The prion protein protease sensitivity, stability and seeding activity in variably protease sensitive prionopathy brain tissue suggests molecular overlaps with sporadic Creutzfeldt-Jakob disease

INTRODUCTION: Variably protease sensitive prionopathy (VPSPr) is a recently described, sporadic human prion disease that is pathologically and biochemically distinct from the currently recognised sporadic Creutzfeldt-Jakob disease (sCJD) subtypes. The defining biochemical features of the abnormal form of the prion protein (PrP(Sc)) in VPSPr are increased sensitivity to proteolysis and the presence of an N- and C-terminally cleaved ~8 kDa protease resistant PrP(Sc) (PrP(res)) fragment. The biochemical and neuropathological profile of VPSPr has been proposed to resemble either Gerstmann–Sträussler–Scheinker syndrome (GSS) or familial CJD with the PRNP-V180I mutation. However, in some cases of VPSPr two protease resistant bands have been observed in Western blots that co-migrate with those of type 2 PrP(res), suggesting that a proportion of the PrP(Sc) present in VPSPr has properties similar to those of sCJD. RESULTS: Here, we have used conformation dependent immunoassay to confirm the presence of PrP(Sc) in VPSPr that is more protease sensitive compared with sCJD. However, CDI also shows that a proportion of PrP(Sc) in VPSPr resists PK digestion of its C-terminus, distinguishing it from GSS associated with ~8 kDa PrP(res), and showing similarity to sCJD. Intensive investigation of a single VPSPr case with frozen tissue from multiple brain regions shows a broad, region-specific spectrum of protease sensitivity and differential stability of PrP(Sc) in the absence of PK treatment. Finally, using protein misfolding cyclic amplification and real-time quaking induced conversion, we show that VPSPr PrP(Sc) has the potential to seed conversion in vitro and that seeding activity is dispersed through a broad range of aggregate sizes. We further propose that seeding activity is associated with the ~19 and ~23 kDa PrP(res) rather than the ~8 kDa fragment. CONCLUSIONS: Therefore, PrP(Sc) in VPSPr is heterogeneous in terms of protease sensitivity and stability to denaturation with the chaotrope GdnHCl and includes a proportion with similar properties to that found in sCJD. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s40478-014-0152-4) contains supplementary material, which is available to authorized users

Dental treatment and risk of variant CJD - a case control study

Abstract Objective: Knowledge of risk factors for variant CJD (vCJD) remains limited, but transmission of prion proteins via re-useable medical devices, including dental instruments, or enhanced susceptibility following trauma to the oral cavity is a concern. This study aimed to identify whether previous dental treatment is a risk factor for development of vCJD. Design: Case control study Methods: Risk factor questionnaires completed by interview with relatives of 130 vCJD patients and with relatives of 66 community and 53 hospital controls were examined by a dental surgeon. Responses regarding dental treatments were analysed. Results: We did not find a statistically significant excess of risk of vCJD associated with dental treatments with the exception of extractions in an unmatched analysis of vCJD cases with community controls (p=0.02). However, this result may be explained by multiple testing. Conclusions: This is the first published study to date to examine potential links between vCJD and dental treatment. There was no convincing evidence found of an increased risk of variant CJD associated with reported dental treatment. However, the power of the study is restricted by the number of vCJD cases to date and does not preclude the possibility that some cases have resulted from secondary transmission via dental procedures. Due to the limitations of the data available, more detailed analyses of dental records are required to fully exclude the possibility of transmission via dental treatment

Bioassay studies support the potential for latrogenic transmission of variant Creutzfeldt Jakob disease through dental procedures

Background: Evidence is required to quantify the potential risks of transmission of variant Creutzfeldt Jakob (vCJD) through dental procedures. Studies, using animal models relevant to vCJD, were performed to address two questions. Firstly, whether oral tissues could become infectious following dietary exposure to BSE? Secondly, would a vCJD-contaminated dental instrument be able to transmit disease to another patient? Methods: BSE-301V was used as a clinically relevant model for vCJD. VM-mice were challenged by injection of infected brain homogenate into the small intestine (Q1) or by five minute contact between a deliberately-contaminated dental file and the gingival margin (Q2). Ten tissues were collected from groups of challenged mice at three or four weekly intervals, respectively. Each tissue was pooled, homogenised and bioassayed in indicator mice. Findings: Challenge via the small intestine gave a transmission rate of 100% (mean incubation 157±17 days). Infectivity was found in both dental pulp and the gingival margin within 3 weeks of challenge and was observed in all tissues tested within the oral cavity before the appearance of clinical symptoms. Following exposure to deliberately contaminated dental files, 97% of mice developed clinical disease (mean incubation 234±33 days). Interpretation: Infectivity was higher than expected, in a wider range of oral tissues, than was allowed for in previous risk assessments. Disease was transmitted following transient exposure of the gingiva to a contaminated dental file. These observations provide evidence that dental procedures could be a route of cross-infection for vCJD and support the enforcement of single-use for certain dental instruments

