Alpha-Synuclein Gene Alterations Modulate Tyrosine Hydroxylase in Human iPSC-Derived Neurons in a Parkinson's Disease Animal Model.

Parkinson's disease (PD) caused by SNCA gene triplication (3XSNCA) leads to early onset, rapid progression, and often dementia. Understanding the impact of 3XSNCA and its absence is crucial. This study investigates the differentiation of human induced pluripotent stem cell (hiPSC)-derived floor-plate progenitors into dopaminergic neurons. Three different genotypes were evaluated in this study: patient-derived hiPSCs with 3XSNCA, a gene-edited isogenic line with a frame-shift mutation on all SNCA alleles (SNCA 4KO), and a normal wild-type control. Our aim was to assess how the substantia nigra pars compacta (SNpc) microenvironment, damaged by 6-hydroxydopamine (6-OHDA), influences tyrosine hydroxylase-positive (Th+) neuron differentiation in these genetic variations. This study confirms successful in vitro differentiation into neuronal lineage in all cell lines. However, the SNCA 4KO line showed unusual LIM homeobox transcription factor 1 alpha (Lmx1a) extranuclear distribution. Crucially, both 3XSNCA and SNCA 4KO lines had reduced Th+ neuron expression, despite initial successful neuronal differentiation after two months post-transplantation. This indicates that while the SNpc environment supports early neuronal survival, SNCA gene alterations-either amplification or knock-out-negatively impact Th+ dopaminergic neuron maturation. These findings highlight SNCA's critical role in PD and underscore the value of hiPSC models in studying neurodegenerative diseases.

Rotenone Induces a Neuropathological Phenotype in Cholinergic-like Neurons Resembling Parkinson's Disease Dementia (PDD).

Parkinson's disease with dementia (PDD) is a neurological disorder that clinically and neuropathologically overlaps with Parkinson's disease (PD) and Alzheimer's disease (AD). Although it is assumed that alpha-synuclein ( α -Syn), amyloid beta (A ß ), and the protein Tau might synergistically induce cholinergic neuronal degeneration, presently the pathological mechanism of PDD remains unclear. Therefore, it is essential to delve into the cellular and molecular aspects of this neurological entity to identify potential targets for prevention and treatment strategies. Cholinergic-like neurons (ChLNs) were exposed to rotenone (ROT, 10 µ M) for 24 h. ROT provokes loss of Δ Ψ m , generation of reactive oxygen species (ROS), phosphorylation of leucine-rich repeated kinase 2 (LRRK2 at Ser935) concomitantly with phosphorylation of α -synuclein ( α -Syn, Ser129), induces accumulation of intracellular A ß (iA ß ), oxidized DJ-1 (Cys106), as well as phosphorylation of TAU (Ser202/Thr205), increases the phosphorylation of c-JUN (Ser63/Ser73), and increases expression of proapoptotic proteins TP53, PUMA, and cleaved caspase 3 (CC3) in ChLNs. These neuropathological features resemble those reproduced in presenilin 1 (PSEN1) E280A ChLNs. Interestingly, anti-oxidant and anti-amyloid cannabidiol (CBD), JNK inhibitor SP600125 (SP), TP53 inhibitor pifithrin- α (PFT), and LRRK2 kinase inhibitor PF-06447475 (PF475) significantly diminish ROT-induced oxidative stress (OS), proteinaceous, and cell death markers in ChLNs compared to naïve ChLNs. In conclusion, ROT induces p- α -Syn, iA ß , p-Tau, and cell death in ChLNs, recapitulating the neuropathology findings in PDD. Our report provides an excellent in vitro model to test for potential therapeutic strategies against PDD. Our data suggest that ROT induces a neuropathologic phenotype in ChLNs similar to that caused by the mutation PSEN1 E280A.

Accumulation of Alpha-Synuclein and Increase in the Inflammatory Response in the substantia nigra, Jejunum, and Colon in a Model of O3 Pollution in Rats.

