Diagnostic accuracy of cerebrospinal fluid protein markers for sporadic Creutzfeldt-Jakob disease in Canada: a 6-year prospective study

Background  

To better characterize the value of cerebrospinal fluid (CSF) proteins as diagnostic markers in a clinical population of subacute encephalopathy patients with relatively low prevalence of sporadic Creutzfeldt-Jakob disease (sCJD), we studied the diagnostic accuracies of several such markers (14-3-3, tau and S100B) in 1000 prospectively and sequentially recruited Canadian patients with clinically suspected sCJD.     

Magnetic resonance spectroscopy and measurement of tau epitopes of autopsy proven sporadic Creutzfeldt-Jakob disease in a patient with non-specific initial EEG, MRI and negative 14-3-3 immunoblot

Limited potential of electroencephalogram (EEG), magnetic resonance images (MRI) and cerebrospinal fluid (CSF) test for 14-3-3 protein in the clinical diagnosis of sporadic Creutzfeldt-Jakob disease (sCJD) resulted in developments in diagnostic premortem tehniques. Recent studies provided evidence that magnetic resonance spectroscopy (MRS) and measurement of total-tau (T-tau) and phospho-tau (P-tau) may be useful to identify patients with CJD. We combined detected metabolic changes in the brain by MRS and measured T-tau and tau-pT181 by ELISA, and tau-pT231 by Westernblot in a patient with autopsy proven sCJD. Our results show that in contrast to negative CSF 14-3-3 protein, nonspecific EEG and MRI, MRS revealed metabolic alterations in regions of the brain that has appeared normal on MRI, and tau tests has shown measurable levels of phosphorylated and non-phosphorylated isoforms in CSF. We conclude that rapidly progressive dementia with negative 14-3-3 test and non-specific initial EEG and MRI must still be considered in the differential diagnosis of the sCJD. Combination of serial functional MRI along with MRS study and measurement of tau ratio could improve the early diagnosis of sCJD. The current case is the first attempt to study results of the use of MRS and tau tests in a case of sCJD with diagnostic dilemma.     

Chinese Specific Characteristics of Sporadic Creutzfeldt-Jakob Disease: A Retrospective Analysis of 57 Cases

Objective

Sporadic Creutzfeldt-Jakob disease (sCJD) is a fatal and transmissible neurodegenerative disorder. However, no studies have reported Chinese specific characteristics of sCJD. We aimed to identify differences in sCJD between Chinese patients and patients from other countries.

Methods

The data from 57 Chinese sCJD patients were retrospectively analyzed, including demographic data, clinical manifestations, laboratory examinations, electroencephalograms (EEGs), diffusion-weighted imaging (DWI) scans, positron emission tomography (PET) scans, and pathological results.

Result

The disease was pathologically confirmed in 11 patients. 39 cases were diagnosed as probable sCJD, and 7 were possible. Of the total cases, 33 were male, and 24 were female. The onset age ranged from 36 to 75 years (mean: 55.5, median: 57). Disease onset before the age of 60 occurred in 57.9% of patients. The disease duration from onset to death ranged 5–22 months (mean: 11.6, median: 11), and 51.9% of patients died 7 to 12 months after disease onset. The majority of patients presented with sub-acute onset with progressive dementia. 3 of the 9 patients who took 14-3-3 protein analysis had positive results (33.3%). The sensitivity of EEG was 79.6% (43/54). For DWI and PET examinations, the sensitivities were 94% (47/50) and 94.1% (16/17), respectively. In seven patients who did not show typical hyper-intensities on the first DWI examination, abnormalities of hypo-metabolism in the cerebral cortex were clearly detected by PET. In 13 out of the 17 patients, PET detected extra abnormal regions in addition to the hyper-intense areas observed in DWI.

Conclusion

This is the first study to indicate that Chinese sCJD patients have a much earlier onset age and a longer disease duration than other populations, which is most likely related to racial differences. The longer disease duration may also be a probable characteristic of Asian populations. PET had high sensitivity for the diagnosis of sCJD.     

