Prion diseases: disease diversity and therapeutics]

More than one hundred victims of iatrogenic CJD with cadaveric dura mater grafting have been recognized in Japan, and the people have been also exposed to a risk of outbreaks of variant CJD. These diseases are distinct from other forms of prion diseases as well as other neuropsychiatric disorders, but on an early clinical stage, their differential diagnoses from other atypical forms of prion diseases are not necessarily easy. Thus, atypical forms of prion diseases were overviewed and discussed here. In addition, data on recent clinical trials of enteral antimalarial drug (quinacrine or quinine) treatment or intracerebroventricular pentosan polysulfate treatment were presented and discussed, because research progress in the therapeutics for prion diseases has been remarkably made on the basis of the prevalence of those acquired forms of prion diseases.     

Review: Creutzfeldt-Jakob disease: prion protein type, disease phenotype and agent strain

The human transmissible spongiform encephalopathies or human prion diseases are one of the most intensively investigated groups of rare human neurodegenerative conditions. They are generally held to be unique in terms of their complex epidemiology and phenotypic variability, but they may also serve as a paradigm with which other more common protein misfolding disorders might be compared and contrasted. The clinico-pathological phenotype of human prion diseases appears to depend on a complex interaction between the prion protein genotype of the affected individual and the physico-chemical properties of the neurotoxic and transmissible agent, thought to comprise of misfolded prion protein. A major focus of research in recent years has been to define the phenotypic heterogeneity of the recognized human prion diseases, correlate this with molecular-genetic features and then determine whether this molecular-genetic classification of human prion disease defines the biological properties of the agent as determined by animal transmission studies. This review seeks to survey the field as it currently stands, summarize what has been learned, and explore what remains to be investigated in order to obtain a more complete scientific understanding of prion diseases and to protect public health.     

Persistent retroviral infection with MoMuLV influences neuropathological signature and phenotype of prion disease

A fundamental step in pathophysiology of prion diseases is the conversion of the host encoded prion protein (PrP^C) into a misfolded isoform (PrP^Sc) that accumulates mainly in neuronal but also non-neuronal tissues. Prion diseases are transmissible within and between species. In a subset of prion diseases, peripheral prion uptake and subsequent transport to the central nervous system are key to disease initiation. The involvement of retroviruses in this process has been postulated based on the findings that retroviral infections enhance the spread of prion infectivity and PrP^Sc from cell to cell in vitro. To study whether retroviral infection influences the phenotype of prion disease or the spread of prion infectivity and PrP^Sc in vivo, we developed a murine model with persistent Moloney murine leukemia retrovirus (MoMuLV) infection with and without additional prion infection. We investigated the pathophysiology of prion disease in MoMuLV and prion-infected mice, monitoring temporal kinetics of PrP^Sc spread and prion infectivity, as well as clinical presentation. Unexpectedly, infection of MoMuLV challenged mice with prions did not change incubation time to clinical prion disease. However, clinical presentation of prion disease was altered in mice infected with both pathogens. This was paralleled by remarkably enhanced astrogliosis and pathognomonic astrocyte morphology in the brain of these mice. Therefore, we conclude that persistent viral infection might act as a disease modifier in prion disease.     

Complement activation in human prion disease

The central event in the neuropathological process of prion diseases (PrD) is the accumulation of abnormal prion protein accompanied by severe neuronal loss. Despite the infectious nature of these diseases, no prominent immune response has been detected yet. However, recent studies have shown that complement, a component of the innate immune system, is involved in the early pathogenesis of experimental prion infection. Here we demonstrate, in the diseased human brains, the presence of active compounds of the complement system, like C1q and C3b, in extracellular disease-associated prion protein deposits and the membrane attack complex in neurons. The neuronal localization of the membrane attack complex correlates well with the severity of disease-specific pathology and TUNEL labeling of neurons, irrespective of genotype or molecular phenotype of human prion diseases.     

