structure of E211K to that of wild type using circular dichroism and completed a thermodynamic analysis of the folding of recombinant wild type and E211K variants of the bovine prion protein. The secondary structure of the E211K variant was essentially indistinguishable from that of wild type. The thermodynamic stability of E211K substitution showed a slight destabilization relative to the wild type consistent with results reported for recombinant human prion protein and its mutant E200K. In addition, the E211K variant exhibits a similarly compact denatured state to that of wild type based upon similar m-value and change in heat capacity of unfolding for the proteins. Together these results indicate that residual structure in the denatured state of bPrP is present in both the wild type protein and BSE associated variant E211K. Given this observation, as well as folding similarities reported for other disease associated variants of PrP it is worth consideration that functional aspects of PrP conformation may play a role in the misfolding process.     

Immunity against prions?

Several recent reports indicate that antibodies directed against the cellular form of the prion protein, PrP^C, might eliminate the transmissible agent of spongiform encephalopathies (the prion) from scrapie-infected cells in vitro, and that a humoral immune response could prevent scrapie pathogenesis in vivo. These findings suggest that immunotherapeutical intervention against prion diseases is not unattainable. Will vaccines and post-exposure strategies based on antibodies ever prove useful against scrapie, bovine spongiform encephalopathy (BSE), or Creutzfeldt–Jakob disease?     

Molecular analysis of carbohydrate N-acetylgalactosamine 4-O sulfotransferase 8 (CHST8) as a candidate gene for bovine spongiform encephalopathy susceptibility

Endogenous prion proteins (PrP) play the central role in the pathogenesis of transmissible spongiform encephalopathies. The carbohydrate N -acetylgalactosamine 4-O sulfotransferase 8 (CHST8) promotes the conversion of the cellular PrP^C into the pathogenic PrP^d. Six sequence variants within the CHST8 gene were identified by comparative sequencing and genotyped for a sample of 623 animals comprising bovine spongiform encephalopathy (BSE)-affected and healthy control cows representing German Fleckvieh (German Simmental), German Holstein (Holstein-Friesian) and Brown Swiss. Significant differences in the allele, genotype and haplotype frequencies between BSE-affected and healthy cows indicate an association of sequence variant g.37254017G>T with the development of the disease in Brown Swiss cattle.     

Cell culture models of transmissible spongiform encephalopathies

Cell cultures represent versatile and useful experimental models of transmissible spongiform encephalopathies. These models include chronically prion infected cell lines, as well as cultures expressing variable amounts of wild-type, mutated or chimeric prion proteins. These cultures have been widely used to investigate the biology of both the normal and the pathological isoform of the prion protein. They have also contributed to the comprehension of the pathogenic processes occurring in transmissible spongiform encephalopathies and in the development of new therapeutic approaches of these diseases.