Lipid Rafts Act as a Common Platform for Amyloid-ß Oligomer-Induced Alzheimer's Disease Pathology.

BACKGROUND: Amyloid-ß (Aß) oligomers induce the overproduction of phosphorylated tau and neurodegeneration. These cascades gradually cause cognitive impairment in Alzheimer's disease (AD). While each pathological event in AD has been studied in detail separately, the spatial and temporal relationships between pathological events in AD remain unclear. OBJECTIVE: We demonstrated that lipid rafts function as a common platform for the pathological cascades of AD. METHODS: Cellular and synaptosomal lipid rafts were prepared from the brains of Aß amyloid model mice (Tg2576 mice) and double transgenic mice (Tg2576 x TgTauP301L mice) and longitudinally analyzed. RESULTS: Aß dimers, the cellular prion protein (PrPc), and Aß dimer/PrPc complexes were detected in the lipid rafts. The levels of Fyn, the phosphorylated NR2B subunit of the N-methyl-D-aspartate receptor, glycogen synthase kinase 3ß, total tau, phosphorylated tau, and tau oligomers increased with Aß dimer accumulation in both the cellular and synaptosomal lipid rafts. Increases in the levels of these molecules were first seen at 6 months of age and corresponded with the early stages of Aß accumulation in the amyloid model mice. CONCLUSION: Lipid rafts act as a common platform for the progression of AD pathology. The findings of this study suggest a novel therapeutic approach to AD, involving the modification of lipid raft components and the inhibition of their roles in the sequential pathological events of AD.

A fatal familial insomnia patient newly diagnosed as having depression: A case report.

INTRODUCTION: Fatal familial insomnia (FFI) is a rare clinical case. The study was mainly to report the clinical symptoms and imaging and genetic characteristics of a FFI case with depression, with relevant literature summarized. PATIENT CONCERNS: A male, aged 57 years old, with mental disorders and progressive memory decline one year before admission. DIAGNOSIS: Clinical manifestations: he had obvious abnormal mental behavior, rapidly progressing dementia symptoms, stubborn insomnia, abnormal movements and laryngeal stridor after falling asleep at night. Imaging and genetic test results: the cranial magnetic resonance imaging showed frontal temporal lobe atrophy; the polysomnography results showed no effective sleep; the 14-3-3 test result of cerebrospinal fluid was negative; the prion protein (PRNP) test showed that the D178N gene locus had mutations. And the patient was finally diagnosed as FFI. INTERVENTIONS: There were no obvious effects in the treatment using medicines such as Risperidone, Olanzapine, Alprazolam, Clonazepam, and Deanxit. OUTCOMES: Mobility dysfunction of the patient was further aggravated. He was no longer able to move around on his own, and there were serious mental disorders. CONCLUSION: PRNP examination is of guiding significance for the diagnosis of the FFI of depression. Hence, it is very necessary to perform PRNP examination in clinical diagnosis of FFI of depression.

Analysis of the Electrophoretic Profiles of Prion Protein in Carcinous and Pericarcinous Lysates of Six Different Types of Cancers.

OBJECTIVE: To find the different electrophoretic profiles of prion protein in carcinous and individual pericarcinous tissues in lysates of gastric, colon, liver, lung, thyroid, and laryngeal cancers. METHODS: Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot were used to test the amounts and electrophoretic patterns of total PrP and the tolerance of PK (protease K) digestion among six various cancer tissue types. RESULTS: A mass of PrP signals with a large molecular weight were identified in the homogenates of peripheral tissues. The amounts and electrophoretic patterns of total PrP did not differ significantly between carcinous and pericarcinous tissues. PrPs in all types of the tested cancer samples were PK sensitive but showed diversity in the tolerance of PK digestion among various tissue types. CONCLUSIONS: The study revealed that the included electrophoretic patterns of carcinous and pericarcinous tissues were almost similar. Unlike PrP-specific immunohistochemical assay, evaluation of PrP electrophoretic patterns in the peripheral organs and tissues by Western blot does not reflect tumor malignancy.

