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Atypical Scrapie, which is not linked to epidemics, is assumed to be an idiopathic spontaneous prion disease in small ruminants. Therefore, its occurrence is unlikely to be controlled through selective breeding or other strategies as it is done for classical scrapie outbreaks. Its spontaneous nature and its sporadic incidence worldwide is reminiscent of the incidence of idiopathic spontaneous prion diseases in humans, which account for more than 85% of the cases in humans. Hence, developing animal models that consistently reproduce this phenomenon of spontaneous PrP misfolding, is of importance to study the pathobiology of idiopathic spontaneous prion disorders. Transgenic mice overexpressing sheep PrP C with I112 polymorphism (TgShI112, 1-2 × PrP levels compared to sheep brain) manifest clinical signs of a spongiform encephalopathy spontaneously as early as 380 days of age. The brains of these animals show the neuropathological hallmarks of prion disease and biochemical analyses of the misfolded prion protein show a ladder-like PrP res pattern with a predominant 7-10 kDa band. Brain homogenates from spontaneously diseased transgenic mice were inoculated in several models to assess their transmissibility and characterize the prion strain generated: TgShI112 (ovine I112 ARQ PrP C ), Tg338 (ovine VRQ PrP C ), Tg501 (ovine ARQ PrP C ), Tg340 (human M129 PrP C ), Tg361 (human V129 PrP C ), TgVole (bank vole I109 PrP C ), bank vole (I109I PrP C ), and sheep (AHQ/ARR and AHQ/AHQ churra-tensina breeds). Our analysis of the results of these bioassays concludes that the strain generated in this model is indistinguishable to that causing atypical scrapie (Nor98). Thus, we present the first faithful model for a bona fide, transmissible, ovine, atypical scrapie prion disease., (© 2022. The Author(s).)

Background: More than 40 pathogenic heterozygous PRNP mutations causing inherited prion diseases have been identified to date. Recessive inherited prion disease has not been described to date., Methods: We describe the clinical and neuropathological data of inherited early-onset prion disease caused by the rare PRNP homozygous mutation R136S. In vitro PrP Sc propagation studies were performed using recombinant-adapted protein misfolding cyclic amplification technique. Brain material from two R136S homozygous patients was intracranially inoculated in TgMet129 and TgVal129 transgenic mice to assess the transmissibility of this rare inherited form of prion disease., Results: The index case presented symptoms of early-onset dementia beginning at the age of 49 and died at the age of 53. Neuropathological evaluation of the proband revealed abundant multicentric PrP plaques and Western blotting revealed a ~ 8 kDa protease-resistant, unglycosylated PrP Sc fragment, consistent with a Gerstmann-Sträussler-Scheinker phenotype. Her youngest sibling suffered from progressive cognitive decline, motor impairment, and myoclonus with onset in her late 30s and died at the age of 48. Genetic analysis revealed the presence of the R136S mutation in homozygosis in the two affected subjects linked to homozygous methionine at codon 129. One sibling carrying the heterozygous R136S mutation, linked to homozygous methionine at codon 129, is still asymptomatic at the age of 74. The inoculation of human brain homogenates from our index case and an independent case from a Portuguese family with the same mutation in transgenic mice expressing human PrP and in vitro propagation of PrP Sc studies failed to show disease transmissibility., Conclusion: In conclusion, biallelic R136S substitution is a rare variant that produces inherited early-onset human prion disease with a Gerstmann-Sträussler-Scheinker neuropathological and molecular signature. Even if the R136S variant is predicted to be "probably damaging", heterozygous carriers are protected, at least from an early onset providing evidence for a potentially recessive pattern of inheritance in human prion diseases., (© 2021. The Author(s).)

E/D 163 polymorphism of dog prion protein (PrP) has been recently proposed as the variant responsible for canid prion resistance. To further investigate the protective role of this variant against prion replication, the transgenic mouse model OvPrP-Tg532 expressing sheep/goat PrP carrying the substitution D 162 (equivalent to D 163 position of dog PrP) was generated and intracranially inoculated with a broad collection of small ruminant prion strains. OvPrP-Tg532 mice showed resistance to classical bovine spongiform encephalopathy (BSE) from sheep and some classical scrapie isolates from sheep and goat but were susceptible to ovine atypical L-BSE and numerous classical scrapie isolates. Strikingly, some of these classical scrapie isolates showed a shift in their prion strain properties. These results suggest that other PrP residues apart from E/D 163 variant of dog PrP or factors distinct than PrP may participate in prion resistance of canids and that different factors may be required for D 162 sheep PrP to provide effective protection to sheep against ruminant prions., (© 2021. The Author(s).)