This work aimed to study the effect of repeated exposure to low doses of ozone on alpha-synuclein and the inflammatory response in the substantia nigra, jejunum, and colon. Seventy-two male Wistar rats were divided into six groups. Each group received one of the following treatments: The control group was exposed to air. The ozone groups were exposed for 7, 15, 30, 60, and 90 days for 0.25 ppm for four hours daily. Afterward, they were anesthetized, and their tissues were extracted and processed using Western blotting, immunohistochemistry, and qPCR. The results indicated a significant increase in alpha-synuclein in the substantia nigra and jejunum from 7 to 60 days of exposure and an increase in NFκB from 7 to 90 days in the substantia nigra, while in the jejunum, a significant increase was observed at 7 and 15 days and a decrease at 60 and 90 days for the colon. Interleukin IL-17 showed an increase at 90 days in the substantia nigra in the jejunum and increases at 30 days and in the colon at 15 and 90 days. Exposure to ozone increases the presence of alpha-synuclein and induces the loss of regulation of the inflammatory response, which contributes significantly to degenerative processes.

Nicotinic Acetylcholine Receptors in Glial Cells as Molecular Target for Parkinson's Disease.

Parkinson's disease (PD) is a progressive neurodegenerative disease characterized by resting tremor, bradykinesia, rigidity, and postural instability that also includes non-motor symptoms such as mood dysregulation. Dopamine (DA) is the primary neurotransmitter involved in this disease, but cholinergic imbalance has also been implicated. Current intervention in PD is focused on replenishing central DA, which provides remarkable temporary symptomatic relief but does not address neuronal loss and the progression of the disease. It has been well established that neuronal nicotinic cholinergic receptors (nAChRs) can regulate DA release and that nicotine itself may have neuroprotective effects. Recent studies identified nAChRs in nonneuronal cell types, including glial cells, where they may regulate inflammatory responses. Given the crucial role of neuroinflammation in dopaminergic degeneration and the involvement of microglia and astrocytes in this response, glial nAChRs may provide a novel therapeutic target in the prevention and/or treatment of PD. In this review, following a brief discussion of PD, we focus on the role of glial cells and, specifically, their nAChRs in PD pathology and/or treatment.

Alpha-synuclein dynamics bridge Type-I Interferon response and SARS-CoV-2 replication in peripheral cells.

BACKGROUND: Increasing evidence suggests a double-faceted role of alpha-synuclein (α-syn) following infection by a variety of viruses, including SARS-CoV-2. Although α-syn accumulation is known to contribute to cell toxicity and the development and/or exacerbation of neuropathological manifestations, it is also a key to sustaining anti-viral innate immunity. Consistently with α-syn aggregation as a hallmark of Parkinson's disease, most studies investigating the biological function of α-syn focused on neural cells, while reports on the role of α-syn in periphery are limited, especially in SARS-CoV-2 infection. RESULTS: Results herein obtained by real time qPCR, immunofluorescence and western blot indicate that α-syn upregulation in peripheral cells occurs as a Type-I Interferon (IFN)-related response against SARS-CoV-2 infection. Noteworthy, this effect mostly involves α-syn multimers, and the dynamic α-syn multimer:monomer ratio. Administration of excess α-syn monomers promoted SARS-CoV-2 replication along with downregulation of IFN-Stimulated Genes (ISGs) in epithelial lung cells, which was associated with reduced α-syn multimers and α-syn multimer:monomer ratio. These effects were prevented by combined administration of IFN-ß, which hindered virus replication and upregulated ISGs, meanwhile increasing both α-syn multimers and α-syn multimer:monomer ratio in the absence of cell toxicity. Finally, in endothelial cells displaying abortive SARS-CoV-2 replication, α-syn multimers, and multimer:monomer ratio were not reduced following exposure to the virus and exogenous α-syn, suggesting that only productive viral infection impairs α-syn multimerization and multimer:monomer equilibrium. CONCLUSIONS: Our study provides novel insights into the biology of α-syn, showing that its dynamic conformations are implicated in the innate immune response against SARS-CoV-2 infection in peripheral cells. In particular, our results suggest that promotion of non-toxic α-syn multimers likely occurs as a Type-I IFN-related biological response which partakes in the suppression of viral replication. Further studies are needed to replicate our findings in neuronal cells as well as animal models, and to ascertain the nature of such α-syn conformations.

In silico analysis of alpha-synuclein protein variants and posttranslational modifications related to Parkinson's disease.