Biological Functions of the Intrinsically Disordered N-Terminal Domain of the Prion Protein: A Possible Role of Liquid-Liquid Phase Separation.

The mammalian prion protein (PrPC) is composed of a large intrinsically disordered N-terminal and a structured C-terminal domain, containing three alpha-helical regions and a short, two-stranded beta-sheet. Traditionally, the activity of a protein was linked to the ability of the polypeptide chain to adopt a stable secondary/tertiary structure. This concept has been extended when it became evident that intrinsically disordered domains (IDDs) can participate in a broad range of defined physiological activities and play a major functional role in several protein classes including transcription factors, scaffold proteins, and signaling molecules. This ability of IDDs to engage in a variety of supramolecular complexes may explain the large number of PrPC-interacting proteins described. Here, we summarize diverse physiological and pathophysiological activities that have been described for the unstructured N-terminal domain of PrPC. In particular, we focus on subdomains that have been conserved in evolution.

V180I genetic Creutzfeldt-Jakob disease with cardiac sympathetic nerve denervation masquerading as Parkinson's disease: A case report.

RATIONALE: Creutzfeldt-Jakob disease (CJD) with a point mutation of valine to isoleucine at codon 180 of the prion protein gene (V180I) is the most frequent form of genetic CJD in Japan. However, peripheral nerve involvement, especially cardiac sympathetic denervation, has not been investigated in cases with V180I genetic CJD.We herein report a genetically confirmed case of V180I genetic CJD presenting with parkinsonism and cardiac sympathetic nerve denervation. PATIENT CONCERNS: The patient was a 79-year-old Japanese woman who presented with subacute progressive gait disturbance and cognitive impairment. Clinical diagnosis of Parkinson's disease (PD) with mild cognitive impairment was initially suspected based on parkinsonism, such as bradykinesia, rigidity and tremor, and reduced accumulation of cardiac meta-iodobenzylguanidine (MIBG) scintigraphy. INTERVENTIONS: Based on parkinsonism and impaired cardiac MIBG findings, levodopa/decarboxylase inhibitor was administered up to 300 mg/day; however, her symptoms were not improved. OUTCOMES: Her motor and cognitive function progressively deteriorated. DIAGNOSIS: Although the patient had no family history of CJD, genetic CJD was diagnosed according to extensive hyperintensities in the bilateral cortices on diffusion-weighted magnetic resonance images, positive tau protein and 14-3-3 protein in the cerebrospinal fluid and a V180I mutation with methionine homozygosity at codon 129 by prion protein gene analysis. LESSONS: We should be aware that reduced uptake of cardiac MIBG scintigraphy in patients presenting with parkinsonism cannot confirm a diagnosis of PD. CJD should be considered when patients show a rapid progressive clinical course with atypical manifestations of PD.

Detection of cutaneous prion protein deposits could help diagnose GPI-anchorless prion disease with neuropathy.

BACKGROUND AND PURPOSE: To investigate prion protein (PrP) deposits in cutaneous tissues of patients of glycosylphosphatidylinositol (GPI)-anchorless prion diseases with neuropathy. METHODS: Cutaneous tissue samples from three patients with GPI-anchorless prion diseases were obtained, two cutaneous biopsy samples from the lower leg of Case 1 (Y162X) and Case 3 (D178fs25), and a cutaneous sample taken from the abdomen during an autopsy of Case 2 (D178fs25). We performed immunohistochemistry for PrP to look for abnormal PrP deposits. RESULTS: PrP deposits were observed in the dermal papilla, the sweat glands, the hair follicles, the arrector pili muscles, and peripheral nerves of all examined cases of GPI-anchorless prion disease with neuropathy. The abnormal PrP accumulation was frequently localized at the basement membrane, and colocalized with laminin. CONCLUSION: Immunohistochemical detection of PrP in cutaneous samples could be used to definitively diagnose GPI-anchorless PrP disease with neuropathy.

Effects of fibrinogen synthesis inhibition on vascular cognitive impairment during traumatic brain injury in mice.

Traumatic brain injury (TBI) is associated with increased blood content of fibrinogen (Fg), called hyperfibrinogenemia (HFg), which results in enhanced cerebrovascular permeability and leads to short-term memory (STM) reduction. Previously, we showed that extravasated Fg was deposited in the vasculo-astrocyte interface and was co-localized with cellular prion protein (PrPC) during mild-to-moderate TBI in mice. These effects were accompanied by neurodegeneration and STM reduction. However, there was no evidence presented that the described effects were the direct result of the HFg during TBI. We now present data indicating that inhibition of Fg synthesis can ameliorate TBI-induced cerebrovascular permeability and STM reduction. Cortical contusion injury (CCI) was induced in C57BL/6J mice. Then mice were treated with either Fg antisense oligonucleotide (Fg-ASO) or with control-ASO for two weeks. Cerebrovascular permeability to fluorescently labeled bovine serum albumin was assessed in cortical venules following evaluation of STM with memory assessement tests. Separately, brain samples were collected in order to define the expression of PrPC via Western blotting while deposition and co-localization of Fg and PrPC, as well as gene expression of inflammatory marker activating transcription factor 3 (ATF3), were characterized with real-time PCR. Results showed that inhibition of Fg synthesis with Fg-ASO reduced overexpression of AFT3, ameliorated enhanced cerebrovascular permeability, decreased expression of PrPC and Fg deposition, decreased formation of Fg-PrPC complexes in brain, and improved STM. These data provide direct evidence that a CCI-induced inflammation-mediated HFg could be a triggering mechanism involved in vascular cognitive impairment seen previously in our studies during mild-to-moderate TBI.

Analysis of the Electrophoretic Profiles of Prion Protein in Carcinous and Pericarcinous Lysates of Six Different Types of Cancers / 生物医学与环境科学（英文）

Objective@#To find the different electrophoretic profiles of prion protein in carcinous and individual pericarcinous tissues in lysates of gastric, colon, liver, lung, thyroid, and laryngeal cancers.@*Methods@#Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot were used to test the amounts and electrophoretic patterns of total PrP and the tolerance of PK (protease K) digestion among six various cancer tissue types.@*Results@#A mass of PrP signals with a large molecular weight were identified in the homogenates of peripheral tissues. The amounts and electrophoretic patterns of total PrP did not differ significantly between carcinous and pericarcinous tissues. PrPs in all types of the tested cancer samples were PK sensitive but showed diversity in the tolerance of PK digestion among various tissue types.@*Conclusions@#The study revealed that the included electrophoretic patterns of carcinous and pericarcinous tissues were almost similar. Unlike PrP-specific immunohistochemical assay, evaluation of PrP electrophoretic patterns in the peripheral organs and tissues by Western blot does not reflect tumor malignancy.

Transmission of CJD from nasal brushings but not spinal fluid or RT-QuIC product.

OBJECTIVE: The detection of prion seeding activity in CSF and olfactory mucosal brushings using real-time quaking-induced conversion assays allows highly accurate clinical diagnosis of sporadic Creutzfeldt-Jakob disease. To gauge transmission risks associated with these biospecimens and their testing, we have bioassayed prion infectivity levels in patients' brain tissue, nasal brushings, and CSF, and assessed the pathogenicity of amplified products of real-time quaking-induced conversion assays seeded with Creutzfeldt-Jakob disease prions. METHODS: We obtained olfactory mucosal brushings and CSF from patients with a final diagnosis of sporadic Creutzfeldt-Jakob disease subtype MM1 (n = 3). Samples were inoculated intracerebrally into Tg66 transgenic mice that overexpress the homologous human 129M prion protein. The mice were evaluated for clinical, neuropathological, and biochemical evidence of prion infection. RESULTS: Patients' brain tissue at 102 to 105 fold dilutions affected 47/48 Tg66 mice. In contrast, maximum acutely tolerable doses of insoluble pellets from their olfactory mucosa brushings caused evidence of prion disease in only 4/28 inoculated mice, and no effects were seen with 10-fold dilutions. No clinical prion disease was observed in mice inoculated with antemortem CSF samples or prion-seeded real-time quaking-induced conversion assay products. INTERPRETATION: Pellets from patients' olfactory mucosa brushings had ≥10,000-fold lower infectivity per unit volume than brain tissue, while CSF lacked detectable infectivity. Nonetheless, the results suggest that appropriate precautions may be warranted in surgical interventions involving the olfactory areas. The lack of pathogenic infectivity in the real-time quaking-induced conversion assay products provides evidence that the assay does not replicate biohazardous prions in vitro.

Rapid quantification of prion proteins using resistive pulse sensing.

Prion diseases are a group of fatal transmissible neurological conditions caused by the change in conformation of intrinsic cellular prion protein (PrPC). We present a rapid assay using aptamers and resistive pulse sensing, RPS, to extract and quantify PrPC from complex sample matrices. We functionalise the surface of superparamagnetic beads, SPBs, with a DNA aptamer. First SPB's termed P-beads, are used to pre-concentrate the analyte from a large sample volume. The PrPC protein is then eluted from the P-beads before aptamer modified sensing beads, S-beads, are added. The velocity of the S-beads through the nanopore reveals the concentration of the PrPC protein. The process is done in under an hour and allows the detection of picomol's of protein.

PrP (122-139) is a covert mitochondrial targeting signal of prion protein and it specifically triggers the perinuclear clustering of mitochondria in neuronal culture cells.

In many neurodegenerative diseases, mitochondria are actively involved in the onset and/or progression of diseases because the energy depletion of the neuronal cells directly leads to the dysfunction and degeneration of cells. In the case of prion diseases, mitochondrial involvement has been reported recently and evidence that prion protein (PrP) is localized in mitochondria is increasing. Despite these findings, the precise molecular mechanism by which PrP targets mitochondria remains unclear. PrP is a secretory protein and does not have a pre-sequence that targets the mitochondria, therefore, we thought that there was a covert signal in the amino acid sequence of PrP. To find the sequence, we constructed various GFP-fused PrP-truncations and colocalization with mitochondria was verified by live-cell imaging. Consequently, we found that 18 amino acids, PrP (122-139), are indispensable for the mitochondrial targeting of PrP. In addition, fluorescent microscopy observation revealed that PrP-localized mitochondria were accumulated at the perinuclear region in neuronal cells such as mouse neuroblastoma Neuro2a (N2a) and prion persistent infection N2a strain (ScN2a), anterograde movement of the mitochondria toward the cell membrane was completely inhibited because of the stacking of PrP on the outer membrane. The cristae formation of perinuclear accumulated mitochondria was disappeared indicating the reduced mitochondrial activity. Surprisingly, PrP-dependent mitochondrial perinuclear accumulation was specifically occurred on neuronal cells, whereas in epithelial HeLa cells and fibroblast COS-7 cells, no perinuclear accumulation observed even after the mitochondrial targeting of PrP.

Use of different RT-QuIC substrates for detecting CWD prions in the brain of Norwegian cervids.

Chronic wasting disease (CWD) is a highly contagious prion disease affecting captive and free-ranging cervid populations. CWD has been detected in United States, Canada, South Korea and, most recently, in Europe (Norway, Finland and Sweden). Animals with CWD release infectious prions in the environment through saliva, urine and feces sustaining disease spreading between cervids but also potentially to other non-cervids ruminants (e.g. sheep, goats and cattle). In the light of these considerations and due to CWD unknown zoonotic potential, it is of utmost importance to follow specific surveillance programs useful to minimize disease spreading and transmission. The European community has already in place specific surveillance measures, but the traditional diagnostic tests performed on nervous or lymphoid tissues lack sensitivity. We have optimized a Real-Time Quaking-Induced Conversion (RT-QuIC) assay for detecting CWD prions with high sensitivity and specificity to try to overcome this problem. In this work, we show that bank vole prion protein (PrP) is an excellent substrate for RT-QuIC reactions, enabling the detection of trace-amounts of CWD prions, regardless of prion strain and cervid species. Beside supporting the traditional diagnostic tests, this technology could be exploited for detecting prions in peripheral tissues from live animals, possibly even at preclinical stages of the disease.