The epidemiological, clinical, and laboratory features of sporadic Creutzfeldt-Jakob disease patients in China: surveillance data from 2006 to 2010

Background

Creutzfeldt-Jakob disease (CJD) is a rare, rapidly progressive fatal central nervous system disorder, which consists of three main catalogues: sporadic, familial, and iatrogenic CJD.

Methodology/Principal Findings

In China, the surveillance for CJD started in 2006, covering 12 provincial Centers for Disease Control and Prevention (CDCs) and 15 hospitals. From 2006 to 2010, 624 suspected patients were referred to China CJD surveillance. The epidemiological, clinical and laboratory features of sporadic CJD (sCJD) were analysed. Both groups of probable and possible sCJD showed highest incidences in the population of 60 to 69 year-olds. The most common presenting symptoms were progressive dementia and mental-related symptoms (neurological symptoms including sleeping turbulence, depression, anxiety and stress). Among the four main clinical manifestations, myoclonus was more frequently observed in the probable sCJD patients. About 2/3 of probable sCJD cases showed positive 14-3-3 in CSF and/or periodic sharp wave complexes (PSWC) in electroencephalography (EEG). The presence of myoclonus was significantly closely related with the appearance of PSWC in EEG. Polymorphisms of codon 129 in PRNP of the notified cases revealed a highly predominant M129M genotype in Han Chinese. Among 23 genetic human prion diseases, ten were D178N/M129M Fatal familial insomnia (FFI) and five were T188K genetic CJD (gCJD), possibly indicating a special distribution of gCJD-related mutations in Han Chinese.

Conclusion

From the period of 2006 to 2010, 261 patients were diagnosed as sCJD and 23 patients were diagnosed as genetic human prion diseases in China. The epidemiological, clinical and laboratory analysis data were consistent with the characteristics of sporadic CJD, which provide insight into the features of CJD in China.     

High sensitivity of an ELISA kit for detection of the gamma-isoform of 14-3-3 proteins: usefulness in laboratory diagnosis of human prion disease

Background  

The gamma-isoform of the 14-3-3 protein (14-3-3 gamma) is expressed in neurons, and could be a specific marker for neuronal damage. This protein has been reported as a detectable biomarker, especially in the cerebrospinal fluid (CSF) of Creutzfeldt-Jakob disease (CJD) patients by Western blotting (WB) or enzyme-linked immunosorbent assays (ELISAs). Western blotting for 14-3-3 gamma is not sensitive, and the reported data are conflicting among publications. An ELISA specific for 14-3-3 gamma is not available.     

MRI findings are often missed in the diagnosis of Creutzfeldt-Jakob disease

Background

Establishing a confident clinical diagnosis before an advanced stage of illness can be difficult in Creutzfeldt-Jakob disease (CJD) but unlike common causes of dementia, prion diseases can often be diagnosed by identifying characteristic MRI signal changes. However, it is not known how often CJD-associated MRI changes are identified at the initial imaging report, whether the most sensitive sequences are used, and what impact MRI-diagnosis has on prompt referral to clinical trial-like studies.

Methods

We reviewed the MRI scans of 103 patients with CJD referred to the National Prion Clinic since 2007 and reviewed the presence of CJD-associated changes, compared these findings with the formal report from the referring centre and reviewed the types of sequence performed.

Results

In sCJD we found CJD-associated MRI changes in 83 of 91 cases (91% sensitivity). However, the referring centres documented CJD-associated MRI changes in 43 of the sCJD cases (47% sensitivity). The most common region not documented by referring centres was the cortex (23 of 68 sCJD cases), but there was a statistically significant discrepancy in all regions (p<0.0001). Patients in whom MRI abnormalities were missed by the referring hospital were more advanced at the time of recruitment to a clinical trial-like study (p=0.03).

Conclusions

CJD-associated MRI changes are often not documented on the formal investigation report at the referring centre. This is important as delay makes enrolment to clinical trials futile because of highly advanced disease. If a diagnosis of CJD is suspected, even if the initial imaging is reported as normal, a specialist MRI review either by an experienced neuroradiologist or by a prion disease specialist unit could facilitate earlier diagnosis.

