Neuroinvasion of α-Synuclein Prionoids after Intraperitoneal and Intraglossal Inoculation.

UNLABELLED: α-Synuclein is a soluble, cellular protein that in a number of neurodegenerative diseases, including Parkinson's disease and multiple system atrophy, forms pathological deposits of protein aggregates. Because misfolded α-synuclein has some characteristics that resemble those of prions, we investigated its potential to induce disease after intraperitoneal or intraglossal challenge injection into bigenic Tg(M83(+/-):Gfap-luc(+/-)) mice, which express the A53T mutant of human α-synuclein and firefly luciferase. After a single intraperitoneal injection with α-synuclein fibrils, four of five mice developed paralysis and α-synuclein pathology in the central nervous system, with a median incubation time of 229 ± 17 days. Diseased mice accumulated aggregates of Sarkosyl-insoluble and phosphorylated α-synuclein in the brain and spinal cord, which colocalized with ubiquitin and p62 and were accompanied by gliosis. In contrast, only one of five mice developed α-synuclein pathology in the central nervous system after intraglossal injection with α-synuclein fibrils, after 285 days. These findings are novel and important because they show that, similar to prions, α-synuclein prionoids can neuroinvade the central nervous system after intraperitoneal or intraglossal injection and can cause neuropathology and disease. IMPORTANCE: Synucleinopathies are neurodegenerative diseases that are characterized by the pathological presence of aggregated α-synuclein in cells of the nervous system. Previous studies have shown that α-synuclein aggregates made of recombinant protein or derived from brains of patients can spread in the central nervous system in a spatiotemporal manner when inoculated into the brains of animals and can induce pathology and neurologic disease, suggesting that misfolded α-synuclein can behave similarly to prions. Here we show that α-synuclein inoculation into the peritoneal cavity or the tongue in mice overexpressing α-synuclein causes neurodegeneration after neuroinvasion from the periphery, which further corroborates the prionoid character of misfolded α-synuclein.

The Anti-amyloid Compound DO1 Decreases Plaque Pathology and Neuroinflammation-Related Expression Changes in 5xFAD Transgenic Mice.

Self-propagating amyloid-ß (Aß) aggregates or seeds possibly drive pathogenesis of Alzheimer's disease (AD). Small molecules targeting such structures might act therapeutically in vivo. Here, a fluorescence polarization assay was established that enables the detection of compound effects on both seeded and spontaneous Aß42 aggregation. In a focused screen of anti-amyloid compounds, we identified Disperse Orange 1 (DO1) ([4-((4-nitrophenyl)diazenyl)-N-phenylaniline]), a small molecule that potently delays both seeded and non-seeded Aß42 polymerization at substoichiometric concentrations. Mechanistic studies revealed that DO1 disrupts preformed fibrillar assemblies of synthetic Aß42 peptides and decreases the seeding activity of Aß aggregates from brain extracts of AD transgenic mice. DO1 also reduced the size and abundance of diffuse Aß plaques and decreased neuroinflammation-related gene expression changes in brains of 5xFAD transgenic mice. Finally, improved nesting behavior was observed upon treatment with the compound. Together, our evidence supports targeting of self-propagating Aß structures with small molecules as a valid therapeutic strategy.

Prion-like propagation of human brain-derived alpha-synuclein in transgenic mice expressing human wild-type alpha-synuclein.

INTRODUCTION: Parkinson's disease (PD) and multiple system atrophy (MSA) are neurodegenerative diseases that are characterized by the intracellular accumulation of alpha-synuclein containing aggregates. Recent increasing evidence suggests that Parkinson's disease and MSA pathology spread throughout the nervous system in a spatiotemporal fashion, possibly by prion-like propagation of alpha-synuclein positive aggregates between synaptically connected areas. Concurrently, intracerebral injection of pathological alpha-synuclein into transgenic mice overexpressing human wild-type alpha-synuclein, or human alpha-synuclein with the familial A53T mutation, or into wild-type mice causes spreading of alpha-synuclein pathology in the CNS. Considering that wild-type mice naturally also express a threonine at codon 53 of alpha-synuclein, it has remained unclear whether human wild-type alpha-synuclein alone, in the absence of endogenously expressed mouse alpha-synuclein, would support a similar propagation of alpha-synuclein pathology in vivo. RESULTS: Here we show that brain extracts from two patients with MSA and two patients with probable incidental Lewy body disease (iLBD) but not phosphate-buffered saline induce prion-like spreading of pathological alpha-synuclein after intrastriatal injection into mice expressing human wild-type alpha-synuclein. Mice were sacrificed at 3, 6, and 9 months post injection and analyzed neuropathologically and biochemically. Mice injected with brain extracts from patients with MSA or probable iLBD both accumulated intraneuronal inclusion bodies, which stained positive for phosphorylated alpha-synuclein and appeared predominantly within the injected brain hemisphere after 6 months. After 9 months these intraneuronal inclusion bodies had spread to the contralateral hemisphere and more rostral and caudal areas. Biochemical analysis showed that brains of mice injected with brain extracts from patients with MSA and probable iLBD contained hyperphosphorylated alpha-synuclein that also seeded aggregation of recombinant human wild-type alpha-synuclein in a Thioflavin T binding assay. CONCLUSIONS: Our results indicate that human wild-type alpha-synuclein supports the prion-like spreading of alpha-synuclein pathology in the absence of endogenously expressed mouse alpha-synuclein in vivo.