The Predictive Value of Endpoint Quaking-Induced Conversion in Creutzfeldt-Jakob Disease.

Creutzfeldt-Jakob disease (CJD) is a fatal neurological illness for which accurate diagnosis is paramount. Real-time quaking-induced conversion (RT-QuIC) is a prion-specific assay with high sensitivity and specificity for CJD. The Canadian endpoint quaking-induced conversion (EP-QuIC) test is similar, but unlike RT-QuIC there is little data regarding its diagnostic utility in clinical practice. In this exploratory predictive value analysis of EP-QuIC in CJD, the negative predictive value (NPV) and positive predictive value (PPV) was 100% and 83%, respectively, with one false-positive result identified. Re-testing this sample with an optimized EP-QuIC protocol eliminated this false-positive result, leading to a PPV of 100%.

La valeur prédictive de la méthode diagnostique dite de conversion provoquée par tremblement au point final dans le cas de la maladie de Creutzfeldt-Jakob. La maladie de Creutzfeldt-Jakob (MCJ) est une maladie neurologique qui entraîne à terme un décès et pour laquelle l'établissement d'un diagnostic précis est primordial. La conversion provoquée par tremblement en temps réel (RT-QuIC en anglais) est une méthode diagnostique qui repose sur la détection de la protéine prion. Dans le cas de la MCJ, cette méthode est réputée posséder une sensibilité et une spécificité élevées. La méthode canadienne dite de conversion provoquée par tremblement au point final (EP-QuIC en anglais) se veut similaire mais, à la différence de la RT-QuIC, il existe peu de données en ce qui concerne son utilité diagnostique dans le cadre d'une pratique clinique. Dans cette analyse prédictive exploratoire de la EP-QuIC en lien avec la MCJ, la valeur prédictive négative (VPN) et la valeur prédictive positive (VPP) ont été respectivement de 100 % et de 83 %, un seul résultat faux-positif ayant été identifié. Le fait de soumettre notre échantillon à un nouveau test effectué à l'aide d'un protocole d'EP-QuIC optimisé a permis d'éliminer ce résultat faux-positif, ce qui a débouché sur une VPP de 100 %.

Psicosis como inicio de enfermedad de Creutzfeldt-Jakob esporádica / Psychosis as debut of sporadic Creutzfeldt-Jakob disease

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Human prion diseases.

PURPOSE OF REVIEW: Prion diseases are rapidly progressive neurodegenerative conditions that can be difficult to diagnose and are transmissible under specific circumstances. The authors will provide background regarding prion disease and focus on diagnostic tools. RECENT FINDINGS: Prion disease is caused by misfolded prion protein. The three possible causes of prion disease include sporadic (85%), genetic (10-15%), and acquired (<1%). Acquired prion diseases include kuru, iatrogenic, and variant Creutzfeldt-Jakob disease. Prion diseases differ in their clinical manifestation, neuropathology, and diagnostic test results. A variety of recent diagnostic tools have evolved that allow more reliable antemortem diagnosis of prion disease such as brain MRI and cerebrospinal fluid real-time quaking-induced conversion. Special infectivity guidelines must be followed when dealing with central nervous system tissue, but only standard precautions are needed for routine clinical care of patients with prion disease. SUMMARY: The only way to definitely diagnose prion disease and determine its type is via neuropathologic examination. However, brain MRI and cerebrospinal fluid real-time quaking-induced conversion have drastically increased diagnostic accuracy and are important tests to use when evaluating patients with suspected prion disease.