     

Deletion of the cruciform binding domain in CBP/14-3-3 displays reduced origin binding and initiation of DNA replication in budding yeast

Background  

Initiation of eukaryotic DNA replication involves many protein-protein and protein-DNA interactions. We have previously shown that 14-3-3 proteins bind cruciform DNA and associate with mammalian and yeast replication origins in a cell cycle dependent manner.     

Wernicke-Korsakoff syndrome as a rare phenotype of sporadic Creutzfeldt-Jakob disease

We reported the case of a patient with Wernicke-Korsakoff syndrome (WKs) as an early clinical manifestation of sporadic Creutzfeld-Jakob disease (sCJD). The 66-year-old female complained of dizziness and imbalance which mostly occurred while walking. A neurological examination revealed a triad of symptoms characteristic for WKs such as gaze paresis, ataxia of limbs and trunk as well as memory disturbances with confabulations. The disturbances increased during the course of the disease, which led to the death of the patient four months after the appearance of the signs. The patient was finally diagnosed with sCJD disease. The most useful ancillary examination results supporting sCJD diagnosis were brain diffusion DWI MRI (diffusion weighted magnetic resonance imaging) and the presence of 14–3-3 protein in CSF (cerebrospinal fluid). Since that manifestation of sCJD is very unique other causes should be taken into consideration while making a final diagnosis.     

14-3-3 theta binding to cell cycle regulatory factors is enhanced by HIV-1 Vpr

Background  

Despite continuing advances in our understanding of AIDS pathogenesis, the mechanism of CD4+ T cell depletion in HIV-1-infected individuals remains unclear. The HIV-1 Vpr accessory protein causes cell death, likely through a mechanism related to its ability to arrest cells in the G₂,M phase. Recent evidence implicated the scaffold protein, 14-3-3, in Vpr cell cycle blockade.     

Regulating Factors of PrPres Glycosylation in Creutzfeldt-Jakob Disease - Implications for the Dissemination and the Diagnosis of Human Prion Strains

Objective

The glycoprofile of pathological prion protein (PrP^res) is widely used as a diagnosis marker in Creutzfeldt-Jakob disease (CJD) and is thought to vary in a strain-specific manner. However, that the same glycoprofile of PrP^res always accumulates in the whole brain of one individual has been questioned. We aimed to determine whether and how PrP^res glycosylation is regulated in the brain of patients with sporadic and variant Creutzfeldt-Jakob disease.

Methods

PrP^res glycoprofiles in four brain regions from 134 patients with sporadic or variant CJD were analyzed as a function of the genotype at codon 129 of PRNP and the Western blot type of PrP^res.

Results

The regional distribution of PrP^res glycoforms within one individual was heterogeneous in sporadic but not in variant CJD. PrP^res glycoforms ratio significantly correlated with the genotype at codon 129 of the prion protein gene and the Western blot type of PrP^res in a region-specific manner. In some cases of sCJD, the glycoprofile of thalamic PrP^res was undistinguishable from that observed in variant CJD.

Interpretation

Regulations leading to variations of PrP^res pattern between brain regions in sCJD patients, involving host genotype and Western blot type of PrP^res may contribute to the specific brain targeting of prion strains and have direct implications for the diagnosis of the different forms of CJD.     

Towards an Age-Dependent Transmission Model of Acquired and Sporadic Creutzfeldt-Jakob Disease

Introduction

Sporadic Creutzfeldt-Jakob disease (sCJD) might be transmitted by surgery. The purpose of this study was to investigate potential susceptibility to sCJD from surgery at juvenile age and in early adulthood.

Methods

From Danish and Swedish national registries we identified 167 definite and probable sCJD cases with onset from 1987 through 2003, and 835 age-, sex- and residence-matched controls along with their surgical histories. Main, anatomically or etiologically classified surgical procedures followed by a ≥20-year lag were analyzed using logistic regression, and stratified by age at first-registered surgical discharge.

Results

The risk of having a diagnosis of CJD depended strongly on age at first surgery with odds ratio (OR) of 12.80 (95% CI 2.56–64.0) in patients <30 years, 3.04 (95% 1.26–7.33) in 30–39 years, and 1.75 (95% CI 0.89–3.45) in ≥40 years, for anatomically classified surgical procedures. Similar figures were obtained for etiologically classified surgical procedures.

Conclusions

Risk of surgical-acquired sCJD depends on age at exposure; this pattern is similar to age-specific profiles reported for CJD accidentally transmitted by human pituitary-derived growth hormone and susceptibility curves for variant CJD estimated after adjustment for dietary exposure to bovine spongiform encephalopathy. There might be an age-at-exposure-related susceptibility to acquire all CJD forms, including sCJD from routine surgery.     

Structural basis of the 14-3-3 protein-dependent activation of yeast neutral trehalase Nth1

Background

Trehalases are highly conserved enzymes catalyzing the hydrolysis of trehalose in a wide range of organisms. The activity of yeast neutral trehalase Nth1 is regulated in a 14-3-3- and a calcium-dependent manner. The Bmh proteins (the yeast 14-3-3 isoforms) recognize phosphorylated Nth1 and enhance its enzymatic activity through an unknown mechanism.

Methods

To investigate the structural basis of interaction between Nth1 and Bmh1, we used hydrogen/deuterium exchange coupled to mass spectrometry, circular dichroism spectroscopy and homology modeling to identify structural changes occurring upon the complex formation.

Results

Our results show that the Bmh1 protein binding affects structural properties of several regions of phosphorylated Nth1: the N-terminal segment containing phosphorylation sites responsible for Nth1 binding to Bmh, the region containing the calcium binding domain, and segments surrounding the active site of the catalytic trehalase domain. The complex formation between Bmh1 and phosphorylated Nth1, however, is not accompanied by the change in the secondary structure composition but rather the change in the tertiary structure.

Conclusions

The 14-3-3 protein-dependent activation of Nth1 is based on the structural change of both the calcium binding domain and the catalytic trehalase domain. These changes likely increase the accessibility of the active site, thus resulting in Nth1 activation.

General significance

The results presented here provide a structural view of the 14-3-3 protein-dependent activation of yeast neutral trehalase Nth1, which might be relevant to understand the process of Nth1 activity regulation as well as the role of the 14-3-3 proteins in the regulation of other enzymes.     

14-3-3 protein directly interacts with the kinase domain of calcium/calmodulin-dependent protein kinase kinase (CaMKK2)

Background

Calcium/calmodulin-dependent protein kinase kinase 2 (CaMKK2) is a member of the Ca²⁺/calmodulin-dependent kinase (CaMK) family involved in adiposity regulation, glucose homeostasis and cancer. This upstream activator of CaMKI, CaMKIV and AMP-activated protein kinase is inhibited by phosphorylation, which also triggers an association with the scaffolding protein 14-3-3. However, the role of 14-3-3 in the regulation of CaMKK2 remains unknown.

Clinical and laboratory features of 14 young Chinese probable sCJD patients

Sporadic Creutzfeldt-Jakob disease (sCJD) occurs frequently in the relatively older population, mainly in the groups of 60–69 and 70–79 year-old. Since 2006 when China performed national CJD surveillance, 14 young probable sCJD patients below 40 year-old were identified, counting for 1.93% of all probable sCJD cases. The clinical features of young probable sCJD cases, including the onset feature, the presence of sCJD-associated signs and the clinical duration, are indistinguishable from those of older patients. Special sCJD-associated abnormalities on EEG and MRI were noticed in 7 and 10 cases. CSF 14–3–3 was positive in 7 cases. CSF RT-QuIC showed positive reactive curves in 9 cases, with short lag phases. PRNP sequencing did not find any mutation. Due to low rate of brain autopsy in China, performances of other CJD-associated examinations as much as possible are extremely important for the distinguish diagnosis of young probable sCJD patients.     

