Τίτλος:
Cell-produced α-synuclein is secreted in a calcium-dependent manner by exosomes and impacts neuronal survival
Γλώσσες Τεκμηρίου:
Αγγλικά
Περίληψη:
α-Synuclein is central in Parkinson's disease pathogenesis. Although initially α-synuclein was considered a purely intracellular protein, recent data suggest that it can be detected in the plasma and CSF of humans and in the culture media of neuronal cells. To address a role of secreted α-synuclein in neuronal homeostasis, we have generated wild-type α-synuclein and β-galactosidase inducible SH-SY5Y cells. Soluble oligomeric and monomeric species of α-synuclein are readily detected in the conditioned media (CM) of these cells at concentrations similar to those observed in human CSF. We have found that, in this model, α-synuclein is secreted by externalized vesicles in a calcium-dependent manner. Electron microscopy and liquid chromatography-mass spectrometry proteomic analysis demonstrate that these vesicles have the characteristic hallmarks of exosomes, secreted intraluminar vesicles of multivesicular bodies. Application of CM containing secreted α-synuclein causes cell death of recipient neuronal cells, which can be reversed after α-synuclein immunodepletion from the CM. High- and low-molecular-weight α-synuclein species, isolated from this CM, significantly decrease cell viability. Importantly, treatment of the CM with oligomer-interfering compounds before application rescues the recipient neuronal cells from the observed toxicity. Our results show for the first time that cell-produced α-synuclein is secreted via an exosomal, calcium-dependent mechanism and suggest that α-synuclein secretion serves to amplify and propagate Parkinson's disease-related pathology. Copyright © 2010 the authors.
Συγγραφείς:
Emmanouilidou, E.
Melachroinou, K.
Roumeliotis, T.
Garbis, S.D.
Ntzouni, M.
Margaritis, L.H.
Stefanis, L.
Vekrellis, K.
Περιοδικό:
Journal of Neuroscience Methods
Λέξεις-κλειδιά:
alpha synuclein; beta galactosidase; calcium; monomer; oligomer, article; cell death; cell lysate; cell proliferation; cell secretion; cell survival; cell viability; concentration (parameters); confocal microscopy; cytotoxicity; electron microscopy; endocytosis; exosome; homeostasis; human; human cell; human cell culture; immunocytochemistry; immunoprecipitation; isotope labeling; liquid chromatography; mass spectrometry; membrane vesicle; nerve cell; Parkinson disease; pathogenesis; priority journal; protein analysis; protein secretion; proteomics, alpha-Synuclein; Analysis of Variance; Animals; beta-Galactosidase; Brefeldin A; Calcium; Cell Death; Cell Differentiation; Cell Line, Tumor; Cell Proliferation; Cell Survival; Cells, Cultured; Cerebral Cortex; Chromatography, High Pressure Liquid; Culture Media, Conditioned; Cytotoxicity Tests, Immunologic; Dose-Response Relationship, Drug; Endocytosis; Exosomes; Gene Expression Regulation, Neoplastic; Humans; Hydrogen Peroxide; Immunoprecipitation; Mass Spectrometry; Microscopy, Confocal; Microscopy, Electron, Transmission; Molecular Weight; Multivesicular Bodies; Nerve Tissue Proteins; Neuroblastoma; Neurons; Peptides; Piperidines; Presenilin-1; Protein Synthesis Inhibitors; Pyrazoles; Rats; Receptors, Transferrin; Serum; Subcellular Fractions; Temperature; Transfection
DOI:
10.1523/JNEUROSCI.5699-09.2010