Aims: The amino acid sequence of prion protein (PrP) is a key determinant in the transmissibility of prion diseases. While PrP sequence is highly conserved among mammalian species, minor changes in the PrP amino acid sequence may confer alterations in the transmissibility of prion diseases. Classical bovine spongiform encephalopathy (C-BSE) is the only zoonotic prion strain reported to date causing variant Creutzfeldt-Jacob disease (vCJD) in humans, although experimental transmission points to atypical L-BSE and some classical scrapie isolates as also zoonotic. The precise molecular elements in the human PrP sequence that limit the transmissibility of prion strains such as sheep/goat scrapie or cervid chronic wasting disease (CWD) are not well known., Methods: The transmissibility of a panel of diverse prions from different species was compared in transgenic mice expressing either wild-type human PrP C (MDE-HuTg340) or a mutated human PrP C harbouring Val 166 -Gln 168 amino acid changes (VDQ-HuTg372) in the β2-α2 loop instead of Met 166 -Glu 168 wild-type variants., Results: VDQ-HuTg372 mice were more susceptible to prions than MDE-HuTg340 mice in a strain-dependent manner., Conclusions: Met 166 -Glu 168 amino acid residues present in wild-type human PrP C are molecular determinants that limit the propagation of most prion strains assayed in the human PrP context., (© 2020 The Authors. Neuropathology and Applied Neurobiology published by John Wiley & Sons Ltd on behalf of British Neuropathological Society.)

Triggering receptor expressed on myeloid cells 2 (TREM2) is an innate immune cell surface receptor that regulates microglial function and is involved in the pathophysiology of several neurodegenerative diseases. Its soluble form (sTREM2) results from shedding of the TREM2 ectodomain. The role of TREM2 in prion diseases, a group of rapidly progressive dementias remains to be elucidated. In the present study, we analysed the expression of TREM2 and its main sheddase ADAM10 in the brain of sporadic Creutzfeldt-Jakob disease (sCJD) patients and evaluated the role of CSF and plasma sTREM2 as a potential diagnostic marker of prion disease. Our data indicate that, compared to controls, TREM2 is increased in sCJD patient brains at the mRNA and protein levels in a regional and subtype dependent fashion, and expressed in a subpopulation of microglia. In contrast, ADAM10 is increased at the protein, but not the mRNA level, with a restricted neuronal expression. Elevated CSF sTREM2 is found in sCJD, genetic CJD with mutations E200K and V210I in the prion protein gene (PRNP), and iatrogenic CJD, as compared to healthy controls (HC) (AUC = 0.78-0.90) and neurological controls (AUC = 0.73-0.85), while CSF sTREM2 is unchanged in fatal familial insomnia. sTREM2 in the CSF of cases with Alzheimer's disease, and multiple sclerosis was not significantly altered in our series. CSF sTREM2 concentrations in sCJD are PRNP codon 129 and subtype-related, correlate with CSF 14-3-3 positivity, total-tau and YKL-40, and increase with disease progression. In plasma, sTREM2 is increased in sCJD compared with HC (AUC = 0.80), displaying positive correlations with plasma total-tau, neurofilament light, and YKL-40. We conclude that comparative study of TREM2 in brain and biological fluids of prion diseases reveals TREM2 to be altered in human prion diseases with a potential value in target engagement, patient stratification, and disease monitoring.

MicroRNAs (miRNAs) may contribute to the development and pathology of many neurodegenerative diseases, including prion diseases. They are also promising biomarker candidates due to their stability in body fluids. We investigated miRNA alterations in a Tg501 mouse model of prion diseases that expresses a transgene encoding the goat prion protein ( PRNP ). Tg501 mice intracranially inoculated with mouse-adapted goat scrapie were compared with age-matched, mock inoculated controls in preclinical and clinical stages. Small RNA sequencing from the cervical spinal cord indicated that miR-223-3p, miR-151-3p, and miR-144-5p were dysregulated in scrapie-inoculated animals before the onset of symptoms. In clinical-stage animals, 23 significant miRNA alterations were found. These miRNAs were predicted to modify the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways including prion disease, extracellular matrix interactions, glutaminergic synapse, axon guidance, and transforming growth factor-beta signaling. MicroRNAs miR-146a-5p (up in cervical spinal cord) and miR-342-3p (down in cervical spinal cord, cerebellum and plasma), both indicated in neurodegenerative diseases earlier, were verified by quantitative real-time polymerase chain reaction (qRT-PCR). Minimal changes observed before the disease onset suggests that most miRNA alterations observed here are driven by advanced prion-associated pathology, possibly limiting their use as diagnostic markers. However, the results encourage further mechanistic studies on miRNA-regulated pathways involved in these neurodegenerative conditions.

