IMPORTANCE Parkinson disease (PD) is a highly prevalent and incurable neurodegenerative disease associated with the accumulation of misfolded α-synuclein (αSyn) aggregates. An important problem in this disease is the lack of a sensitive, specific, and noninvasive biochemical diagnosis to help in clinical evaluation, monitoring of disease progression, and early differential diagnosis from related neurodegenerative diseases. OBJECTIVE To develop a novel assay with high sensitivity and specificity to detect small quantities of αSyn aggregates circulating in cerebrospinal fluid (CSF) of patients affected by PD and related synucleinopathies. DESIGN, SETTING, AND PARTICIPANTS The strategy evaluated in this proof-of-concept study uses the protein misfolding cyclic amplification (PMCA) technology that detects minute amounts of misfolded oligomers by taking advantage of their ability to nucleate further aggregation, enabling a very high amplification of the signal. The technology was first adapted with synthetic αSyn oligomers prepared in vitro and used to screen in 2 blinded cohorts of CSF samples from German and Japanese patients with PD (n = 76) and individuals serving as controls affected by other neurologic disorders (n = 65), neurodegenerative diseases (n = 18), and Alzheimer disease (n = 14). The kinetics of αSyn aggregation were measured by αSyn-PMCA in the presence of CSF samples from the participants to detect αSyn oligomeric seeds present in this biological fluid. The assays were conducted from November 15, 2013, to August 28, 2015. MAIN OUTCOMES AND MEASURES Kinetic parameters correlated with disease severity at the time of sample collection, measured by the Hoehn and Yahr scale, with the lowest grade indicating unilateral involvement with minimal or no functional impairment, and the highest grade defining patients with complete confinement to wheelchair or bed. RESULTS Studies with synthetic αSyn aggregates showed that αSyn-PMCA enabled to detect as little as 0.1 pg/mL of αSyn oligomers. The αSyn-PMCA signal was directly proportional to the amount of αSyn oligomers added to the reaction. A blinded study of CSF samples correctly identified patients affected by PD with an overall sensitivity of 88.5% (95% CI, 79.2%-94.6%) and specificity of 96.9% (95% CI, 89.3%-99.6%). The αSyn-PMCA results for different patients correlated with the severity of the clinical symptoms of PD (Japanese cohort: r s = −0.54, P = .006; German cohort: r s = −0.36, P = .02). CONCLUSIONS AND RELEVANCE The findings suggest that detection of αSyn oligomers by αSyn-PMCA in the CSF of patients affected by PD may offer a good opportunity for a sensitive and specific biochemical diagnosis of the disease. Further studies are needed to investigate the usefulness of αSyn-PMCA to monitor disease progression and for preclinical identification of patients who may develop PD.
IntroductionAlzheimer’s disease (AD) is the most dominant neurodegenerative disorder that causes dementia, and no effective treatments are available. To study its pathogenesis and develop therapeutics, animal models representing its pathologies are needed. Although many animal species develop senile plaques (SP) composed of amyloid-β (Aβ) proteins that are identical to those found in humans, none of them exhibit neurofibrillary tangles (NFT) and subsequent neurodegeneration, which are integral parts of the pathology of AD.ResultsThe present study shows that Aβ accumulation, NFT formation, and significant neuronal loss all emerge naturally in the hippocampi of aged domestic cats. The NFT that form in the cat brain are identical to those seen in human AD in terms of their spatial distribution, the cells they affect, and the tau isoforms that comprise them. Interestingly, aged cats do not develop mature argyrophilic SP, but instead accumulate intraneuronal Aβ oligomers in their hippocampal pyramidal cells, which might be due to the amino acid sequence of felid Aβ.ConclusionsThese results suggest that Aβ oligomers are more important than SP for NFT formation and the subsequent neurodegeneration. The domestic cat is a unique animal species that naturally replicates various AD pathologies, especially Aβ oligomer accumulation, NFT formation, and neuronal loss.Electronic supplementary materialThe online version of this article (doi:10.1186/s40478-015-0258-3) contains supplementary material, which is available to authorized users.
Amyloid β-protein (Aβ42) oligomerization is an early event in Alzheimer’s disease (AD). Current diagnostic methods using sequence-specific antibodies against less toxic fibrillar and monomeric Aβ42 run the risk of overdiagnosis. Hence, conformation-specific antibodies against neurotoxic Aβ42 oligomers have garnered much attention for developing more accurate diagnostics. Antibody 24B3, highly specific for the toxic Aβ42 conformer that has a turn at Glu22 and Asp23, recognizes a putative Aβ42 dimer, which forms stable and neurotoxic oligomers more potently than the monomer. 24B3 significantly rescues Aβ42-induced neurotoxicity, whereas sequence-specific antibodies such as 4G8 and 82E1, which recognizes the N-terminus, do not. The ratio of toxic to total Aβ42 in the cerebrospinal fluid of AD patients is significantly higher than in control subjects as measured by sandwich ELISA using antibodies 24B3 and 82E1. Thus, 24B3 may be useful for AD diagnosis and therapy.
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