Molecular pathology of human prion disease

Human prion diseases are associated with a range of clinical presentations and are classified by both clinicopathological syndrome and aetiology with sub-classification according to molecular criteria. Considerable experimental evidence suggests that phenotypic diversity in human prion disease relates in significant part to the existence of distinct human prion strains encoded by abnormal PrP isoforms with differing physicochemical properties. To date, however, the conformational repertoire of pathological isoforms of wild-type human PrP and the various forms of mutant human PrP has not been fully defined. Efforts to produce a unified international classification of human prion disease are still ongoing. The ability of genetic background to influence prion strain selection together with knowledge of numerous other factors that may influence clinical and neuropathological presentation strongly emphasises the requirement to identify distinct human prion strains in appropriate transgenic models, where host genetic variability and other modifiers of phenotype are removed. Defining how many human prion strains exist allied with transgenic modelling of potentially zoonotic prion strains will inform on how many human infections may have an animal origin. Understanding these relationships will have direct translation to protecting public health.     

Creutzfeldt–Jakob disease

It is valuable to summarize the milestone study of prion diseases done in Japan for review in the journal Neuropathology in 2000. Many studies done in Japan promote world prion research activity, and also influence further research projects in other groups abroad. In this review the author focuses on the transmission experiment, the discovery of abnormal prion protein localization in the synaptic structures or follicular dendritic cells, and the genetic analysis of prion protein gene for the establishment of familial prion diseases.     

Phenotypic variability of sporadic human prion disease and its molecular basis: past,present, and future

Human prion diseases are rare neurodegenerative disorders related to prion protein misfolding that can occur as sporadic, familial or acquired forms. In comparison to other more common neurodegenerative disorders, prion diseases show a wider range of phenotypic variation and largely transmit to experimental animals, a feature that led to the isolation and characterization of different strains of the transmissible agent or prion with distinct biological properties. Biochemically distinct PrP^Sc types have been demonstrated which differ in their size after proteinase cleavage, glycosylation pattern, and possibly other features related to their conformation. These PrP^Sc types, possibly enciphering the prion strains, together with the naturally occurring polymorphism at codon 129 in the prion protein gene have a major influence on the disease phenotype. In the sporadic form, the most common but perhaps least understood form of human prion disease, there are at least six major combinations of codon 129 genotype and prion protein isotype, which are significantly related to distinctive clinical–pathological subgroups of the disease. In this review, we provide an update on the current knowledge and classification of the disease subtypes of the sporadic human prion diseases as defined by molecular features and pathological changes. Furthermore, we discuss the molecular basis of phenotypic variability taking into account the results of recent transmission studies that shed light on the extent of prion strain variation in humans.     

Neurodegeneration and oxidative stress: prion disease results from loss of antioxidant defence

Prion diseases or transmissible spongiform encephalopathies (TSEs) are rare neurodegenerative disorders that can be acquired either by direct transmission, inherited through dominant mutations in the prion protein gene or via an unknown sporadic cause. This latter group constitutes the vast majority of cases. Like many neurodegenerative diseases the hallmarks of oxidative damage can be readily detected throughout the brain of the affected individual. However, unlike most other neurodegenerative diseases, prion diseases are connected with a dramatic loss of antioxidant defence. As abnormal protein accumulates in the diseased brain there is both an increase of oxidative substances and a loss of the defences that keep them in check. In particular the normal cellular prion protein has been shown to be an antioxidant. Conversion of this protein to the protease resistant isoform is accompanied by a loss of this antioxidant activity. This change creates a paradox as the loss of activity is not accompanied by a loss of protein expression. It is likely that this prevents other cellular defences from responding sufficiently to protect neurons from the heightened oxidative burden. Recent experiments with transgenic mice have shown that when prion protein expression is switched off during the course of prion disease, cell death is dramatically halted and the mouse recovers from the disease. This result clearly illustrates that the continued expression of non-function prion protein is essential for disease progression. This implies that the presence of this abnormal protein during prion disease causes a failure of cellular antioxidant defence. This failed defence is the fundamental cause of the massive neurodegeneration that results in the fatal nature of TSEs. The role of oxidative stress in TSEs and other neurodegenerative disorders are discussed in this review.     