Parkinson's disease (PD) is among the most prevalent neurodegenerative disorders, affecting over 10 million people worldwide. The protein encoded by the SNCA gene, alpha-synuclein (ASYN), is the major component of Lewy body (LB) aggregates, a histopathological hallmark of PD. Mutations and posttranslational modifications (PTMs) in ASYN are known to influence protein aggregation and LB formation, possibly playing a crucial role in PD pathogenesis. In this work, we applied computational methods to characterize the effects of missense mutations and PTMs on the structure and function of ASYN. Missense mutations in ASYN were compiled from the literature/databases and underwent a comprehensive predictive analysis. Phosphorylation and SUMOylation sites of ASYN were retrieved from databases and predicted by algorithms. ConSurf was used to estimate the evolutionary conservation of ASYN amino acids. Molecular dynamics (MD) simulations of ASYN wild-type and variants A30G, A30P, A53T, and G51D were performed using the GROMACS package. Seventy-seven missense mutations in ASYN were compiled. Although most mutations were not predicted to affect ASYN stability, aggregation propensity, amyloid formation, and chaperone binding, the analyzed mutations received relatively high rates of deleterious predictions and predominantly occurred at evolutionarily conserved sites within the protein. Moreover, our predictive analyses suggested that the following mutations may be possibly harmful to ASYN and, consequently, potential targets for future investigation: K6N, T22I, K34E, G36R, G36S, V37F, L38P, G41D, and K102E. The MD analyses pointed to remarkable flexibility and essential dynamics alterations at nearly all domains of the studied variants, which could lead to impaired contact between NAC and the C-terminal domain triggering protein aggregation. These alterations may have functional implications for ASYN and provide important insight into the molecular mechanism of PD, supporting the design of future biomedical research and improvements in existing therapies for the disease.

Correlato neurobiológico de los síntomas premotores del parkinson: el origen en la teoría de Braak / Neurobiological correlate of parkinson's premotor symptoms: the origin in Braak's theory / Correlação neurobiológica dos sintomas pré-motores de parkinson: a origem na teoria de Braak

El diagnóstico de enfermedad de Parkinson (ED) se basa en las principales manifestaciones motoras: bradicinesia en combinación con temblor en reposo, rigidez o ambos. Cuando se realiza el diagnóstico basado en la sintomatología motora clínica típica ya se han perdido hasta el 60 % de las neuronas dopaminérgicas de la sustancia negra pars compacta mesencefálica. La identificación de los síntomas premotores son un marcador temprano para sospechar la aparición futura de la enfermedad, así como su progresión y gravedad. La hipótesis sobre la patogénesis que mejor expone la progresión de la enfermedad es la teoría de Braak. Esta se basa en la aparición y presencia de cuerpos de Lewy en diferentes estructuras anatómicas, las cuales representadas en cada uno de sus seis estadios y podrían ser la explicación biológica de los síntomas premotores, motores y no motores. La detección temprana de los síntomas premotores puede tener repercusiones positivas en el enfoque, seguimiento, diagnóstico y tratamiento de la EP. El propósito de este artículo es identificar las aproximaciones neurológicas descritas por la teoría de Braak para los síntomas premotores de la enfermedad de Parkinson de acuerdo con la literatura publicada en los últimos 20 años.

The diagnosis of Parkinson's disease (PD) is based on the main motor manifestations: bradykinesia in combination with tremor at rest, rigidity, or both. When the diagnosis is made based on typical clinical motor symptoms, up to 60 % of the dopaminergic neurons of the mesencephalic substantia nigra pars compacta have already been lost. The identification of premotor symptoms is an early marker to suspect the future appearance of the disease, as well as its progression and severity. The hypothesis about the pathogenesis that best exposes the progression of the disease is Braak's theory. It is based on the appearance and presence of Lewy bodies in different anatomical structures, which are represented in each of its six stages and could be the biological explanation biological of premotor, motor, and non-motor symptoms. Early detection of premotor symptoms can have positive repercussions in the approach, follow-up, diagnosis and treatment of PD. The purpose of this article is to identify the neurological approaches described by Braak's theory for the premotor symptoms of Parkinson's disease according to the literature published in the last 20 years.

O diagnóstico da doença de Parkinson (DP) baseia-se nas principais manifestações motoras: bradicinesia combinada com tremor de repouso, rigidez ou ambos. Quando o diagnóstico é feito com base em sintomas clínicos motores típicos, até 60% dos neurônios dopaminérgicos da substância negra pars compacta mesencefálica já foram perdidos. A identificação de sintomas pré-motores é um marcador precoce para suspeitar do futuro aparecimento da doença, bem como da sua progressão e gravidade. A hipótese sobre a patogênese que melhor expõe a progressão da doença é a teoria de Braak. Isto se baseia no aparecimento e presença de corpos de Lewy em diferentes estruturas anatômicas, que estão representados em cada uma de suas seis etapas e podem ser a explicação biológica dos sintomas pré-motores, motores e não motores. A detecção precoce de sintomas pré-motores pode repercutir positivamente na abordagem, acompanhamento, diagnóstico e tratamento da DP. O objetivo deste artigo é identificar as abordagens neurológicas descritas pela teoria de Braak para os sintomas pré-motores da doença de Parkinson de acordo com a literatura publicada nos últimos 20 anos.