Prion replication results from the autocatalytic templated assisted conversion of the host-encoded prion protein PrP C into misfolded, polydisperse PrP Sc conformers. Structurally distinct PrP Sc conformers can give rise to multiple prion strains. Within and between prion strains, the biological activity (replicative efficacy and specific infectivity) of PrP Sc assemblies is size dependent and thus reflects an intrinsic structural heterogeneity. The contribution of such PrP Sc heterogeneity across species prion adaptation, which is believed to be based on fit adjustment between PrP Sc template(s) and host PrP C , has not been explored. To define the structural-to-fitness PrP Sc landscape, we measured the relative capacity of size-fractionated PrP Sc assemblies from different prion strains to cross mounting species barriers in transgenic mice expressing foreign PrP C . In the absence of a transmission barrier, the relative efficacy of the isolated PrP Sc assemblies to induce the disease is like the efficacy observed in the homotypic context. However, in the presence of a transmission barrier, size fractionation overtly delays and even abrogates prion pathogenesis in both the brain and spleen tissues, independently of the infectivity load of the isolated assemblies. Altering by serial dilution PrP Sc assembly content of non-fractionated inocula aberrantly reduces their specific infectivity, solely in the presence of a transmission barrier. This suggests that synergy between structurally distinct PrP Sc assemblies in the inoculum is requested for crossing the species barrier. Our data support a mechanism whereby overcoming prion species barrier requires complementation between structurally distinct PrP Sc assemblies. This work provides key insight into the "quasispecies" concept applied to prions, which would not necessarily rely on prion substrains as constituent but on structural PrP Sc heterogeneity within prion population.

Classical bovine spongiform encephalopathy (BSE) is the only zoonotic prion disease described to date. Although the zoonotic potential of atypical BSE prions have been partially studied, an extensive analysis is still needed. We conducted a systematic study by inoculating atypical BSE isolates from different countries in Europe into transgenic mice overexpressing human prion protein (PrP): TgMet 129 , TgMet/Val 129 , and TgVal 129 . L-type BSE showed a higher zoonotic potential in TgMet 129 mice than classical BSE, whereas Val 129 -PrP variant was a strong molecular protector against L-type BSE prions, even in heterozygosis. H-type BSE could not be transmitted to any of the mice. We also adapted 1 H- and 1 L-type BSE isolate to sheep-PrP transgenic mice and inoculated them into human-PrP transgenic mice. Atypical BSE prions showed a modification in their zoonotic ability after adaptation to sheep-PrP producing agents able to infect TgMet 129 and TgVal 129 , bearing features that make them indistinguishable of sporadic Creutzfeldt-Jakob disease prions.

Prions are unique agents that challenge the molecular biology dogma by transmitting information on the protein level. They cause neurodegenerative diseases that lack of any cure or treatment called transmissible spongiform encephalopathies. The function of the normal form of the prion protein, the exact mechanism of prion propagation between species as well as at the cellular level and neuron degeneration remains elusive. However, great amount of information known for all these aspects has been achieved thanks to the use of animal models and more precisely to transgenic mouse models. In this chapter, the main contributions of these powerful research tools in the prion field are revised., (© 2020 Elsevier Inc. All rights reserved.)

Cynomolgus macaque has been used for the evaluation of the zoonotic potential of prion diseases, especially for classical-Bovine Spongiform Encephalopathy (classical-BSE) infectious agent. PrP amino acid sequence is considered to play a key role in the susceptibility to prion strains and only one amino acid change may alter this susceptibility. Macaque and human-PrP sequences have only nine amino acid differences, but the effect of these amino acid changes in the susceptibility to dissimilar prion strains is unknown. In this work, the transmissibility of a panel of different prions from several species was compared in transgenic mice expressing either macaque-PrP C (TgMac) or human-PrP C (Hu-Tg340). Similarities in the transmissibility of most prion strains were observed suggesting that macaque is an adequate model for the evaluation of human susceptibility to most of the prion strains tested. Interestingly, TgMac were more susceptible to classical-BSE strain infection than Hu-Tg340. This differential susceptibility to classical-BSE transmission should be taken into account for the interpretation of the results obtained in macaques. It could notably explain why the macaque model turned out to be so efficient (worst case model) until now to model human situation towards classical-BSE despite the limited number of animals inoculated in the laboratory experiments.