Update on Creutzfeldt-Jakob disease

PURPOSE OF REVIEW: Prion diseases are transmissible fatal neurodegenerative disorders in which infectivity is associated with the accumulation of PrP(Sc), a disease-related isoform of normal cellular prion protein. The recent emergence of variant Creutzfeldt-Jakob disease has led to major public health concerns, and the need for the development of effective treatments. As PrP(Sc) is associated both with pathology and infectivity, therapeutic approaches to date have largely aimed at preventing its accumulation, but this strategy has produced only modest results in animal models. The link between PrP(Sc) and neurotoxicity is unclear, and alternative pathological processes need to be considered. Here we focus on the latest progress in therapeutic strategies and potential mechanisms of prion neurotoxicity. RECENT FINDINGS: Passive immunisation with anti-prion protein antibodies prevents peripheral prion replication and blocks progression to clinical disease in peripherally infected mice. A new approach, in which neuronal cellular prion protein is depleted in mice with established neuroinvasive prion infection, prevents the onset of clinical disease, blocks neuronal cell loss and reverses early spongiform pathology. This dramatic protective effect occurs despite the continued build-up of extraneuronal PrP(Sc) and continued replication of prion infectivity, effectively producing a sub-clinical state. SUMMARY: New insights into the mechanisms of neurotoxicity in prion diseases support the concept that PrP(Sc) itself is not directly neurotoxic. They suggest that neuronal prion propagation results in the production of a toxic intermediate or depletion of a key constituent. Prevention of the formation of such a species rather than PrP(Sc) accumulation itself is a clear target for prion therapeutics.     

Creutzfeldt-Jakob disease in Hungary

Human prion diseases or transmissible spongiform encephalopathies are progressive fatal neuropsychiatric diseases. In addition to the evaluation of clinical features, a common diagnostic procedure includes examination of the protein 14-3-3 in the cerebrospinal fluid, performing EEG to detect periodic sharp wave complexes with triphasic morphology, and cranial MRI to demonstrate high signal intensity in the basal ganglia or thalamus. The definite diagnosis requires a neuropathological examination. The analysis of the prion protein gene (PRNP) is initiated mainly after suspicion of a positive family history or an atypical presentation. In Hungary collecting data and setting up the neuropathological diagnosis in suspect prion disease cases originates from the late 1960s. Systematic surveillance was established in 1994 and since 2001 reporting of Creutzfeldt-Jakob disease has been compulsory. According to our database, the incidence of genetic prion disease is increased in Hungary. The most frequent mutation in the PRNP is at codon 200. This might be linked to migration from the Slovakian focus. Acquired forms of prion disease were not detected in our country. The surveillance system is based on referrals from clinicians and pathologists and the aim is to perform the neuropathological examination and analysis of the PRNP on the majority of suspect cases.     

Creutzfeldt-Jakob disease and scrapie prions

Creutzfeldt-Jakob disease, kuru, and Gerstmann-Str?ussler syndrome are transmissible degenerative diseases of the central nervous system caused by novel infectious pathogens designated prions. Scrapie is a neurodegenerative disease of sheep and goats and is also caused by prions. Experimental scrapie has been extensively studied in hamsters and mice. The scrapie prion protein (PrPSc) is the only component of the infectious scrapie prion identified, to date. Scrapie infectivity and PrPSc copartition into membranes, rods, and liposomes raising the possibility that only PrPSc might be required for infection; however, a second component such as a small nucleic acid cannot be eliminated. PrPSc is encoded by a single copy cellular gene and not by a hypothetical nucleic acid within purified prion preparations. Normal, uninfected cells express the cellular prion protein (PrPc). Both PrPSc and PrPc appear to be translated from the same 2.1-kb mRNA. The N-terminal amino acid sequences of hamster PrPC and PrPSc are identical; both correspond to that predicted by the translated prion protein (PrP) gene sequence. While the chemical difference between PrPc and PrPSc remains unknown, the organization of the PrP gene argues that it results from a posttranslational event. Six posttranslational modifications of both PrP isoforms have been identified: (1) cleavage of an N-terminal signal peptide, (2) an intramolecular disulfide bond, (3) an N-linked oligosaccharide attached to Asn 181, (4) a second oligosaccharide attached to Asn 197, (5) cleavage of a C-terminal hydrophobic peptide, and (6) a phosphatidylinositol glycolipid attached to the C-terminus. The mouse PrP gene is on chromosome 2 and is linked to a gene controlling the scrapie incubation time (Prn-i). PrP genes from inbred mice with short and long incubation times differ by two amino acids, a finding consistent with but not proving that PrP modulates susceptibility to scrapie. PrPSc stimulation of a posttranslational process which converts PrPc or its precursor into PrPSc is one possible mechanism for prion replication. This is consistent with observations showing that human prion diseases are manifest as infectious, sporadic and genetic disorders.     