Accumulation of prion protein in the vagus nerve in creutzfeldt-jakob disease.

Disease-associated proteins are thought to propagate along neuronal processes in neurodegenerative diseases. To detect disease-associated prion protein (PrPSc ) in the vagus nerve in different forms and molecular subtypes of Creutzfeldt-Jakob disease (CJD), we applied 3 different anti-PrP antibodies. We screened the vagus nerve in 162 sporadic and 30 genetic CJD cases. Four of 31 VV-2 type sporadic CJD and 7 of 30 genetic CJD cases showed vagal PrPSc immunodeposits with distinct morphology. Thus, PrPSc in CJD affects the vagus nerve analogously to α-synuclein in Parkinson disease. The morphologically diverse deposition of PrPSc in genetic and sporadic CJD argues against uniform mechanisms of propagation of PrPSc . Ann Neurol 2019;85:782-787.

Intermolecular crosslinking of abnormal prion protein is efficiently induced by a primuline-sensitized photoreaction.

In prion diseases, infectious pathogenic particles that are composed of abnormal prion proteins (PrPSc) accumulate in the brain. PrPSc is biochemically characterized by its protease-resistance core (PrPres), but its structural features have not been fully elucidated. Here, we report that primuline, a fluorescent dye with photosensitization activity, dramatically enhances UV-irradiation-induced SDS-resistant PrPSc/res oligomer formation that can be detected by immunoblot analysis of prion-infected materials. This oligomer formation occurs specifically with PrPSc/res but not with normal prion protein, and it was demonstrated using purified PrPSc/res as well as unpurified materials. The oligomer formation proceeded in both primuline-dose- and UV irradiation time-dependent manners. Treatment with urea or formic acid did not break oligomers into monomers. Neither did the presence of aromatic amino acids modify oligomer formation. Analysis with a panel of anti-prion protein antibodies showed that the antibodies against the N-terminal region of PrPres were less reactive in the dimer than the monomer. These findings suggest that the primuline-sensitized photoreaction enhances intermolecular crosslinking of PrPSc/res molecules at a hydrophobic area of the N-terminal region of PrPres. In the screening of other compounds, photoreactive compounds such as luciferin exhibited a similar but lower activity with respect to oligomer formation than primuline. The enhanced photoreaction with these compounds will be useful for evaluating the structural features of PrPSc/res, especially the interactions between PrPSc/res molecules.

Melatonin protects mesenchymal stem cells from autophagy-mediated death under ischaemic ER-stress conditions by increasing prion protein expression.

OBJECT: The purpose of this study was to explore whether melatonin could protect mesenchymal stem cells (MSCs) against ischaemic injury, by inhibiting endoplasmic reticulum (ER) stress and autophagy both in vivo and in vitro. MATERIALS AND METHODS: To confirm the protective effect of melatonin against ER stress in MSCs, markers of cell viability, apoptosis and autophagy were analysed. To further investigate the regenerative effect of melatonin-treated MSCs in ischaemic tissues, a murine hindlimb ischaemic model was established. RESULTS: Under oxidative stress conditions, treatment with melatonin suppressed the activation of ER stress-associated proteins and autophagy-associated proteins acting through upregulation of cellular prion protein (PrPC ) expression. Consequently, inhibition of apoptotic cell death occurred. Melatonin also promoted the activation of MnSOD and catalase activities in MSCs. In a murine hindlimb ischaemia model, melatonin-treated MSCs also enhanced the functional limb recovery as well as neovascularization. These beneficial effects of melatonin were all blocked by knock-down of PrPC expression. CONCLUSION: Melatonin protects against ER stress/autophagy-induced apoptotic cell death by augmenting PrPC expression. Thus, melatonin-treated MSCs could be a potential cell-based therapeutic agent for ER stress-induced ischaemic diseases, and melatonin-induced PrPC might be a key molecule in ameliorating ER stress and autophagy.