Is there any correlation between alpha-synuclein levels in tears and retinal layer thickness in Parkinson's disease?

PURPOSE: To determine the total alpha-synuclein (αSyn) reflex tears and its association with retinal layers thickness in Parkinson's disease (PD). METHODS: Fifty-two eyes of 26 PD subjects and 52 eyes of age-and sex-matched healthy controls were included. Total αSyn in reflex tears was quantified using a human total αSyn enzyme-linked immunosorbent assay (ELISA) kit. The retinal thickness was evaluated with spectral-domain optical coherence tomography. The Movement Disorder Society-Unified Parkinsons Disease Rating Scale (MDS-UPDRS), Non-Motor Symptoms Scale (NMSS), and Montreal Cognitive Assessment (MoCA) were used to assess motor, non-motor, and cognition. RESULTS: In PD, total αSyn levels were increased compared to control subjects [1.76pg/mL (IQR 1.74-1.80) vs 1.73pg/mL (IQR 1.70-1.77), p < 0.004]. The nerve fiber layer, ganglion cell layer, internal plexiform layer, inner nuclear layer, and outer nuclear layer were thinner in PD in comparison with controls (p < 0.05). The outer plexiform layer and retinal pigment epithelium were thicker in PD (p < 0.05). The total αSyn levels positively correlated with the central volume of the inner nuclear layer (r = 0.357, p = 0.009). CONCLUSION: Total αSyn reflex tear levels were increased in subjects with PD compared to controls. PD patients showed significant thinning of the inner retinal layers and thickening of outer retinal layers in comparison with controls. Total αSyn levels positively correlate with the central volume of the inner nuclear layer in PD. The combination of these biomarkers might have a possible role as a diagnostic tool in PD subjects.

Revisiting the role of 3-nitrotyrosine residues in the formation of alpha-synuclein oligomers and fibrils.

Nitration of tyrosine residues in alpha-synuclein (a-syn) has been detected in different synucleinopathies, including Parkinson's disease. The potential role of 3-nitrotyrosine formation in a-syn, as an oxidative post-translational modification, is still elusive. In this work, we generated well-characterized tyrosine nitrated a-syn monomers and studied their capability to form oligomers and fibrils. We constructed tyrosine to phenylalanine mutants, containing a single tyrosine residue, a-syn mutant Y(125/133/136)F and Y(39/125/133)F) and assessed the impact in a-syn biophysical properties. Nitrated wild-type a-syn and the Y-F mutants, with one 3-nitrotyrosine residue in either the protein's N-terminal or C-terminal region, showed inhibition of fibril formation but retained the capacity of oligomer formation. The inhibition of a-syn fibrillation occurs even when an important amount of unmodified a-syn is still present. We characterized oligomers from both nitrated and non-nitrated forms of the wild-type protein and the mutant forms obtained. Our results indicate that the formation of 3-nitrotyrosine in a-syn could induce an off-pathway oligomer formation which may have an important impact in the development of synucleinopathies.

Alpha-synuclein dynamics bridge Type-I Interferon response and SARS-CoV-2 replication in peripheral cells