A polymorphism in the YWHAH gene encoding 14-3-3 eta that is not associated with sporadic Creutzfeldt–Jakob disease (CJD)

14-3-3 proteins are abundantly expressed in the brain, particularly neuronal tissue and are thought to serve multiple biological functions involved in neuronal development and cell growth and death. Recent studies have shown associations of 14-3-3 genes with neurodegenerative disorders based on their chromosomal linkage to these diseases and to regulatory functions for the nervous system. Although the role of 14-3-3 proteins in the pathogenesis of prion diseases remains unknown, the detection of altered levels of isoforms of the 14-3-3 protein in the cerebrospinal fluid is considered a biomarker for diagnosis of sporadic Creutzfeldt–Jakob disease (sCJD). To identify other susceptibility genes for prion disease, we examined nucleotide variations in YWHAH, a gene encoding 14-3-3 eta. This case–control study included 182 sCJD patients and 206 healthy Koreans. Polymerase chain reaction was used to amplify open reading frame and some 3′-untranslated region (UTR) in exon 2, and direct sequencing was carried out. One polymorphism, 753 G/A, was detected in the 3′-UTR of exon 2 on the YWHAH. The genotype distribution and allele frequencies of the YWHAH 753 G/A polymorphism were not significantly different between controls and sCJD patients. This finding indicates that YWHAH 753 G/A polymorphism is unlikely to be linked to genetic susceptibility or have a modifying effect in sCJD. On analysis stratified by the prion protein gene 129 or 219 genotype, no significant relation was found in genotype and allele frequencies of the YWHAH 753G/A. This is the first genetic association study of YWHAH with sCJD populations.     

Saliva levels of Abeta1-42 as potential biomarker of Alzheimer's disease: a pilot study

Background  

Simple, non-invasive tests for early detection of degenerative dementia by use of biomarkers are urgently required. However, up to the present, no validated extracerebral diagnostic markers for the early diagnosis of Alzheimer disease (AD) are available. The clinical diagnosis of probable AD is made with around 90% accuracy using modern clinical, neuropsychological and imaging methods. A biochemical marker that would support the clinical diagnosis and distinguish AD from other causes of dementia would therefore be of great value as a screening test. A total of 126 samples were obtained from subjects with AD, and age-sex-matched controls. Additionally, 51 Parkinson's disease (PD) patients were used as an example of another neurodegenerative disorder. We analyzed saliva and plasma levels of β amyloid (Aβ) using a highly sensitive ELISA kit.     

Colocalization of 14-3-3 proteins with SOD1 in Lewy body-like hyaline inclusions in familial amyotrophic lateral sclerosis cases and the animal model

Background and Purpose

Cu/Zn superoxide dismutase (SOD1) is a major component of Lewy body-like hyaline inclusion (LBHI) found in the postmortem tissue of SOD1-linked familial amyotrophic lateral sclerosis (FALS) patients. In our recent studies, 14-3-3 proteins have been found in the ubiquitinated inclusions inside the anterior horn cells of spinal cords with sporadic amyotrophic lateral sclerosis (ALS). To further investigate the role of 14-3-3 proteins in ALS, we performed immunohistochemical analysis of 14-3-3 proteins and compared their distributions with those of SOD1 in FALS patients and SOD1-overexpressing mice.

Methods

We examined the postmortem brains and the spinal cords of three FALS cases (A4V SOD1 mutant). Transgenic mice expressing the G93A mutant human SOD1 (mutant SOD1-Tg mice), transgenic mice expressing the wild-type human SOD1 (wild-type SOD1-Tg mice), and non-Tg wild-type mice were also subjected to the immunohistochemical analysis.

