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?-Synuclein propagates from mouse brain to grafted dopaminergic neurons and seeds aggregation in cultured human cells.

ABSTRACT: Post-mortem analyses of brains from patients with Parkinson disease who received fetal mesencephalic transplants show that ?-synuclein-containing (?-syn-containing) Lewy bodies gradually appear in grafted neurons. Here, we explored whether intercellular transfer of ?-syn from host to graft, followed by seeding of ?-syn aggregation in recipient neurons, can contribute to this phenomenon. We assessed ?-syn cell-to-cell transfer using microscopy, flow cytometry, and high-content screening in several coculture model systems. Coculturing cells engineered to express either GFP- or DsRed-tagged ?-syn resulted in a gradual increase in double-labeled cells. Importantly, ?-syn-GFP derived from 1 neuroblastoma cell line localized to red fluorescent aggregates in other cells expressing DsRed-?-syn, suggesting a seeding effect of transmitted ?-syn. Extracellular ?-syn was taken up by cells through endocytosis and interacted with intracellular ?-syn. Next, following intracortical injection of recombinant ?-syn in rats, we found neuronal uptake was attenuated by coinjection of an endocytosis inhibitor. Finally, we demonstrated in vivo transfer of ?-syn between host cells and grafted dopaminergic neurons in mice overexpressing human ?-syn. In summary, intercellularly transferred ?-syn interacts with cytoplasmic ?-syn and can propagate ?-syn pathology. These results suggest that ?-syn propagation is a key element in the progression of Parkinson disease pathology.

SUBMITTER: Hansen C

PROVIDER: S-EPMC3026723 | biostudies-literature | 2011 Feb

REPOSITORIES: biostudies-literature

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α-Synuclein propagates from mouse brain to grafted dopaminergic neurons and seeds aggregation in cultured human cells.

Hansen Christian C Angot Elodie E Bergström Ann-Louise AL Steiner Jennifer A JA Pieri Laura L Paul Gesine G Outeiro Tiago F TF Melki Ronald R Kallunki Pekka P Fog Karina K Li Jia-Yi JY Brundin Patrik P

The Journal of clinical investigation 20110118 2

Post-mortem analyses of brains from patients with Parkinson disease who received fetal mesencephalic transplants show that α-synuclein-containing (α-syn-containing) Lewy bodies gradually appear in grafted neurons. Here, we explored whether intercellular transfer of α-syn from host to graft, followed by seeding of α-syn aggregation in recipient neurons, can contribute to this phenomenon. We assessed α-syn cell-to-cell transfer using microscopy, flow cytometry, and high-content screening in severa ...[more]

PMID: 21245577

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Project description:Alpha-synuclein pathology is associated with dopaminergic neuronal loss in the substantia nigra (SN) of Parkinson's patients. Working across human and mouse models, we investigated mechanisms by which the accumulation of soluble α-synuclein oligomers leads to neurodegeneration. Biochemical analysis of the midbrain of α-synuclein overexpressing BAC-transgenic male and female mice revealed age- and region-dependent mitochondrial dysfunction and accumulation of damaged proteins downstream of the RE1 Silencing Transcription Factor (REST). Vulnerable SN dopaminergic neurons displayed low REST levels compared with neighboring protected SN GABAergic neurons, which correlated with the accumulation of α-synuclein oligomers and disrupted mitochondrial morphology. Consistent with a protective role, REST levels were reduced in patient induced pluripotent stem cell-derived dopaminergic neurons carrying the SNCA-Triplication mutation, which accumulated α-synuclein oligomers and mitochondrial damage, and displayed REST target gene dysregulation. Furthermore, CRISPR-mediated REST KO induced mitochondrial dysfunction and impaired mitophagy in vitro Conversely, REST overexpression attenuated mitochondrial toxicity and mitochondrial morphology disruption through the transcription factor PGC-1α. Finally, decreased α-synuclein oligomer accumulation and mitochondrial dysfunction in mice correlated with nuclear REST and PGC-1α in protected SN GABAergic neurons compared with vulnerable dopaminergic neurons. Our findings show that increased levels of α-synuclein oligomers cause dopaminergic neuronal-specific dysfunction through mitochondrial toxicity, which can be attenuated by REST in an early model of Parkinsonian pathology. These findings highlight REST as a mediator of dopaminergic vulnerability in PD.SIGNIFICANCE STATEMENT Understanding early Parkinsonian pathophysiology through studies of advanced preclinical models is fundamental to the translation of disease-modifying therapies. Here we show disease-relevant levels of α-synuclein expression in mice leads to accumulation of α-synuclein oligomers in the absence of overt aggregation, and mitochondrial dysfunction in dopaminergic neurons lacking the RE1 Silencing Transcription Factor. Our findings identify the mechanism of action of RE1 Silencing Transcription Factor and PGC-1α as mediators of dopaminergic vulnerability in α-synuclein BAC-transgenic mice and induced pluripotent stem cell-derived dopaminergic cultures, highlighting their potential as therapeutic targets.

Project description:Synucleinopathies are characterized by the accumulation of insoluble ?-synuclein (?Syn). To test whether ?Syn aggregates modulate synaptic activity, we used a recently developed model in primary neurons for inducing ?Syn pathology. We demonstrated that preformed fibrils (PFFs) generated with recombinant human ?Syn compromised synaptic activity in a time- and dose-dependent manner and that the magnitude of these deficits correlated with the formation of ?Syn pathology in cultured excitatory hippocampal neurons from both sexes of mice. Remarkably, acute passive infusion of ?Syn PFFs from whole-cell patch-clamp pipette decreased mEPSC frequency within 10 min followed by induction of ?Syn pathology within 1 d. Moreover, by direct addition of ?Syn PFFs into culture medium, the formation of misfolded ?Syn inclusions dramatically compromised the colocalization of synaptic markers and altered dynamic changes of dendritic spines, but the viability of neurons was not affected up to 7 d post-treatment with ?Syn PFFs. Our data indicate that intraneuronal ?Syn fibrils impaired the initiation of synaptogenesis and their physiological functions, thereby suggesting that targeting synaptic dysfunction in synucleinopathies may provide a promising therapeutic direction.SIGNIFICANCE STATEMENT Under pathological conditions, the presynaptic protein ?-synuclein (?Syn) aggregates to form intraneuronal inclusions. To understand how and to what extent ?Syn aggregates modulate synaptic activity before neuron loss, we demonstrate that ?Syn preformed fibrils (PFFs) reduced synaptic activity in a dose- and time-dependent manner. The magnitude of these deficits correlated with the deposition of ?Syn pathology, which dramatically compromised the colocalization of synaptic markers and altered the dendritic spine dynamics. The present work further highlights the impact of ?Syn PFFs on synaptogenesis and physiological function, which may be applicable to other types of synucleinopathies.

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?-Synuclein propagates from mouse brain to grafted dopaminergic neurons and seeds aggregation in cultured human cells.

Publications

α-Synuclein propagates from mouse brain to grafted dopaminergic neurons and seeds aggregation in cultured human cells.

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