BACKGROUND: Increasing evidence suggests a double-faceted role of alpha-synuclein (α-syn) following infection by a variety of viruses, including SARS-CoV-2. Although α-syn accumulation is known to contribute to cell toxicity and the development and/or exacerbation of neuropathological manifestations, it is also a key to sustaining anti-viral innate immunity. Consistently with α-syn aggregation as a hallmark of Parkinson's disease, most studies investigating the biological function of α-syn focused on neural cells, while reports on the role of α-syn in periphery are limited, especially in SARS-CoV-2 infection. RESULTS: Results herein obtained by real time qPCR, immunofluorescence and western blot indicate that α-syn upregulation in peripheral cells occurs as a Type-I Interferon (IFN)-related response against SARS-CoV-2 infection. Noteworthy, this effect mostly involves α-syn multimers, and the dynamic α-syn multimer:monomer ratio. Administration of excess α-syn monomers promoted SARS-CoV-2 replication along with downregulation of IFN-Stimulated Genes (ISGs) in epithelial lung cells, which was associated with reduced α-syn multimers and α-syn multimer:monomer ratio. These effects were prevented by combined administration of IFN-ß, which hindered virus replication and upregulated ISGs, meanwhile increasing both α-syn multimers and α-syn multimer:monomer ratio in the absence of cell toxicity. Finally, in endothelial cells displaying abortive SARS-CoV-2 replication, α-syn multimers, and multimer:monomer ratio were not reduced following exposure to the virus and exogenous α-syn, suggesting that only productive viral infection impairs α-syn multimerization and multimer:monomer equilibrium. CONCLUSIONS: Our study provides novel insights into the biology of α-syn, showing that its dynamic conformations are implicated in the innate immune response against SARS-CoV-2 infection in peripheral cells. In particular, our results suggest that promotion of non-toxic α-syn multimers likely occurs as a Type-I IFN-related biological response which partakes in the suppression of viral replication. Further studies are needed to replicate our findings in neuronal cells as well as animal models, and to ascertain the nature of such α-syn conformations.

PrP meets alpha-synuclein: Molecular mechanisms and implications for disease.

The discovery of prions has challenged dogmas and has revolutionized our understanding of protein-misfolding diseases. The concept of self-propagation via protein conformational changes, originally discovered for the prion protein (PrP), also applies to other proteins that exhibit similar behavior, such as alpha-synuclein (aSyn), a central player in Parkinson's disease and in other synucleinopathies. aSyn pathology appears to spread from one cell to another during disease progression, and involves the misfolding and aggregation of aSyn. How the transfer of aSyn between cells occurs is still being studied, but one important hypothesis involves receptor-mediated transport. Interestingly, recent studies indicate that the cellular prion protein (PrPC ) may play a crucial role in this process. PrPC has been shown to act as a receptor/sensor for protein aggregates in different neurodegenerative disorders, including Alzheimer's disease and amyotrophic lateral sclerosis. Here, we provide a comprehensive overview of the current state of knowledge regarding the interaction between aSyn and PrPC and discuss its role in synucleinopathies. We examine the properties of PrP and aSyn, including their structure, function, and aggregation. Additionally, we discuss the current understanding of PrPC 's role as a receptor/sensor for aSyn aggregates and identify remaining unanswered questions in this area of research. Ultimately, we posit that exploring the interaction between aSyn and PrPC may offer potential treatment options for synucleinopathies.

Meta-analysis to Implement Alpha-Synuclein in Extracellular Vesicles as a Potential Biomarker for Parkinsons Disease.

Background: In Parkinson's disease (PD), exosomes carry α-synuclein (α-syn), a fibrillar protein aggregates with potential value as a biomarker. Objective: Evidence on blood levels of exosomal α-syn in PD patients and controls was reviewed for their consistency. Methods: Thirty-six studies on exosomal α-syn concentrations in PD were identified in a systematic literature search and meta-analysis. Results: Both raw and ratio-adjusted blood exosomal α-syn levels were consistently higher in PD patients than in controls. The standardized mean difference (SMD) was 1.54 (0.18-2.90, CI95%, p < 0.01) and 1.53 (0.23-2.83, CI95%, p < 0.01), respectively. Conclusion: Our results suggest that exosomal α-syn concentrations could be a useful biomarker for PD.

Meta-analysis to Implement Alpha-Synuclein in Extracellular Vesicles as a Potential Biomarker for Parkinson's Disease

ABSTRACT Background: In Parkinson's disease (PD), exosomes carry α-synuclein (α-syn), a fibrillar protein aggregates with potential value as a biomarker. Objective: Evidence on blood levels of exosomal α-syn in PD patients and controls was reviewed for their consistency. Methods: Thirty-six studies on exosomal α-syn concentrations in PD were identified in a systematic literature search and meta-analysis. Results: Both raw and ratio-adjusted blood exosomal α-syn levels were consistently higher in PD patients than in controls. The standardized mean difference (SMD) was 1.54 (0.18-2.90, CI95%, p < 0.01) and 1.53 (0.23-2.83, CI95%, p < 0.01), respectively. Conclusion: Our results suggest that exosomal α-syn concentrations could be a useful biomarker for PD.