Gastrointestinal stromal tumour and hypoglycemia in a Fjord pony: Case report

<p>Abstract</p> <p>Background</p> <p>Neoplasia may cause hypoglycemia in different species including the horse, but hypoglycemia has not previously been reported in the horse associated with gastrointestinal stromal tumours.</p> <p>Case presentation</p> <p>A case of a gastrointestinal stromal tumour in a Fjord pony with severe recurrent hypoglycemia is presented. The mechanism causing the hypoglycemia was not established.</p> <p>Conclusion</p> <p>This case indicates that a gastrointestinal stromal tumour may cause hypoglycemia also in the horse.</p

Rapidly progressive dementia with thalamic degeneration and peculiar cortical prion protein immunoreactivity, but absence of proteinase K resistant PrP: a new disease entity?

BACKGROUND: Human prion diseases are a group of rare fatal neurodegenerative conditions with well-developed clinical and neuropathological diagnostic criteria. Recent observations have expanded the spectrum of prion diseases beyond the classically recognized forms. RESULTS: In the present study we report six patients with a novel, apparently sporadic disease characterised by thalamic degeneration and rapidly progressive dementia (duration of illness 2-12 months; age at death: 55-81 years). Light and electron microscopic immunostaining for the prion protein (PrP) revealed a peculiar intraneuritic distribution in neocortical regions. Proteinase K resistant PrP (PrPres) was undetectable by Western blotting in frontal cortex from the three cases with frozen tissue, even after enrichment for PrPres by centrifugation or by phosphotungstic acid precipitation. Conformation-dependent immunoassay analysis using a range of PK digestion conditions (and no PK digestion) produced only very limited evidence of meaningful D-N (denatured/native) values, indicative of the presence of disease-associated PrP (PrPSc) in these cases, when the results were compared with appropriate negative control groups. CONCLUSIONS: Our observation expands the spectrum of conditions associated with rapidly progressive dementia and may have implications for the understanding of the pathogenesis of prion diseases

All clinically-relevant blood components transmit prion disease following a single blood transfusion: a sheep model of vCJD

Variant CJD (vCJD) is an incurable, infectious human disease, likely arising from the consumption of BSE-contaminated meat products. Whilst the epidemic appears to be waning, there is much concern that vCJD infection may be perpetuated in humans by the transfusion of contaminated blood products. Since 2004, several cases of transfusion-associated vCJD transmission have been reported and linked to blood collected from pre-clinically affected donors. Using an animal model in which the disease manifested resembles that of humans affected with vCJD, we examined which blood components used in human medicine are likely to pose the greatest risk of transmitting vCJD via transfusion. We collected two full units of blood from BSE-infected donor animals during the pre-clinical phase of infection. Using methods employed by transfusion services we prepared red cell concentrates, plasma and platelets units (including leucoreduced equivalents). Following transfusion, we showed that all components contain sufficient levels of infectivity to cause disease following only a single transfusion and also that leucoreduction did not prevent disease transmission. These data suggest that all blood components are vectors for prion disease transmission, and highlight the importance of multiple control measures to minimise the risk of human to human transmission of vCJD by blood transfusion

