Dataset Information

Detecting Alpha Synuclein Seeding Activity in Formaldehyde-Fixed MSA Patient Tissue by PMCA.

ABSTRACT: Alpha synuclein (?-syn) is central to the pathogenesis of a group of neurodegenerative disorders known as synucleinopathies, including Parkinson's disease (PD), dementia with Lewy bodies (DLB), and multiple system atrophy (MSA). Aggregation of ?-syn is the pathologic hallmark of these disorders and is intimately associated with the pathogenic changes. The prion-like hypothesis postulates that the aggregated ?-syn provides a template to seed the aggregation of normal ?-syn and spread the pathology. Thus far, it remains unclear whether aggregated ?-syn can be a useful biomarker for diagnosis and/or tracking disease progression, which is mainly due to the lack of a suitable biochemical assay. The protein misfolding cyclic amplification (PMCA) technique is known for its enormous amplification power to detect the seeding activity of protein aggregates such as prions. In this study, we adapted PMCA for detecting the seeding activity of ?-syn. By extensively optimizing the PMCA parameters, we developed a protocol that is able to sensitively and quantitatively detect the seeding activity of as little as 100 attomoles (10^-16 mol) of ?-syn aggregate. Using our protocol, we detected ?-syn seeding activity from a histologically positive, formaldehyde-fixed MSA sample, but not with the histologically negative, formaldehyde-fixed control sample. Our results confirmed that the ?-syn in MSA patient's brain does contain seeding activity, which remains active even after fixation. Moreover, we also established that PMCA with sonication is a sensitive and quantitative method for detecting ?-syn seeding activity, which can be further adapted to more accessible patients' samples to evaluate ?-syn aggregates as a biomarker for synucleinopathies.

SUBMITTER: Becker K

PROVIDER: S-EPMC6153717 | biostudies-literature | 2018 Nov

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Detecting Alpha Synuclein Seeding Activity in Formaldehyde-Fixed MSA Patient Tissue by PMCA.

Becker Katelyn K Wang Xinhe X Vander Stel Kayla K Chu Yaping Y Kordower Jeffrey J Ma Jiyan J

Molecular neurobiology 20180327 11

Alpha synuclein (α-syn) is central to the pathogenesis of a group of neurodegenerative disorders known as synucleinopathies, including Parkinson's disease (PD), dementia with Lewy bodies (DLB), and multiple system atrophy (MSA). Aggregation of α-syn is the pathologic hallmark of these disorders and is intimately associated with the pathogenic changes. The prion-like hypothesis postulates that the aggregated α-syn provides a template to seed the aggregation of normal α-syn and spread the patholog ...[more]

PMID: 29589283

Similar Datasets

Project description:Tauopathies such as Alzheimer's disease (AD) feature progressive intraneuronal deposition of aggregated tau protein. The cause is unknown, but in experimental systems trans-cellular propagation of tau pathology resembles prion pathogenesis. Tau aggregate inoculation into mice produces transmissible pathology, and tau forms distinct strains, i.e. conformers that faithfully replicate and create predictable patterns of pathology in vivo. The prion model predicts that tau seed formation will anticipate neurofibrillary tau pathology. To test this idea requires simultaneous assessment of seed titer and immunohistochemistry (IHC) of brain tissue, but it is unknown whether tau seed titer can be determined in formaldehyde-fixed tissue. We have previously created a cellular biosensor system that uses flow cytometry to quantify induced tau aggregation and thus determine seed titer. In unfixed tissue from PS19 tauopathy mice that express 1 N,4R tau (P301S), we have measured tau seeding activity that precedes the first observable histopathology by many months. Additionally, in fresh frozen tissue from human AD subjects at early to mid-neurofibrillary tangle stages (NFT I-IV), we have observed tau seeding activity in cortical regions predicted to lack neurofibrillary pathology. However, we could not directly compare the same regions by IHC and seeding activity in either case. We now describe a protocol to extract and measure tau seeding activity from small volumes (.04 mm3) of formaldehyde-fixed tissue immediately adjacent to that used for IHC. We validated this method with the PS19 transgenic mouse model, and easily observed seeding well before the development of phospho-tau pathology. We also accurately isolated two tau strains, DS9 and DS10, from fixed brain tissues in mice. Finally, we have observed robust seeding activity in fixed AD brain, but not controls. The successful coupling of classical IHC with seeding and strain detection should enable detailed study of banked brain tissue in AD and other tauopathies.

Dataset Information

Detecting Alpha Synuclein Seeding Activity in Formaldehyde-Fixed MSA Patient Tissue by PMCA.

Publications

Detecting Alpha Synuclein Seeding Activity in Formaldehyde-Fixed MSA Patient Tissue by PMCA.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets