Isabelle Quadrio

Genetic Creutzfeldt-Jakob disease associated with the E200K mutation: characterization of a complex proteinopathy

The E200K mutation is the most frequent prion protein gene (PRNP) mutation detected worldwide that is associated with Creutzfeldt-Jakob disease (CJD) and thought to have overlapping features with sporadic CJD, yet detailed neuropathological studies have not been reported. In addition to the prion protein, deposition of tau, α-synuclein, and amyloid-β has been reported in human prion disease. To describe the salient and concomitant neuropathological alterations, we performed a systematic clinical, neuropathological, and biochemical study of 39 individuals carrying the E200K PRNP mutation originating from different European countries. The most frequent clinical symptoms were dementia and ataxia followed by myoclonus and various combinations of further symptoms, including vertical gaze palsy and polyneuropathy. Neuropathological examination revealed relatively uniform anatomical pattern of tissue lesioning, predominating in the basal ganglia and thalamus, and also substantia nigra, while the deposition of disease-associated PrP was more influenced by the codon 129 constellation, including different or mixed types of PrPres detected by immunoblotting. Unique and prominent intraneuronal PrP deposition involving brainstem nuclei was also noted. Systematic examination of protein depositions revealed parenchymal amyloid-β in 53.8%, amyloid angiopathy (Aβ) in 23.1%, phospho-tau immunoreactive neuritic profiles in 92.3%, neurofibrillary degeneration in 38.4%, new types of tau pathology in 33.3%, and Lewy-type α-synuclein pathology in 15.4%. TDP-43 and FUS immunoreactive protein deposits were not observed. This is the first demonstration of intensified and combined neurodegeneration in a genetic prion disease due to a single point mutation, which might become an important model to decipher the molecular interplay between neurodegeneration-associated proteins.

Risk of Alzheimer's disease biological misdiagnosis linked to cerebrospinal collection tubes.

Tau proteins and amyloid-β (Aβ) peptides are the current recognized cerebrospinal fluid (CSF) biomarkers used as an aid in the diagnosis of Alzheimers disease (AD). However, there is no consensus on their clinical use due to non-qualified cut-off values, probably related to the observed high pre-analytical and analytical variability. Standardized pre-analytical protocols have therefore been proposed. Importantly, these recommend the use of polypropylene collection/sampling tubes while, to date, no broad comparison of these types of tubes has been conducted. In this study, we first compared, as part of a real clinical workflow, the impact of four different collection tubes on the CSF concentration of Aβ peptides (Aβ42, Aβ40) and total (hTau) and phosphorylated (P-Tau181P) tau proteins measured using routine ELISA kits. We then extended this study to 11 polypropylene tubes used by different clinical laboratories, and investigated their plastic polymer composition using differential scanning calorimetry and Fourier Transformed Infrared spectroscopy. Significant concentration variations linked solely to the use of different types of tubes were observed. This was particularly marked for Aβ peptides, with >50% disparity occurring in less than five minutes. Polymer composition analysis revealed that most polypropylene tubes were in fact copolymers with at least polyethylene. There was no clear correlation between tube composition and pre-analytical behavior. Our results show that the use of polypropylene tubes does not guarantee satisfactory pre-analytical behavior. They also point to collection/sampling tubes being a major pre-analytical source of variability that could impact the significance of AD biological diagnosis.

Impact of chronic Helicobacter pylori infection on Alzheimer's disease: preliminary results

Recent case-control studies reported an association between H. pylori infection and Alzheimers disease (AD). Our aim was to compare cognitive impairment, neuroinflammation, and cerebrovascular lesion load in a group of AD patients according to their H. pylori status. For the 53 AD patients included, we assessed: clinical data (vascular comorbidities and cognitive assessment), biological data (especially fibrinogen, homocysteine levels, apolipoprotein E4 genotype; cerebrospinal fluid [CSF] total tau protein [Tau], phospho-tau(181) protein [pTau(181)]), and amyloid beta peptide levels, serum/CSF-cytokines (interleukin [IL]-1β, IL-6, IL-8, tumor necrosis factor [TNF]-α) and pepsinogen I/pepsinogen II (PgI/PgII) ratio, and cerebrovascular lesion load (magnetic resonance imaging [MRI] fluid-attenuated inversion recovery [FLAIR] with the Fazekas and Schmidt scale). H. pylori infection was diagnosed by enzyme-linked immunosorbent assay (ELISA) and immunoblot test. H. pylori infection was associated with a decreased Mini Mental State Examination (MMS) (p = 0.024), and higher CSF pTau(181) (p = 0.014) and tau (p = 0.021) levels. A decreased PgI/II ratio (i.e., an increased gastric atrophy) was associated with the infection (p = 0.005). Homocysteine levels were positively correlated to Fazekas score (r = 0.34; p = 0.032) and to H. pylori immunoglobulin (Ig)G levels (r = 0.44; p = 0.001). Higher CSF cytokine levels (IL-8, p = 0.003; TNF-α, p = 0.019) were associated with the infection, but systemic inflammation results were controversial. Finally, in multivariate analysis, a lower MMSE score (odds ratio [OR], 0.83 [0.72-0.97]; p = 0.017), plasma IL-1β level (OR, 0.31 [0.11-0.87]; p = 0.025), an increased gastric atrophy, i.e., a lower PgI/PgII ratio (OR, 0.63 [0.43-0.93]; p = 0.020) were still associated with the infection. AD patients infected by H. pylori tended to be more cognitively impaired. Studies are needed to attest to the impact of H. pylori infection on AD course, especially on cerebrovascular lesions and neuroinflammation.

Impact of harmonization of collection tubes on Alzheimer's disease diagnosis

The objective of this study was to analyze differences in biomarker outcomes before and after harmonization of cerebrospinal fluid (CSF) collection tubes in Alzheimers disease (AD) diagnosis.

Cerebrospinal Fluid Collection Tubes: A Critical Issue for Alzheimer Disease Diagnosis

To the Editor: Total tau protein (hTau),1 its phosphorylated isoform (p-Tau181P), and Aβ1–42 peptides are the currently accepted cerebrospinal fluid (CSF) biomarkers used as aids in the diagnosis of Alzheimer disease (1). Although polypropylene (PP) was previously reported as the best material for CSF collection tubes (2), heterogeneity in CSF Aβ1–42 values was observed with different PP sampling tubes (3). Because the recommendation to use PP tubes did not lead to standardization of clinical cutoff values (4), we decided to fully address this issue by comparing various types of tubes within an actual clinical work flow and by analyzing the material of different commercially available PP tubes. In the framework of an ethically approved study, we collected CSF samples from 12 patients directly (from the lumbar puncture needle) into 2 PP tubes [BD catalog no. 352096 (BD-PP); Sarstedt catalog no. 62.610.201 (ST-PP)], 1 hemolysis polyethylene tube [Fisher Scientific catalog no. ref.W1773X (HE-PE)], and 1 polystyrene tube [BD catalog no. 352095 (BD-PS)]. CSF biomarker concentrations were measured in parallel in these 4 types of tubes with Innogenetics INNOTEST® kits. We extended this analysis by comparing the results obtained with 11 different commercially available collection tubes labeled as “PP” for a series of fresh (unfrozen) …

Association of Cerebrospinal Fluid Prion Protein Levels and the Distinction Between Alzheimer Disease and Creutzfeldt-Jakob Disease

IMPORTANCE Although typical forms of Alzheimer disease (AD) and Creutzfeldt-Jakob disease (CJD) are clinically distinguishable, atypical AD phenotypes may pose a diagnostic challenge. The major biological diagnostic biomarker for identifying CJD, 14-3-3 protein in cerebrospinal fluid (CSF), unfortunately lacks specificity when confronting a rapid dementia presentation. OBJECTIVE To assess the relevance of total CSF prion protein (t-PrP) levels in the differential biological diagnosis between atypical AD phenotypes and CJD. DESIGN, SETTING, AND PARTICIPANTS A retrospective study in an autopsy-confirmed cohort of 82 patients was performed to evaluate the relevance of CSF t-PrP to distinguish 30 definite cases of AD from 52 definite cases of CJD. Next, CSF t-PrP concentration was measured in a cohort of 104 patients including 55 patients with probable AD, 26 with probable sporadic CJD, and 23 control patients for whom 14-3-3 protein, total tau, phosphorylated tau 181 (P-tau181), and Aβ1-42 were available. We investigated 46 patients diagnosed as having probable AD who presented atypical phenotypes. A diagnosis strategy was proposed to classify atypical AD phenotypes with suspicion of CJD based on a decision tree combining CSF biomarkers. MAIN OUTCOMES AND MEASURES We determined CSF t-PrP levels for all patients. We calculated the ratio of total tau and P-tau181 and determined the diagnostic accuracy of each biomarker alone or in combination. We calculated the misclassification rate for each biomarker that corresponded to the percentage of patients within the group of atypical AD phenotypes wrongly classified as CJD. RESULTS In patients with CJD, CSF t-PrP concentrations were decreased compared with control participants and patients with AD. When considering the differential diagnosis of CJD compared with atypical AD phenotypes, CSF t-PrP determination reached 82.1% sensitivity and 91.3% specificity. The misclassification rate of atypical AD phenotypes decreased from 43.5%, obtained when using the CSF 14-3-3 protein determination alone, to only 4.3% when calculating the ratio total tau/(P-tau181 × t-PrP). The proposed classification tree permitted correct classification of 98.4% of the patients. CONCLUSIONS AND RELEVANCE For unusual phenotypes of AD, especially cases presenting with a biological ambiguity suggesting CJD, determination of CSF t-PrP levels increased diagnostic accuracy. The use of CSF t-PrP levels may be beneficial in clinical practice in addition to the current classic biomarkers.