Gut-microbiome-brain axis: the crosstalk between the vagus nerve, alpha-synuclein and the brain in Parkinson's disease.

This critical review of the literature shows that there is a close link between the microbiome, the gut, and the brain in Parkinson's disease. The vagus nerve, the main component of the parasympathetic nervous system, is involved in the regulation of immune response, digestion, heart rate, and control of mood. It can detect microbiota metabolites through its afferents, transferring this gut information to the central nervous system. Preclinical and clinical studies have shown the important role played by the gut microbiome and gut-related factors in disease development and progression, as well as treatment responses. These findings suggest that the gut microbiome may be a valuable target for new therapeutic strategies for Parkinson's disease. More studies are needed to better understand the underlying biology and how this axis can be modulated for the patient's benefit.

Three-dimensional neuroimmune co-culture system for modeling Parkinson's disease microenvironmentsin vitro.

Parkinson's disease (PD) is a complex and multifaceted neurodegenerative disorder that results from multiple environmental factors and multicellular interactions. Although several PD neuropathologies have been identified and described, the thorough understanding of PD pathophysiology and research has been largely limited by the absence of reliablein vitromodels that truly recapitulate PD microenvironments. Here, we propose a neuroimmune co-culture system that models PD neuropathologies by combining relevant multicellular interactions with environments that mimic the brain. This system is composed of: (i) 3D bioprinted cultures of mature human dopaminergic (DA) neurons grown on extracellular matrix (ECM)-derived scaffolds doped with electroconductive nanostructures, and (ii) a direct co-culture of human astrocytes and differentiated monocytes that models neuroinflammatory responses. When co-cultured in a transwell format, these two compartments recreate relevant multicellular environments that model PD pathologies after exposure to the neurotoxin A53Tα-synuclein. With immunofluorescent staining and gene expression analyses, we show that functional and mature DA 3D networks are generated within our ECM-derived scaffolds with superior performance to standard 2D cultures. Moreover, by analyzing cytokine secretion, cell surface markers, and gene expression, we define a human monocyte differentiation scheme that allows the appearance of both monocyte-derived macrophages and dendritic cell phenotypes, as well as their optimal co-culture ratios with human astrocytes to recreate synergistic neuroinflammatory responses. We show that the combined response of both compartments to A53Tα-synuclein stimulates the formation of intracellularα-synuclein aggregates, induces progressive mitochondrial dysfunction and reactive oxygen species production, downregulates the expression of synaptic, DA, and mitophagy-related genes, and promotes the initiation of apoptotic processes within the DA networks. Most importantly, these intracellular pathologies were comparable or superior to those generated with a rotenone-stimulated 2D control that represents the current standard forin vitroPD models and showed increased resilience towards these neurotoxic insults, allowing the study of disease progression over longer time periods than current models. Taken together, these results position the proposed model as a superior alternative to current 2D models for generating PD-related pathologiesin vitro.

JM-20, a Benzodiazepine-Dihydropyridine Hybrid Molecule, Inhibits the Formation of Alpha-Synuclein-Aggregated Species.

Studies showed that JM-20, a benzodiazepine-dihydropyridine hybrid molecule, protects against rotenone and 6-hydroxydopamine neurotoxicity. However, its protective effects against cytotoxicity induced by endogenous neurotoxins involved in Parkinson's disease (PD) pathogenesis have never been investigated. In this study, we evaluated the ability of JM-20 to inhibit alpha-synuclein (aSyn) aggregation. We also evaluated the interactions of JM-20 with aSyn by molecular docking and molecular dynamics and assessed the protective effect of JM-20 against aminochrome cytotoxicity. We demonstrated that JM-20 induced the formation of heterogeneous amyloid fibrils, which were innocuous to primary cultures of mesencephalic cells. Moreover, JM-20 reduced the average size of aSyn positive inclusions in H4 cells transfected with SynT wild-type and synphilin-1-V5, but not in HEK cells transfected with synphilin-1-GFP. In silico studies showed the interaction between JM-20 and the aSyn-binding site. Additionally, we showed that JM-20 protects SH-SY5Y cells against aminochrome cytotoxicity. These results reinforce the potential of JM-20 as a neuroprotective compound for PD and suggest aSyn as a molecular target for JM-20.

Extracellular Alpha-Synuclein: Mechanisms for Glial Cell Internalization and Activation.