Results

In all the FALS patients, LBHIs were observed in the cytoplasm of the anterior horn cells, and these inclusions were immunopositive intensely for pan 14-3-3, 14-3-3β, and 14-3-3γ. In the mutant SOD1-Tg mice, a high degree of immunoreactivity for misfolded SOD1 (C4F6) was observed in the cytoplasm, with an even greater degree of immunoreactivity present in the cytoplasmic aggregates of the anterior horn cells in the lumbar spinal cord. Furthermore, we have found increased 14-3-3β and 14-3-3γ immunoreactivities in the mutant SOD1-Tg mice. Double immunofluorescent staining showed that C4F6 and 14-3-3 proteins were partially co-localized in the spinal cord with FALS and the mutant SOD1-Tg mice. In comparison, the wild-type SOD1-Tg and non-Tg wild-type mice showed no or faint immunoreactivity for C4F6 and 14-3-3 proteins (pan 14-3-3, 14-3-3β, and 14-3-3γ) in any neuronal compartments.

Discussion

These results suggest that 14-3-3 proteins may be associated with the formation of SOD1-containing inclusions, in FALS patients and the mutant SOD1-Tg mice.     

Phosphorylation-dependent protein interaction with Trypanosoma brucei 14-3-3 proteins that display atypical target recognition

Background

The 14-3-3 proteins are structurally conserved throughout eukaryotes and participate in protein kinase signaling. All 14-3-3 proteins are known to bind to evolutionally conserved phosphoserine-containing motifs (modes 1 and/or 2) with high affinity. In Trypanosoma brucei, 14-3-3I and II play pivotal roles in motility, cytokinesis and the cell cycle. However, none of the T. brucei 14-3-3 binding proteins have previously been documented.

Methodology/Principal Findings

Initially we showed that T. brucei 14-3-3 proteins exhibit far lower affinity to those peptides containing RSxpSxP (mode 1) and RxY/FxpSxP (mode 2) (where x is any amino acid residue and pS is phosphoserine) than human 14-3-3 proteins, demonstrating the atypical target recognition by T. brucei 14-3-3 proteins. We found that the putative T. brucei protein phosphatase 2C (PP2c) binds to T. brucei 14-3-3 proteins utilizing its mode 3 motif (–pS/pTx_1-2-COOH, where x is not Pro). We constructed eight chimeric PP2c proteins replacing its authentic mode 3 motif with potential mode 3 sequences found in Trypanosoma brucei genome database, and tested their binding. As a result, T. brucei 14-3-3 proteins interacted with three out of eight chimeric proteins including two with high affinity. Importantly, T. brucei 14-3-3 proteins co-immunoprecipitated with an uncharacterized full-length protein containing identified high-affinity mode 3 motif, suggesting that both proteins form a complex in vivo. In addition, a synthetic peptide derived from this mode 3 motif binds to T. brucei 14-3-3 proteins with high affinity.

Conclusion/Significance

Because of the atypical target recognition of T. brucei 14-3-3 proteins, no 14-3-3-binding proteins have been successfully identified in T. brucei until now whereas over 200 human 14-3-3-binding proteins have been identified. This report describes the first discovery of the T. brucei 14-3-3-binding proteins and their binding motifs. The high-affinity phosphopeptide will be a powerful tool to identify novel T. brucei 14-3-3-binding proteins.     

Structure of the p53 C-terminus bound to 14-3-3: Implications for stabilization of the p53 tetramer

The adaptor protein 14-3-3 binds to and stabilizes the tumor suppressor p53 and enhances its anti-tumour activity. In the regulatory C-terminal domain of p53 several 14-3-3 binding motifs have been identified. Here, we report the crystal structure of the extreme C-terminus (residues 385-393, p53pT387) of p53 in complex with 14-3-3σ at a resolution of 1.28 Å. p53pT387 is accommodated by 14-3-3 in a yet unrecognized fashion implying a rationale for 14-3-3 binding to the active p53 tetramer. The structure exhibits a potential binding site for small molecules that could stabilize the p53/14-3-3 protein complex suggesting the possibility for therapeutic intervention.

Structured summary

MINT-7711943: 14-3-3 sigma (uniprotkb:P31947) and p53 (uniprotkb:P04637) bind (MI:0407) by X-ray crystallography (MI:0114)MINT-7711931: 14-3-3 sigma (uniprotkb:P31947) and p53 (uniprotkb:P04637) bind (MI:0407) by isothermal titration calorimetry (MI:0065)