Pre-analytical and analytical factors influencing Alzheimer's disease cerebrospinal fluid biomarker variability

A panel of cerebrospinal fluid (CSF) biomarkers including total Tau (t-Tau), phosphorylated Tau protein at residue 181 (p-Tau) and β-amyloid peptides (Aβ42 and Aβ40), is frequently used as an aid in Alzheimers disease (AD) diagnosis for young patients with cognitive impairment, for predicting prodromal AD in mild cognitive impairment (MCI) subjects, for AD discrimination in atypical clinical phenotypes and for inclusion/exclusion and stratification of patients in clinical trials. Due to variability in absolute levels between laboratories, there is no consensus on medical cut-off value for the CSF AD signature. Thus, for full implementation of this core AD biomarker panel in clinical routine, this issue has to be solved. Variability can be explained both by pre-analytical and analytical factors. For example, the plastic tubes used for CSF collection and storage, the lack of reference material and the variability of the analytical protocols were identified as important sources of variability. The aim of this review is to highlight these pre-analytical and analytical factors and describe efforts done to counteract them in order to establish cut-off values for core CSF AD biomarkers. This review will give the current state of recommendations.

CSF neopterin level as a diagnostic marker in primary central nervous system lymphoma

BACKGROUND The diagnosis of primary central nervous system lymphoma (PCNSL) can be challenging. PCNSL lesions are frequently located deep within the brain, and performing a cerebral biopsy is not always feasible. The aim of this study was to investigate the diagnostic value of CSF neopterin, a marker of neuroinflammation, in immunocompetent patients with suspected PCNSL. METHODS We retrospectively reviewed the characteristics of 124 patients with brain tumor (n = 82) or an inflammatory CNS disorder (n = 42) in whom CSF neopterin levels were assessed. Twenty-eight patients had PCNSL, 54 patients had another type of brain tumor (glioma n = 36, metastasis n = 13, other n = 5), and 13 patients had a pseudotumoral inflammatory brain lesion. RESULTS CSF neopterin levels were significantly higher in the patients with PCNSL than in those with other brain tumors (41.8 vs 5.1 nmol/L, P < .001), those with pseudotumoral inflammatory brain lesions (41.8 vs 4.3 nmol/L, P < .001), and those with nontumefactive inflammatory CNS disorders (41.8 vs 3.8 nmol/L, P < .001). In the 95 patients with space-occupying brain lesions, at a cutoff of 10 nmol/L, the sensitivity of this approach was 96% and the specificity was 93% for the diagnosis of PCNSL. The positive and negative predictive values were 84% and 98%, respectively. CONCLUSION Assessing CSF neopterin levels in patients with a suspected brain tumor might be helpful for the positive and differential diagnosis of PCNSL. A prospective study is warranted to confirm these results.

Chasing the Effects of Pre-Analytical Confounders – A Multicenter Study on CSF-AD Biomarkers

Introduction Core cerebrospinal fluid (CSF) biomarkers – Aβ42, Tau, and phosphorylated Tau (pTau) – have been recently incorporated in the revised criteria for Alzheimer’s disease (AD). However, their widespread clinical application lacks standardization. Pre-analytical sample handling and storage play an important role in the reliable measurement of these biomarkers across laboratories. Aim In this study, we aim to surpass the efforts from previous studies, by employing a multicenter approach to assess the impact of less studied CSF pre-analytical confounders in AD-biomarkers quantification. Methods Four different centers participated in this study and followed the same established protocol. CSF samples were analyzed for three biomarkers (Aβ42, Tau, and pTau) and tested for different spinning conditions [temperature: room temperature (RT) vs. 4°C; speed: 500 vs. 2000 vs. 3000 g], storage volume variations (25, 50, and 75% of tube total volume), as well as freezing-thaw cycles (up to five cycles). The influence of sample routine parameters, inter-center variability, and relative value of each biomarker (reported as normal/abnormal) was analyzed. Results Centrifugation conditions did not influence biomarkers levels, except for samples with a high CSF total protein content, where either non-centrifugation or centrifugation at RT, compared to 4°C, led to higher Aβ42 levels. Reducing CSF storage volume from 75 to 50% of total tube capacity decreased Aβ42 concentration (within analytical CV of the assay), whereas no change in Tau or pTau was observed. Moreover, the concentration of Tau and pTau appears to be stable up to five freeze–thaw cycles, whereas Aβ42 levels decrease if CSF is freeze-thawed more than three times. Conclusion This systematic study reinforces the need for CSF centrifugation at 4°C prior to storage and highlights the influence of storage conditions in Aβ42 levels. This study contributes to the establishment of harmonized standard operating procedures that will help reducing inter-lab variability of CSF-AD biomarkers evaluation.

Oral Transmission of L-type Bovine Spongiform Encephalopathy in Primate Model

We report transmission of atypical L-type bovine spongiform encephalopathy to mouse lemurs after oral or intracerebral inoculation with infected bovine brain tissue. After neurologic symptoms appeared, transmissibility of the disease by both inoculation routes was confirmed by detection of disease-associated prion protein in samples of brain tissue.