Elevated manganese levels in blood and CNS in human prion disease

Prion disease or transmissible spongiform encephalopathies are neurodegenerative disorders of humans and other mammals. They are fatal and difficult to diagnose. Previous studies have suggested that some prion diseases cause elevation of manganese in the blood and brain. In the current study we analysed blood and brain samples from humans to determine whether elevation in manganese is a specific characteristic of Creutzfeldt-Jakob disease, the most common form of human prion disease. Analysis of manganese in the blood of normal humans showed that concentrations vary little with age or sex. Analysis of other diseases, including other neurodegenerative disease showed that only CJD showed an elevation in manganese and copper. Other diseases that showed elevated manganese included blood-brain barrier disorders and haemochromatosis. However, CJD could be easily distinguished from these diseases. This implies that increased blood manganese in prion disease is a highly specific characteristic of the disease.     

Creutzfeldt-Jakob disease and other human transmissible spongiform encephalopathies. Part I

In the first part of this work the main problems of prion diseases--also called transmissible cerebral amyloidoses (TCA) or subacute (transmissible) encephalopathies (SSE, TSE)--and clinical symptoms of Creutzfeldt-Jakob disease are presented. Some problems of neuropathology of Creutzfeldt-Jakob disease and basic informations about other human prion diseases will be presented in the second part. The growth of the interest in prion diseases during last years is caused by the problem of bovine spongiform encephalopathy (BSE or "mad cow disease") and its transmission into a human. The new variant of Creutzfeldt-Jakob disease (nvCJD) has appeared. Prion diseases: Gerstmann-Str?ussler-Scheinker syndrome (GSS), kuru, fatal familial insomnia (FFI) and particularly the most frequent of them--Creutzfeldt-Jakob disease (CJD)--have nonspecific, sometimes variable clinical (psychopathological and neurological) symptoms. The imaging, EEG, cerebrospinal fluid tests and other laboratory tests are not specific either and their diagnostic value is limited. Neuropathological studies are needed but their interpretation is often difficult. The only certain diagnostic marker for TSE is the presence of PrP(Sc), the prion protein, which is presently believed to be a direct cause for all transmissible cerebral amyloidoses (TCA).     

Prion disease with a 144 base pair insertion: unusual cerebellar prion protein immunoreactivity

Sporadic, acquired, and genetic human prion diseases are characterized neuropathologically by distinct deposition patterns of the abnormal, disease-associated form of the prion protein (PrP^sc). In addition to mutations in the prion protein gene (PRNP), PrP^sc immunostaining patterns correlate with molecular phenotypes of prion diseases defined by the PRNP polymorphism at codon 129 and with protease-resistant PrP classified by Western blotting. Some point or insertional PRNP mutations share similar clinical and neuropathological phenotypes, whereas others show great variability even within the same family. Here we report a patient who presented clinically as sporadic Creutzfeldt-Jakob disease (CJD). Histologically moderate spongiform change was seen in cerebral and cerebellar cortical areas. Neuronal loss was restricted mainly to the occipital cortex and the basal ganglia. Surprisingly, numerous eosinophilic globular structures were noted in the molecular layer and the parahippocampal gyrus. These globules showed intense PrP immunopositivity using anti-PrP antibodies against different epitopes. They were stained with PAS but lacked congophilia and birefringence in polarized light. Ultrastructurally, globules were composed of 21-nm-thick intermingled filaments without dense core. Genetic analysis revealed a PRNP 144 base pair insertion. Our case reinforces the importance of molecular genetic diagnosis, especially in those patients who lack a family history of prion disease and show unusual neuropathological changes. It also widens the phenotypic spectrum of prion diseases. The phenotypic variability within the same mutation suggests further, yet uncharacterized, genetic or epigenetic influence on phenotype in these diseases.     

