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Analysis of α-synuclein species enriched from cerebral cortex of humans with sporadic dementia with Lewy bodies.

ABSTRACT: Since researchers identified α-synuclein as the principal component of Lewy bodies and Lewy neurites, studies have suggested that it plays a causative role in the pathogenesis of dementia with Lewy bodies and other 'synucleinopathies'. While α-synuclein dyshomeostasis likely contributes to the neurodegeneration associated with the synucleinopathies, few direct biochemical analyses of α-synuclein from diseased human brain tissue currently exist. In this study, we analysed sequential protein extracts from a substantial number of patients with neuropathological diagnoses of dementia with Lewy bodies and corresponding controls, detecting a shift of cytosolic and membrane-bound physiological α-synuclein to highly aggregated forms. We then fractionated aqueous extracts (cytosol) from cerebral cortex using non-denaturing methods to search for soluble, disease-associated high molecular weight species potentially associated with toxicity. We applied these fractions and corresponding insoluble fractions containing Lewy-type aggregates to several reporter assays to determine their bioactivity and cytotoxicity. Ultimately, high molecular weight cytosolic fractions enhances phospholipid membrane permeability, while insoluble, Lewy-associated fractions induced morphological changes in the neurites of human stem cell-derived neurons. While the concentrations of soluble, high molecular weight α-synuclein were only slightly elevated in brains of dementia with Lewy bodies patients compared to healthy, age-matched controls, these observations suggest that a small subset of soluble α-synuclein aggregates in the brain may drive early pathogenic effects, while Lewy body-associated α-synuclein can drive neurotoxicity.

SUBMITTER: Sanderson JB

PROVIDER: S-EPMC7130446 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Analysis of α-synuclein species enriched from cerebral cortex of humans with sporadic dementia with Lewy bodies.

Sanderson John B JB De Suman S Jiang Haiyang H Rovere Matteo M Jin Ming M Zaccagnini Ludovica L Hays Watson Aurelia A De Boni Laura L Lagomarsino Valentina N VN Young-Pearse Tracy L TL Liu Xinyue X Pochapsky Thomas C TC Hyman Bradley T BT Dickson Dennis W DW Klenerman David D Selkoe Dennis J DJ Bartels Tim T

Brain communications 20200211 1

Since researchers identified α-synuclein as the principal component of Lewy bodies and Lewy neurites, studies have suggested that it plays a causative role in the pathogenesis of dementia with Lewy bodies and other 'synucleinopathies'. While α-synuclein dyshomeostasis likely contributes to the neurodegeneration associated with the synucleinopathies, few direct biochemical analyses of α-synuclein from diseased human brain tissue currently exist. In this study, we analysed sequential protein extra ...[more]

PMID: 32280944

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Project description:Dementia with Lewy bodies is characterized by the accumulation of Lewy bodies and Lewy neurites in the CNS, both of which are composed mainly of aggregated α-synuclein phosphorylated at Ser129. Although phosphorylated α-synuclein is believed to exert toxic effects at the synapse in dementia with Lewy bodies and other α-synucleinopathies, direct evidence for the precise synaptic localization has been difficult to achieve due to the lack of adequate optical microscopic resolution to study human synapses. In the present study we applied array tomography, a microscopy technique that combines ultrathin sectioning of tissue with immunofluorescence allowing precise identification of small structures, to quantitatively investigate the synaptic phosphorylated α-synuclein pathology in dementia with Lewy bodies. We performed array tomography on human brain samples from five patients with dementia with Lewy bodies, five patients with Alzheimer's disease and five healthy control subjects to analyse the presence of phosphorylated α-synuclein immunoreactivity at the synapse and their relationship with synapse size. Main analyses were performed in blocks from cingulate cortex and confirmed in blocks from the striatum of cases with dementia with Lewy bodies. A total of 1 318 700 single pre- or postsynaptic terminals were analysed. We found that phosphorylated α-synuclein is present exclusively in dementia with Lewy bodies cases, where it can be identified in the form of Lewy bodies, Lewy neurites and small aggregates (<0.16 µm3). Between 19% and 25% of phosphorylated α-synuclein deposits were found in presynaptic terminals mainly in the form of small aggregates. Synaptic terminals that co-localized with small aggregates of phosphorylated α-synuclein were significantly larger than those that did not. Finally, a gradient of phosphorylated α-synuclein aggregation in synapses (pre > pre + post > postsynaptic) was observed. These results indicate that phosphorylated α-synuclein is found at the presynaptic terminals of dementia with Lewy bodies cases mainly in the form of small phosphorylated α-synuclein aggregates that are associated with changes in synaptic morphology. Overall, our data support the notion that pathological phosphorylated α-synuclein may disrupt the structure and function of the synapse in dementia with Lewy bodies.

