Enzyme-linked immunosorbent assays for alpha-synuclein with species and multimeric state specificities

Lee, He Jin; Bae, Eun Jin; Jang, Ara; Ho, Dong Hwan; Cho, Eun Duk; Suk, Ji Eun; Yun, Yeo‐Min; Lee, Seung-Jae

doi:10.1016/j.jneumeth.2011.05.020

Cited by 24 publications

(25 citation statements)

References 38 publications

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“…ELISA was performed as described previously 55 with anti-α-synuclein antibodies Ab62 (1 μg/ml) and biotinylated Ab274 (1 μg/ml) as a capture and reporter, respectively.…”

Section: Methodsmentioning

confidence: 99%

Neuron-released oligomeric α-synuclein is an endogenous agonist of TLR2 for paracrine activation of microglia

et al. 2013

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Abnormal aggregation of α-synuclein and sustained microglial activation are important contributors to the pathogenic processes in Parkinson's disease. However, the relationship between disease-associated protein aggregation and microglia-mediated neuroinflammation remains unknown. Here, using a combination of in silico, in vitro, and in vivo approaches, we show that extracellular α-synuclein released from neuronal cells is an endogenous agonist for toll-like receptor 2 (TLR2), which activates inflammatory responses in microglia. TLR2 ligand activity of α-synuclein is conformation-sensitive; only specific types of oligomer can interact with and activate TLR2. This paracrine interaction between neuron-released oligomeric α-synuclein and TLR2 in microglia suggests that both of these proteins are novel therapeutic targets for modification of neuroinflammation in Parkinson's disease and related neurological diseases.

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“…ELISA was performed as described previously 55 with anti-α-synuclein antibodies Ab62 (1 μg/ml) and biotinylated Ab274 (1 μg/ml) as a capture and reporter, respectively.…”

Section: Methodsmentioning

confidence: 99%

Neuron-released oligomeric α-synuclein is an endogenous agonist of TLR2 for paracrine activation of microglia

et al. 2013

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“…The assay was conducted as previously described (30). Among the antibodies used, Ab274 is specific to human asynuclein, while Ab62 and syn-1 recognize both human and mouse a-synuclein.…”

Section: Enzyme-linked Immunosorbent Assaymentioning

confidence: 99%

“…Antibodies 274 and 62 were produced as previously described (30). Fluorescence dyeconjugated goat anti-rabbit IgG was obtained from Jackson Immunoresearch Laboratories.…”

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confidence: 99%

Lipid Peroxidation Product 4-Hydroxy-2-Nonenal Promotes Seeding-Capable Oligomer Formation and Cell-to-Cell Transfer of α-Synuclein

Bae

Park

et al. 2013

Antioxidants & Redox Signaling

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Aims: Abnormal accumulation of a-synuclein aggregates is one of the key pathological features of many neurodegenerative movement disorders and dementias. These pathological aggregates propagate into larger brain regions as the disease progresses, with the associated clinical symptoms becoming increasingly severe and complex. However, the factors that induce a-synuclein aggregation and spreading of the aggregates remain elusive. Herein, we have evaluated the effects of the major lipid peroxidation byproduct 4-hydroxy-2-nonenal (HNE) on a-synuclein oligomerization and cell-to-cell transmission of this protein. Results: Incubation with HNE promoted the oligomerization of recombinant human a-synuclein via adduct formation at the lysine and histidine residues. HNE-induced a-synuclein oligomers evidence a little b-sheet structure and are distinct from amyloid fibrils at both conformation and ultrastructure levels. Nevertheless, the HNE-induced oligomers are capable of seeding the amyloidogenesis of monomeric a-synuclein under in vitro conditions. When neuronal cells were treated with HNE, both the translocation of a-synuclein into vesicles and the release of this protein from cells were increased. Neuronal cells can internalize HNE-modified a-synuclein oligomers, and HNE treatment increased the cell-to-cell transfer of a-synuclein proteins. Innovation and Conclusion: These results indicate that HNE induces the oligomerization of a-synuclein through covalent modification and promotes the cell-to-cell transfer of seedingcapable oligomers, thereby contributing to both the initiation and spread of a-synuclein aggregates. Antioxid. Redox Signal. 18,[770][771][772][773][774][775][776][777][778][779][780][781][782][783]

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“…The primary antibodies used are as follows: α-synuclein monoclonal antibody (Syn-1; BD Biosciences, San Diego, CA, USA), human α-synuclein specific monoclonal antibody 274 from our laboratory [20], anti-glial fibrillary acidic protein (GFAP) antibody (Abcam, Cambridge, MA, USA), and anti-MHC-II antibody (eBioscience, San Diego, CA, USA).…”

Section: Methodsmentioning

confidence: 99%

Transmission of Synucleinopathies in the Enteric Nervous System of A53T Alpha-Synuclein Transgenic Mice

et al. 2011

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Parkinson's disease (PD) and dementia with Lewy bodies (DLB) are characterized by abnormal deposition of α-synuclein aggregates in many regions of the central and peripheral nervous systems. Accumulating evidence suggests that the α-synuclein pathology initiates in a few discrete regions and spreads to larger areas in the nervous system. Recent pathological studies of PD patients have raised the possibility that the enteric nervous system is one of the initial sites of α-synuclein aggregation and propagation. Here, we evaluated the induction and propagation of α-synuclein aggregates in the enteric nervous system of the A53T α-synuclein transgenic mice after injection of human brain tissue extracts into the gastric walls of the mice. Western analysis of the brain extracts showed that the DLB extract contained detergent-stable α-synuclein aggregates, but the normal brain extract did not. Injection of the DLB extract resulted in an increased deposition of α-synuclein in the myenteric neurons, in which α-synuclein formed punctate aggregates over time up to 4 months. In these mice, inflammatory responses were increased transiently at early time points. None of these changes were observed in the A53T mice injected with saline or the normal brain extract, nor were these found in the wild type mice injected with the DLB extract. These results demonstrate that pathological α-synuclein aggregates present in the brain of DLB patient can induce the aggregation of endogenous α-synuclein in the myenteric neurons in A53T mice, suggesting the transmission of synucleinopathy lesions in the enteric nervous system.

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Enzyme-linked immunosorbent assays for alpha-synuclein with species and multimeric state specificities

Cited by 24 publications

References 38 publications

Neuron-released oligomeric α-synuclein is an endogenous agonist of TLR2 for paracrine activation of microglia

Neuron-released oligomeric α-synuclein is an endogenous agonist of TLR2 for paracrine activation of microglia

Lipid Peroxidation Product 4-Hydroxy-2-Nonenal Promotes Seeding-Capable Oligomer Formation and Cell-to-Cell Transfer of α-Synuclein

Transmission of Synucleinopathies in the Enteric Nervous System of A53T Alpha-Synuclein Transgenic Mice

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