Prion Shedding from Olfactory Neurons into Nasal Secretions

This study investigated the role of prion infection of the olfactory mucosa in the shedding of prion infectivity into nasal secretions. Prion infection with the HY strain of the transmissible mink encephalopathy (TME) agent resulted in a prominent infection of the olfactory bulb and the olfactory sensory epithelium including the olfactory receptor neurons (ORNs) and vomeronasal receptor neurons (VRNs), whose axons comprise the two olfactory cranial nerves. A distinct glycoform of the disease-specific isoform of the prion protein, PrPSc, was found in the olfactory mucosa compared to the olfactory bulb, but the total amount of HY TME infectivity in the nasal turbinates was within 100-fold of the titer in the olfactory bulb. PrPSc co-localized with olfactory marker protein in the soma and dendrites of ORNs and VRNs and also with adenylyl cyclase III, which is present in the sensory cilia of ORNs that project into the lumen of the nasal airway. Nasal lavages from HY TME-infected hamsters contained prion titers as high as 103.9 median lethal doses per ml, which would be up to 500-fold more infectious in undiluted nasal fluids. These findings were confirmed using the rapid PrPSc amplification QuIC assay, indicating that nasal swabs have the potential to be used for prion diagnostics. These studies demonstrate that prion infection in the olfactory epithelium is likely due to retrograde spread from the olfactory bulb along the olfactory and vomeronasal axons to the soma, dendrites, and cilia of these peripheral neurons. Since prions can replicate to high levels in neurons, we propose that ORNs can release prion infectivity into nasal fluids. The continual turnover and replacement of mature ORNs throughout the adult lifespan may also contribute to prion shedding from the nasal passage and could play a role in transmission of natural prion diseases in domestic and free-ranging ruminants

A novel seven-octapeptide repeat insertion in the prion protein gene (PRNP) in a Dutch pedigree with Gerstmann–Sträussler–Scheinker disease phenotype: comparison with similar cases from the literature

Human prion diseases can be sporadic, inherited or acquired by infection and show considerable phenotypic heterogeneity. We describe the clinical, histopathological and pathological prion protein (PrPSc) characteristics of a Dutch family with a novel 7-octapeptide repeat insertion (7-OPRI) in PRNP, the gene encoding the prion protein (PrP). Clinical features were available in four, neuropathological features in three and biochemical characteristics in two members of this family. The clinical phenotype was characterized by slowly progressive cognitive decline, personality change, lethargy, depression with anxiety and panic attacks, apraxia and a hypokinetic-rigid syndrome. Neuropathological findings consisted of numerous multi- and unicentric amyloid plaques throughout the cerebrum and cerebellum with varying degrees of spongiform degeneration. Genetic and molecular studies were performed in two male family members. One of them was homozygous for valine and the other heterozygous for methionine and valine at codon 129 of PRNP. Sequence analysis identified a novel 168 bp insertion [R2–R2–R2–R2–R3g–R2–R2] in the octapeptide repeat region of PRNP. Both patients carried the mutation on the allele with valine at codon 129. Western blot analysis showed type 1 PrPSc in both patients and detected a smaller ~8 kDa PrPSc fragment in the cerebellum in one patient. The features of this Dutch kindred define an unusual neuropathological phenotype and a novel PRNP haplotype among the previously documented 7-OPRI mutations, further expanding the spectrum of genotype–phenotype correlations in inherited prion diseases

Impact of the clinical context on the 14-3-3 test for the diagnosis of sporadic CJD

BACKGROUND: The 14-3-3 test appears to be a valuable aid for the clinical diagnosis of sporadic Creutzfeldt-Jakob disease (sCJD) in selected populations. However, its usefulness in routine practice has been challenged. In this study, the influence of the clinical context on the performance of the 14-3-3 test for the diagnosis of sCJD is investigated through the analysis of a large prospective clinical series. METHODS: Six hundred seventy-two Spanish patients with clinically suspected sCJD were analyzed. Clinical classification at sample reception according to the World Health Organization's (WHO) criteria (excluding the 14-3-3 test result) was used to explore the influence of the clinical context on the pre-test probabilities, and positive (PPV) and negative (NPV) predictive values of the 14-3-3 test. RESULTS: Predictive values of the test varied greatly according to the initial clinical classification: PPV of 98.8%, 96.5% and 45.0%, and NPV of 26.1%, 66.6% and 100% for probable sCJDi (n = 115), possible sCJDi (n = 73) and non-sCJDi (n = 484) cases, respectively. According to multivariate and Bayesian analyses, these values represent an improvement of diagnostic certainty compared to clinical data alone. CONCLUSION: In three different contexts of sCJD suspicion, the 14-3-3 assay provides useful information complementary to clinical and electroencephalographic (EEG) data. The test is most useful supporting a clinical impression, whilst it may show deceptive when it is not in agreement with clinical data