Prion diseases are fatal neurodegenerative disorders caused by misfolding of the normal prion protein into an infectious cellular pathogen. Clinically characterized by rapidly progressive dementia and accounting for 85% of human prion disease cases, sporadic Creutzfeldt-Jakob disease (sCJD) is the prevalent human prion disease. Although sCJD neuropathological hallmarks are well-known, associated molecular alterations are elusive due to rapid progression and absence of preclinical stages. To investigate transcriptome alterations during disease progression, we utilized tg340- PRNP 129MM mice infected with postmortem material from sCJD patients of the most susceptible genotype (MM1 subtype), a sCJD model that faithfully recapitulates the molecular and pathological alterations of the human disease. Here we report that transcriptomic analyses from brain cortex in the context of disease progression, reveal epitranscriptomic alterations (specifically altered RNA edited pathway profiles, eg., ER stress, lysosome) that are characteristic and possibly protective mainly for preclinical and clinical disease stages. Our results implicate regulatory epitranscriptomic mechanisms in prion disease neuropathogenesis, whereby RNA-editing targets in a humanized sCJD mouse model were confirmed in pathological human autopsy material., Competing Interests: The authors declare no conflict of interest., (Copyright © 2019 the Author(s). Published by PNAS.)

Prion diseases are caused by the conversion of physiological PrP C into the pathogenic misfolded protein PrP Sc , conferring new properties to PrP Sc that vary upon prion strains. In this work, we analyze the thermostability of three prion strains (BSE, RML and 22L) that were heated at 98 °C for 2 hours. PrP Sc resistance to proteinase K (PrP res ), residual infectivity by mouse bioassay and in vitro templating activity by protein misfolding cyclic amplification (PMCA) were studied. Heated strains showed a huge loss of PrP res and a radically different infectivity loss: RML was the most thermolabile strain (6 to 7 log10 infectivity loss), followed by 22L (5 log10) while BSE was the most thermostable strain with low or null infectivity reduction showing a clear dissociation between PrP res and infectivity. These results indicate that thermostability is a strain-specific feature, measurable by PMCA and mouse bioassay, and a great tool to distinguish prion strains.

Among the ever-growing number of self-replicating proteins involved in neurodegenerative diseases, the prion protein PrP remains the most infamous for its central role in transmissible spongiform encephalopathies (TSEs). In these diseases, pathogenic prions propagate through a seeding mechanism, where normal PrP C molecules are converted into abnormally folded scrapie isoforms termed PrP Sc . Since its discovery over 30 years ago, much advance has contributed to define the host-encoded cellular prion protein PrP C as a critical relay of prion-induced neuronal cell demise. A current consensual view is that the conversion of PrP C into PrP Sc in neuronal cells diverts the former from its normal function with subsequent molecular alterations affecting synaptic plasticity. Here, we report that prion infection is associated with reduced expression of key effectors of the Notch pathway in vitro and in vivo, recapitulating changes fostered by the absence of PrP C . We further show that both prion infection and PrP C depletion promote drastic alterations in the expression of a defined set of Eph receptors and their ephrin ligands, which represent important players in synaptic function. Our data indicate that defects in the Notch and Eph axes can be mitigated in response to histone deacetylase inhibition in PrP C -depleted as well as prion-infected cells. We thus conclude that infectious prions cause a loss-of-function phenotype with respect to Notch and Eph signaling and that these alterations are sustained by epigenetic mechanisms.

Co-occurrence of different prion strains into the same host has been recognized as a natural phenomenon for several sporadic Creutzfeldt-Jakob disease (sCJD) patients and natural scrapie cases. The final outcome of prion coinfection is not easily predictable. In addition to the usual factors that influence prion conversion, the replication of one strain may entail positive or negative consequences to the other. The main aim of this study was to gain insights into the prion coinfection and interference concepts and their potential therapeutic implications. Here, different mouse models were challenged with several combinations of prion strains coupled on the basis of the lengths of their incubation periods and the existence/absence of a species barrier in the tested animal model. We found that nontransmissible strains can interfere the replication of fully transmissible strains when there is a species transmission barrier involved, as happened with the combination of a mouse (22L) and a human (sCJD) strain. However, this phenomenon seems to be strain dependent, since no interference was observed when the human strain coinoculated was vCJD. For the other combinations tested in this study, the results suggest that both strains replicate independently without effect on the replication of one over the other. It is common that the strain with more favorable conditions (e.g., a higher speed of disease development or the absence of a species barrier) ends being the only one detectable at the terminal stage of the disease. However, this does not exclude the replication of the least favored strain, leading to situations of the coexistence of prion strains. IMPORTANCE As a general conclusion, the outcome of prion coinfection is strongly dependent on the strain combination and the model utilized and is therefore difficult to predict. The coexistence of several prion strains may remain undetected if one of the strains has more favorable conditions to replicate in the host. The use of several models (such as a transgenic mouse expressing PrP from different species) to analyze field prion isolates is recommended to avoid this situation. The inference effect exerted by nonreplicative prion strains should be considered an interesting tool to advance in new therapeutic strategies for treating prion diseases; it may even be a proper therapeutic strategy., (Copyright © 2018 American Society for Microbiology.)