Project description:BackgroundThe objective of this study was to investigate the discriminating value of a range of CSF α-synuclein species for dementia with Lewy bodies compared with Alzheimer's disease, PD, and cognitively normal controls.MethodsWe applied our recently published enzyme-linked immunosorbent assays to measure the CSF levels of total α-synuclein, oligomeric α-synuclein, and phosphorylated α-synuclein in dementia with Lewy bodies (n = 42), Alzheimer's disease (n = 39), PD (n = 46), and controls (n = 78). General linear models corrected for age and sex were performed to assess differences in α-synuclein levels between groups. We used backward-elimination logistic regression analysis to investigate the combined discriminating value of the different CSF α-synuclein species and Alzheimer's disease biomarkers.ResultsCSF levels of total α-synuclein were lower in dementia with Lewy bodies and PD compared with Alzheimer's disease as well as controls (P < 0.001). In contrast, CSF levels of oligomeric α-synuclein were higher in dementia with Lewy bodies and PD compared with Alzheimer's disease (P < 0.05) and controls (P < 0.001). No group differences were found for phosphorylated α-synuclein. In dementia with Lewy bodies and PD, CSF total α-synuclein levels positively correlated with tau and phosphorylated tau (both r > 0.40, P < 0.01), but not with amyloid-β1-42 . The optimal combination to differentiate dementia with Lewy bodies from controls consisted of amyloid-β1-42 , tau, total α-synuclein, oligomeric α-synuclein, age, and sex (AUC, 0.90). To differentiate dementia with Lewy bodies from Alzheimer's disease, the combination of tau and oligomeric α-synuclein resulted in an AUC of 0.83. CSF α-synuclein species do not contribute to the differentiation of dementia with Lewy bodies from PD.ConclusionsCSF α-synuclein species could be useful as part of a biomarker panel for dementia with Lewy bodies. Evaluating both oligomeric α-synuclein and total α-synuclein in CSF helps in the diagnosis of dementia with Lewy bodies. © 2018 The Authors. Movement Disorders published by Wiley Periodicals, Inc. on behalf of International Parkinson and Movement Disorder Society.

Project description:In patients with suspected dementia with Lewy bodies, the detection of the disease-associated α-synuclein in easily accessible tissues amenable to be collected using minimally invasive procedures remains a major diagnostic challenge. This approach has the potential to take advantage of modern molecular assays for the diagnosis of α-synucleinopathy and, in turn, to optimize the recruitment and selection of patients in clinical trials, using drugs directed at counteracting α-synuclein aggregation. In this study, we explored the diagnostic accuracy of α-synuclein real-time quaking-induced conversion assay by testing olfactory mucosa and CSF in patients with a clinical diagnosis of probable (n = 32) or prodromal (n = 5) dementia with Lewy bodies or mixed degenerative dementia (dementia with Lewy bodies/Alzheimer's disease) (n = 6). Thirty-eight patients with non-α-synuclein-related neurodegenerative and non-neurodegenerative disorders, including Alzheimer's disease (n = 10), sporadic Creutzfeldt-Jakob disease (n = 10), progressive supranuclear palsy (n = 8), corticobasal syndrome (n = 1), fronto-temporal dementia (n = 3) and other neurological conditions (n = 6) were also included, as controls. All 81 patients underwent olfactory swabbing while CSF was obtained in 48 participants. At the initial blinded screening of olfactory mucosa samples, 38 out of 81 resulted positive while CSF was positive in 19 samples out of 48 analysed. After unblinding of the results, 27 positive olfactory mucosa were assigned to patients with probable dementia with Lewy bodies, five with prodromal dementia with Lewy bodies and three to patients with mixed dementia, as opposed to three out 38 controls. Corresponding results of CSF testing disclosed 10 out 10 positive samples in patients with probable dementia with Lewy bodies and six out of six with mixed dementia, in addition to three out of 32 for controls. The accuracy among results of real-time quaking-induced conversion assays and clinical diagnoses was 86.4% in the case of olfactory mucosa and 93.8% for CSF. For the first time, we showed that α-synuclein real-time quaking-induced conversion assay detects α-synuclein aggregates in olfactory mucosa of patients with dementia with Lewy bodies and with mixed dementia. Additionally, we provided preliminary evidence that the combined testing of olfactory mucosa and CSF raised the concordance with clinical diagnosis potentially to 100%. Our results suggest that nasal swabbing might be considered as a first-line screening procedure in patients with a diagnosis of suspected dementia with Lewy bodies followed by CSF analysis, as a confirmatory test, when the result in the olfactory mucosa is incongruent with the initial clinical diagnosis.

Dataset Information

Analysis of α-synuclein species enriched from cerebral cortex of humans with sporadic dementia with Lewy bodies.

Publications

Analysis of α-synuclein species enriched from cerebral cortex of humans with sporadic dementia with Lewy bodies.

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