Ultrasensitive Detection of Aggregated ?-Synuclein in Glial Cells, Human Cerebrospinal Fluid, and Brain Tissue Using the RT-QuIC Assay: New High-Throughput Neuroimmune Biomarker Assay for Parkinsonian Disorders.

ABSTRACT: Adult-onset neurodegenerative disorders, like Parkinson's disease (PD) and dementia with Lewy bodies (DLB), that share the accumulation of aggregated ?-synuclein (?Synagg) as their hallmark molecular pathology are collectively known as ?-synucleinopathies. Diagnosing ?-synucleinopathies requires the post-mortem detection of ?Synagg in various brain regions. Recent efforts to measure ?Synagg in living patients include quantifying ?Synagg in different biofluids as a biomarker for PD. We adopted the real-time quaking-induced conversion (RT-QuIC) assay to detect very low levels of ?Synagg. We first optimized RT-QuIC for sensitivity, specificity, and reproducibility by using monomeric recombinant human wild-type ?Syn as a substrate and ?Synagg as the seed. Next, we exposed mouse microglia to ?Syn pre-formed fibrils (?SynPFF) for 24 h. RT-QuIC assay revealed that the ?SynPFF is taken up rapidly by mouse microglia, within 30 min, and cleared within 24 h. We then evaluated the ?Syn RT-QuIC assay for detecting ?Synagg in human PD, DLB, and Alzheimer's disease (AD) post-mortem brain homogenates (BH) along with PD and progressive supranuclear palsy (PSP) cerebrospinal fluid (CSF) samples and then determined protein aggregation rate (PAR) for ?Synagg. The PD and DLB BH samples not only showed significantly higher ?Synagg PAR compared to age-matched healthy controls and AD, but RT-QuIC was also highly reproducible with 94% sensitivity and 100% specificity. Similarly, PD CSF samples demonstrated significantly higher ?Synagg PAR compared to age-matched healthy controls, with 100% sensitivity and specificity. Overall, the RT-QuIC assay accurately detects ?Synagg seeding activity, offering a potential tool for antemortem diagnosis of ?-synucleinopathies and other protein-misfolding disorders. Graphical Abstract A schematic representation of ?Syn RT-QuIC assay.

SUBMITTER: Manne S 

PROVIDER: S-EPMC6669119 | biostudies-literature |

REPOSITORIES: biostudies-literature