Reelin is an extracellular glycoprotein involved in key cellular processes in developing and adult nervous system, including regulation of neuronal migration, synapse formation, and plasticity. Most of these roles are mediated by the intracellular phosphorylation of disabled-1 (Dab1), an intracellular adaptor molecule, in turn mediated by binding Reelin to its receptors. Altered expression and glycosylation patterns of Reelin in cerebrospinal and cortical extracts have been reported in Alzheimer's disease. However, putative changes in Reelin are not described in natural prionopathies or experimental models of prion infection or toxicity. With this is mind, in the present study, we determined that Reelin protein and mRNA levels increased in CJD human samples and in mouse models of human prion disease in contrast to murine models of prion infection. However, changes in Reelin expression appeared only at late terminal stages of the disease, which prevent their use as an efficient diagnostic biomarker. In addition, increased Reelin in CJD and in in vitro models does not correlate with Dab1 phosphorylation, indicating failure in its intracellular signaling. Overall, these findings widen our understanding of the putative changes of Reelin in neurodegeneration.

Bank vole is a rodent species that shows differential susceptibility to the experimental transmission of different prion strains. In this work, the transmission features of a panel of diverse prions with distinct origins were assayed both in bank vole expressing methionine at codon 109 (Bv109M) and in transgenic mice expressing physiological levels of bank vole PrP C (the BvPrP-Tg407 mouse line). This work is the first systematic comparison of the transmission features of a collection of prion isolates, representing a panel of diverse prion strains, in a transgenic-mouse model and in its natural counterpart. The results showed very similar transmission properties in both the natural species and the transgenic-mouse model, demonstrating the key role of the PrP amino acid sequence in prion transmission susceptibility. However, differences in the PrP Sc types propagated by Bv109M and BvPrP-Tg407 suggest that host factors other than PrP C modulate prion strain features., Importance: The differential susceptibility of bank voles to prion strains can be modeled in transgenic mice, suggesting that this selective susceptibility is controlled by the vole PrP sequence alone rather than by other species-specific factors. Differences in the phenotypes observed after prion transmissions in bank voles and in the transgenic mice suggest that host factors other than the PrP C sequence may affect the selection of the substrain replicating in the animal model., (Copyright © 2016, American Society for Microbiology. All Rights Reserved.)

Bovine spongiform encephalopathy (BSE) created a global European crisis in the 1980s and 90s, with very serious health and economic implications. Classical BSE now appears to be under control, to a great extent as a result of a global research effort that identified the sources of prions in meat and bone meal (MBM) and developed new animal-testing tools that guided policy. Priority ( www.prionpriority.eu ) was a European Union (EU) Framework Program 7 (FP7)-funded project through which 21 European research institutions and small and medium enterprises (SMEs) joined efforts between 2009 and 2014, to conduct coordinated basic and applied research on prions and prion diseases. At the end of the project, the Priority consortium drafted a position paper ( www.prionpriority.eu/Priority position paper) with its main conclusions. In the present opinion paper, we summarize these conclusions. With respect to the issue of re-introducing ruminant protein into the feed-chain, our opinion is that sustaining an absolute ban on feeding ruminant protein to ruminants is essential. In particular, the spread and impact of non-classical forms of scrapie and BSE in ruminants is not fully understood and the risks cannot be estimated. Atypical prion agents will probably continue to represent the dominant form of prion diseases in the near future in Europe. Atypical L-type BSE has clear zoonotic potential, as demonstrated in experimental models. Similarly, there are now data indicating that the atypical scrapie agent can cross various species barriers. More epidemiological data from large cohorts are necessary to reach any conclusion on the impact of its transmissibility on public health. Re-evaluations of safety precautions may become necessary depending on the outcome of these studies. Intensified searching for molecular determinants of the species barrier is recommended, since this barrier is key for important policy areas and risk assessment. Understanding the structural basis for strains and the basis for adaptation of a strain to a new host will require continued fundamental research, also needed to understand mechanisms of prion transmission, replication and how they cause nervous system dysfunction and death. Early detection of prion infection, ideally at a preclinical stage, also remains crucial for development of effective treatment strategies.

Transmissible spongiform encephalopaties (TSEs) are fatal neurodegenerative diseases characterized by the aggregation and accumulation of the misfolded prion protein in the brain. Other proteins such as β-amyloid, tau or Serum Amyloid-A (SAA) seem to share with prions some aspects of their pathogenic mechanism; causing a variety of so called prion-like diseases in humans and/or animals such as Alzheimer's, Parkinson's, Huntington's, Type II diabetes mellitus or amyloidosis. The question remains whether these misfolding proteins have the ability to self-propagate and transmit in a similar manner to prions. In this review, we describe the prion and prion-like diseases affecting animals as well as the recent findings suggesting the prion-like transmissibility of certain non-prion proteins., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Variably protease-sensitive prionopathy (VPSPr), a recently identified and seemingly sporadic human prion disease, is distinct from Creutzfeldt-Jakob disease (CJD) but shares features of Gerstmann-Sträussler-Scheinker disease (GSS). However, contrary to exclusively inherited GSS, no prion protein (PrP) gene variations have been detected in VPSPr, suggesting that VPSPr might be the long-sought sporadic form of GSS. The VPSPr atypical features raised the issue of transmissibility, a prototypical property of prion diseases. We inoculated VPSPr brain homogenate into transgenic mice expressing various levels of human PrP (PrPC). On first passage, 54% of challenged mice showed histopathologic lesions, and 34% harbored abnormal PrP similar to that of VPSPr. Surprisingly, no prion disease was detected on second passage. We concluded that VPSPr is transmissible; thus, it is an authentic prion disease. However, we speculate that normal human PrPC is not an efficient conversion substrate (or mouse brain not a favorable environment) and therefore cannot sustain replication beyond the first passage.

Aims: Transmissible spongiform encephalopathies, also called prion diseases, are characterized by the cerebral accumulation of misfolded prion protein (PrP(SC) ) and subsequent neurodegeneration. However, despite considerable research effort, the molecular mechanisms underlying prion-induced neurodegeneration are poorly understood. Here, we explore the hypothesis that prions induce dysfunction of the PI3K/Akt/GSK-3 signalling pathway., Methods: We employed two parallel approaches. Using cell cultures derived from mouse primary neurones and from a human neuronal cell line, we identified common elements that were modified by the neurotoxic fragment of PrP(106-126) . These studies were then complemented by comparative analyses in a mouse model of prion infection., Results: The presence of a polymerized fragment of the prion protein (PrP(106-126) ) or of a prion strain altered PI3K-mediated signalling, as evidenced by Akt inhibition and GSK-3 activation. PI3K activation by the addition of insulin or the expression of a constitutively active Akt mutant restored normal levels of Akt and GSK-3 activity. These changes were correlated with a reduction in caspase activity and an increase in neuronal survival. Moreover, we found that activation of caspase 3, Erk and GSK-3 are common features of PrP(106-126) -mediated neurotoxicity in cellular systems and prion infection in the mouse cerebellum, while activation of caspase 12 and JNK was observed in cellular models., Conclusions: Our findings in cell culture and in vivo models of prion disease demonstrate marked alterations to the PI3K/Akt/GSK-3 pathway and suggest that two additional pathways contribute to PrP-induced neurotoxicity as responsible of JNK and caspase 12 activation., (© 2013 British Neuropathological Society.)

The specific characteristics of Transmissible Spongiform Encephalopathy (TSE) strains may be altered during passage across a species barrier. In this study we investigated the biochemical and biological characteristics of Bovine Spongiform Encephalopathy (BSE) after transmission in both natural host species (cattle, sheep, pigs and mice) and in transgenic mice overexpressing the corresponding cellular prion protein (PrPC) in comparison with other non-BSE related prions from the same species. After these passages, most features of the BSE agent remained unchanged. BSE-derived agents only showed slight modifications in the biochemical properties of the accumulated PrPSc, which were demonstrated to be reversible upon re-inoculation into transgenic mice expressing bovine-PrPC. Transmission experiments in transgenic mice expressing bovine, porcine or human-PrP revealed that all BSE-derived agents were transmitted with no or a weak transmission barrier. In contrast, a high species barrier was observed for the non-BSE related prions that harboured an identical PrP amino acid sequence, supporting the theory that the prion transmission barrier is modulated by strain properties (presumably conformation-dependent) rather than by PrP amino acid sequence differences between host and donor. As identical results were observed with prions propagated either in natural hosts or in transgenic mouse models, we postulate that the species barrier and its passage consequences are uniquely governed by the host PrPC sequence and not influenced by other host genetic factors. The results presented herein reinforce the idea that the BSE agent is highly promiscuous, infecting other species, maintaining its properties in the new species, and even increasing its capabilities to jump to other species including humans. These data are essential for the development of an accurate risk assessment for BSE.