Decreased cell surface prion protein in mouse models of prion disease

Transmissible spongiform encephalopathies are infectious neurodegenerative diseases caused by prions, composed of ordered aggregates of misfolded cellular prion protein. Neural antigen density of prion protein, Thy-1 and glial fibrillary acidic protein was analyzed using flow cytometry of dissociated mouse brain cells after inoculation with mouse-adapted transmissible spongiform encephalopathy agents. Transmissible spongiform encephalopathy gliosis was demonstrated by increased intracellular immunoreactivity for glial fibrillary acidic protein compared with controls. Immunoreactivity for cell surface prion protein was reduced 2.8-3.8-fold compared with control brain cells, whereas surface Thy-1 protein was reduced 1.5-4-fold. Double-staining protocols revealed loss of brain cells highly immunoreactive for prion protein and Thy-1, with a preferential reduction of prion protein, suggesting that prion protein expression, trafficking or consumption may be affected early in disease.     

Molecular pathology, classification, and diagnosis of sporadic human prion disease variants

Human prion diseases are a unique group of transmissible neurodegenerative diseases that occur as sporadic, familial or acquired disorders and show a wide range of phenotypic variation. The latter has been attributed to the existence of distinct strains of the agent or prion, and the genetic background of the host, namely the primary sequence of the gene encoding the prion protein, which is the site of mutations and polymorphisms. The characterization of distinct isoforms of the abnormal prion protein in the brain of affected patients, which has been shown to correlate with the disease phenotype, has recently led to the concept of molecular strain typing, in which the different prion protein isoforms or "types", possibly enciphering the strain variability in their conformation, may serve as surrogate markers for individual prion strains. In sporadic Creutzfeldt-Jakob disease, the most common human prion disease, there are at least six distinct clinico-pathological disease phenotypes that largely correlate at a molecular level with two prion protein types with distinctive physicochemical properties and the genotype at the methionine/valine polymorphic codon 129 in the prion protein gene. Recent results of transmission studies indicate that five prion strains with distinctive biological properties can be isolated from these six disease variants. It has also been shown that about a third of sporadic cases show a mixed phenotype and the co-occurrence of prion protein types. The origin of prion strains and their co-occurrence as well as the mechanisms underlying the strain-specific neuronal targeting remain largely unexplained and their understanding constitute, together with the development of successful therapies and more sensitive and specific clinical biomarkers, the major challenges that this disease poses for the future.     

What went wrong in BSE? From prion disease to public disaster

The recent report of 10 cases of a new variant of Creutzfeldt-Jakob disease (CJD) which could be related to bovine spongiform encephalopathy (BSE) has precipitated alarm throughout Europe. The beef trade in the UK has collapsed and the European beef market has been seriously damaged. What went wrong? Much of the difficulty of handling the BSE epidemic arose from the 4–5 year incubation period which made it difficult to ascertain whether measures taken to contain the epidemic had been effective. Public consternation and scientific equivocation arose because these prion diseases are unlike any other group of infectious diseases. Rather than being caused by a conventional micro-organism, the primary pathogenic event consists of the transformation of a normal protein (the prion protein) into an abnormal form, which can transmit disease. Prion disease is endemic in humans and sheep where it is associated with polymorphisms or mutations within the prion protein gene. Although the disease in these cases arises spontaneously, it produces an infectious prion protein. Under certain circumstances, abnormal prion protein contaminates other animals or humans resulting in epidemics of acquired prion disease. This review describes the events of the BSE epidemic and considers the difficulties in assessing the current risk to human health.     