We generated transgenic mice expressing bovine cellular prion protein (PrP(C)) with a leucine substitution at codon 113 (113L). This protein is homologous to human protein with mutation 102L, and its genetic link with Gerstmann-Sträussler-Scheinker syndrome has been established. This mutation in bovine PrP(C) causes a fully penetrant, lethal, spongiform encephalopathy. This genetic disease was transmitted by intracerebral inoculation of brain homogenate from ill mice expressing mutant bovine PrP to mice expressing wild-type bovine PrP, which indicated de novo generation of infectious prions. Our findings demonstrate that a single amino acid change in the PrP(C) sequence can induce spontaneous generation of an infectious prion disease that differs from all others identified in hosts expressing the same PrP(C) sequence. These observations support the view that a variety of infectious prion strains might spontaneously emerge in hosts displaying random genetic PrP(C) mutations.

Prion diseases, which are mostly represented in humans by Creutzfeldt-Jakob disease, are transmissible neurodegenerative disorders characterized by vacuolization and neuronal loss, as well as by the accumulation of an abnormal form of the prion protein. These disorders have yet no effective treatment, and drugs that block prion replication in vitro do not significantly slow down the progression of the disease when used in vivo at late stages. Cell therapy that has been already tested in other neurodegenerative disorders therefore represents an interesting alternative approach. In this study, we showed for the first time in prion diseases that intracerebral transplantation of fetal neural stem cells significantly extended both incubation and survival time. This result was dependant on the time window chosen for the engraftment and was obtained with both genetically modified and wild-type stem cells, therefore forging a path toward efficient stem cell therapy for human prion diseases.

Objective: The objective of the study is to report 2 new genotypic forms of protease-sensitive prionopathy (PSPr), a novel prion disease described in 2008, in 11 subjects all homozygous for valine at codon 129 of the prion protein (PrP) gene (129VV). The 2 new PSPr forms affect individuals who are either homozygous for methionine (129MM) or heterozygous for methionine/valine (129MV)., Methods: Fifteen affected subjects with 129MM, 129MV, and 129VV underwent comparative evaluation at the National Prion Disease Pathology Surveillance Center for clinical, histopathologic, immunohistochemical, genotypical, and PrP characteristics., Results: Disease duration (between 22 and 45 months) was significantly different in the 129VV and 129MV subjects. Most other phenotypic features along with the PrP electrophoretic profile were similar but distinguishable in the 3 129 genotypes. A major difference laid in the sensitivity to protease digestion of the disease-associated PrP, which was high in 129VV but much lower, or altogether lacking, in 129MV and 129MM. This difference prompted the substitution of the original designation with "variably protease-sensitive prionopathy" (VPSPr). None of the subjects had mutations in the PrP gene coding region., Interpretation: Because all 3 129 genotypes are involved, and are associated with distinguishable phenotypes, VPSPr becomes the second sporadic prion protein disease with this feature after Creutzfeldt-Jakob disease, originally reported in 1920. However, the characteristics of the abnormal prion protein suggest that VPSPr is different from typical prion diseases, and perhaps more akin to subtypes of Gerstmann-Sträussler-Scheinker disease.

Prion diseases are transmissible neurodegenerative disorders for which no therapeutic or prophylactic regimens exist. Passive immunization with appropriate antibodies directed against the cellular form of the prion protein (PrPC) can delay the onset of prion disease after peripheral infection, but mechanisms and parameters determining their in vivo efficacy remain unknown. In the present study, we characterized the main pharmacokinetic properties of anti-PrP antibodies in different mouse models expressing various levels of PrPC (Prnp(0/0), C57BL/6 and tga20 mice) in correlation with therapeutic effect. Plasma levels of free antibodies, total endogenous PrPC and PrPC-antibody complexes were monitored after a single intraperitoneal monoclonal antibody (mAb) injection. Efficacy in delaying PrPSc peripheral accumulation seemed to be associated with mAb capacity to form long-lasting complexes with endogenous PrPC in the plasma. In agreement with previous observations on cellular models of transmissible spongiform encephalopathy infection, we observed that injection of anti-PrP antibodies induced a large (up to 100-fold) increase in circulating PrPC. Finally, the most efficient antibody extended the lifespan of infected animals greatly. These results allowed us to define critical characteristics of anti-PrP mAbs associated with therapeutic efficacy and could constitute a useful reference for designing optimized passive immunotherapies for prion diseases.

In the present report, the selective detection of sheep PrP haplotypes by monoclonal antibody 2A11 is described. It is showed that the substitution of glutamine by arginine but not by histidine at ovine PrP position 171 abolishes completely the recognition of either PrP(c) or PrP(d) by mAb 2A11, in such a way that the application of this antibody allows the unambiguous discrimination of R(171) homozygotes. On the basis of the high resistance to classical scrapie and bovine spongiform encephalophaty (BSE) infection associated to the R(171) PrP haplotype, animals bearing the ARR allele are currently selected within the scrapie national plan initiated in Great Britain. A 2A11-based immuno enzymatic test have been developed and evaluated using a panel of plasma and sera from sheep of different PrP genotypes and breeds. The test allows the efficient discrimination of R(171) homozygotes, R(171) heterozygotes and non-R(171) carriers, therefore offering a rapid, cheap and easy to use alternative method to select sheep for their resistance to scrapie.

Transmissible spongiform encephalopathies (TSEs) can be ameliorated by prion protein (PrP)-specific antibodies, but active immunization is complicated by immune tolerance to the normal cellular host protein (PrP(C)). Here, we show that DNA immunization of wild-type mice can break immune tolerance against the prion protein, resulting in the induction of PrP-specific antibody and T-cell responses. PrP immunogenicity was increased by fusion to the lysosomal targeting signal from LIMPII (lysosomal integral membrane protein type II). Although mice immunized with a PrP-LIMPII DNA vaccine showed a dramatic delay in the onset of early disease signs after intracerebral challenge, immunization against PrP also had some deleterious effects. These results clearly confirm the feasibility of using active immunization to protect against TSEs and, in the absence of effective treatments, indicate a suitable alternative for combating the spread of these diseases.

Objective: We review the current knowledge relative to the role of different immune system components and their contribution the spread of prions throughout the infected organism., Development: During the last years research made on transmissible spongiform encephalopathies conducted to the assumption that changes in the structure of a determined protein (prion protein, PrP) may become self-propagative. The generally assumed finding of an abnormally folded protein as the causative agent, able to propagate in hosts different from it was originated, is changing our views of transmissible diseases. The concept of infection resulting from pathogen propagation within a susceptible host has been steadily linked to the ability of nucleic acids to replicate their own coded information. In the prionosis, term comprising all known prion disorders, the information seems to be coded within the own protein conformation. This phenomena, once again, challenges the well-established principles of molecular biology., Conclusions: Propagation of prions uses strategies not previously recognized, and depends upon prion conformation and/or specific host restrictions. Thus, certain sheep and goat prion strains (scrapie) or human (variant of Creutzfeldt-Jakob disease) invade the brain after an initial interaction with the immune system of affected individuals. The involvement of the lymphoreticular system or, at least, certain host's immune competence seems to be a prerequisite for neuro-invasion after a natural infection or peripheral inoculation. On the other hand, proper identification of immune cell types involved may open the possibility for post exposure prophylaxis.

Introduction: The transmissible spongiform encephalopathies affect several species of higher animals apart from man. Amongst these, undoubtedly the best known is that affecting cattle, since the association between consumption of beef and its derivatives and the appearance of a variant of Creutzfeldt-Jakob disease in humans has been established., Development: This type of pathology has been well known for many years in the ovine family, particularly in sheep and goats. In spite of the establishment of hypotheses linking the cause of the appearance of spongiform encephalopathy in cows with an interspecies jump of the disease between the ovine and bovine families, this has not yet been proved. Concomitant with the epidemic of bovine spongiform encephalopathy, other species of animals were reported to have been affected by an identical disease. These included farmed mink, deer, elks, cats and two species of exotic African ungulates (the nyala and the greater kudu). All cases were described in animals kept in captivity for human consumption. Thus it would seem that there was a common cause of the disease in all cases., Conclusion: In this paper we describe the most relevant aspects of the appearance of this disorder in animals, including the symptoms, epidemiology and pathology typical of each specific condition.