Neuronal apoptosis in Creutzfeldt-Jakob disease

Neuronal loss is a salient feature of prion diseases; however, its causes and mechanisms are unclear The possibility that it could occur through an apoptotic process has been postulated and is consistent with the lack of inflammation in prion disorders as supported by experimental studies. In order to test this hypothesis in humans, we examined samples of frontal and temporal cerebral cortex, striatum, thalamus, and cerebellum from 16 patients who died from Creutzfeldt-Jakob disease. They included 5 sporadic cases, 5 familial, 3 iatrogenic, and 3 cases with the new variant. These were compared with age and sex matched controls. Using in situ end labelling, we identified apoptotic neurons in all the cases of Creutzfeldt-Jakob disease. A single labelled neuron was found in the eldest control. Apoptotic neurons were mostly found in damaged regions and their presence and abundance seemed to correlate closely with neuronal loss. This supports the view that apoptosis of neurons is a feature of prion diseases and may contribute to the neuronal loss which is one of the main characteristics of these conditions. Neuronal apoptosis also correlated well with microglial activation, as demonstrated by the expression of major histocompatibility complex class II antigens, and axonal damage, as identified by beta-amyloid protein precursor immunostaining. In contrast, we found no obvious relationship between the topography and severity of neuronal apoptosis and the type, topography, and abundance of prion protein deposits as demonstrated by immunocytochemistry.     

Synaptic pathology and cell death in the cerebellum in Creutzfeldt-Jakob disease

Prion protein (PrP(c)) is a cell membrane glycoprotein particularly abundant in the synapses. Prion diseases are characterized by the replacement of the normal PrPc by a protease-resistant, sheet-containing isoform (PrP(CJD), PrP(Sc), PrP(BSE)) that is pathogenic. Creutzfeldt-Jakob disease (CJD) in humans, scrapie (Sc) in sheep and goats, and bovine spongiform encephalopathy (BSE) in cattle are typical prion diseases. Classical CJD can be presented as sporadic, infectious or familial, whereas the new variant of CJD (nvCJD) is considered a BSE-derived human disease. Spongiform degeneration, glial proliferation, involving astrocytes and microglia, neuron loss and abnormal PrP deposition are the main neuopathological findings in most human and animal prion diseases. Yet recent data point to synapses as principal targets of abnormal PrP deposition. Loss of synapses is an early abnormality in experimental scrapie. Decreased expression of crucial proteins linked to exocytosis and neurotransmission, covering synaptophysin, synaptosomal-associated protein of 25,000 mol wt (SNAP-25), synapsins, syntaxins and Rab3a occurs in the cerebral cortex and cerebellum in sporadic CJD. Moreover, impairment of glomerular synapses and attenuation of parallel fiber pre-synaptic terminals on Purkinje cell dendrites is a cardinal consequence of abnormal PrP metabolism in CJD. Accumulation of synaptic proteins in the soma and axonal torpedoes of Purkinje cells suggests additional impairment of axonal transport. Increase in nuclear DNA vulnerability leading to augmented numbers of cells bearing nuclear DNA fragments is a common feature in the brains of humans affected by prion diseases examined at post-mortem, but also in archival biopsy samples processed with the method of in situ end-labeling of nuclear DNA fragmentation. This form of cell death is reminiscent of apoptosis found in experimental scrapie in rodents. It is not clear that all forms of cell death in human and animal prion diseases are due to apoptosis. Yet new observations have shown cleaved (active) caspase-3 (17 kDa), a main executioner of apoptosis, expressed in scattered cells in the brains of mice with experimental scrapie and in the cerebellum of patients with sporadic CJD. Together, these data suggest activation of the caspase pathway of apoptosis in human and animal prion diseases.     

A novel mutation (G114V) in the prion protein gene in a family with inherited prion disease

Inherited prion diseases are characterized by mutations in the PRNP gene encoding the prion protein (PrP). We report a novel missense mutation in the PRNP gene (resulting in a G114V mutation in PrP) in members of a Uruguayan family with clinical and histopathologic features of prion disease. Affected individuals were characterized by an early age at onset, initial neuropsychiatric symptoms, late dementia with prominent pyramidal and extrapyramidal symptoms, and long disease duration.