Alpha-synuclein (α-syn) is a small protein composed of 140 amino acids and belongs to the group of intrinsically disordered proteins. It is a soluble protein that is highly expressed in neurons and expressed at low levels in glial cells. The monomeric protein aggregation process induces the formation of oligomeric intermediates and proceeds towards fibrillar species. These α-syn conformational species have been detected in the extracellular space and mediate consequences on surrounding neurons and glial cells. In particular, higher-ordered α-syn aggregates are involved in microglial and oligodendrocyte activation, as well as in the induction of astrogliosis. These phenomena lead to mitochondrial dysfunction, reactive oxygen and nitrogen species formation, and the induction of an inflammatory response, associated with neuronal cell death. Several receptors participate in cell activation and/or in the uptake of α-syn, which can vary depending on the α-syn aggregated state and cell types. The receptors involved in this process are of outstanding relevance because they may constitute potential therapeutic targets for the treatment of PD and related synucleinopathies. This review article focuses on the mechanism associated with extracellular α-syn uptake in glial cells and the consequent glial cell activation that contributes to the neuronal death associated with synucleinopathies.

BAG2 prevents Tau hyperphosphorylation and increases p62/SQSTM1 in cell models of neurodegeneration.

BACKGROUND: Protein aggregates are pathological hallmarks of many neurodegenerative diseases, however the physiopathological role of these aggregates is not fully understood. Protein quality control has a pivotal role for protein homeostasis and depends on specific chaperones. The co-chaperone BAG2 can target phosphorylated Tau for degradation by an ubiquitin-independent pathway, although its possible role in autophagy was not yet elucidated. In view of this, the aim of the present study was to investigate the association among protein aggregation, autophagy and BAG2 levels in cultured cells from hippocampus and locus coeruleus as well as in SH-SY5Y cell line upon different protein aggregation scenarios induced by rotenone, which is a flavonoid used as pesticide and triggers neurodegeneration. METHODS AND RESULTS: The present study showed that rotenone exposure at 0.3 nM for 48 h impaired autophagy prior to Tau phosphorylation at Ser199/202 in hippocampus but not in locus coeruleus cells, suggesting that distinct neuron cells respond differently to rotenone toxicity. Rotenone induced Tau phosphorylation at Ser199/202, together with a decrease in the endogenous BAG2 protein levels in SH-SY5Y and hippocampus cell culture, which indicates that rotenone and Tau hyperphosphorylation can affect this co-chaperone. Finally, it has been shown that BAG2 overexpression, increased p62/SQSTM1 levels in cells from hippocampus and locus coeruleus, stimulated LC3II recycling as well as prevented the raise of phosphorylated Tau at Ser199/202 in hippocampus. CONCLUSIONS: Results demonstrate a possible role for BAG2 in degradation pathways of specific substrates and its importance for the study of cellular aspects of neurodegenerative diseases.

Comparing the copper binding features of alpha and beta synucleins.

Amyloid aggregation of α-synuclein (AS) is one of the hallmarks of Parkinson's disease (PD). Copper ions specifically bind at the N-terminus of AS, accelerating protein aggregation. Its protein homolog ß-synuclein (BS) is also a copper binding protein, but it inhibits AS aggregation. Here, a comparative spectroscopic study of the Cu2+ binding properties of AS and BS has been performed, using electronic absorption, circular dichroism (CD) and electronic paramagnetic resonance (EPR). Our comparative spectroscopic study reveals striking similarities between the Cu2+ binding features of the two proteins. The Cu2+ binding site at the N-terminal group of BS protein, modeled by the BS (1-15) fragment is identical to that of AS; however, its rate of reduction is three times faster as compared to the AS site, consistent with BS having an additional Met residue in its Met1-Xn-Met5-Xn-Met10 motif. The latter is also evident in the cyclic voltammetry studies of the Cu-BS complex. On the other hand, the Cu2+ binding features of the His site in both proteins, as modeled by AS(45-55) and BS(60-70), are identical, indicating that the shift in the His position does not affect its coordination features. Finally, replacement of Glu46 by Ala does not alter Cu2+ binding to the His site, suggesting that the familial PD E46K mutation would not impact copper-induced aggregation. While further studies of the redox activity of copper bound to His50 in AS are required to understand the role of this site in metal-mediated aggregation, our study contributes to a better understanding of the bioinorganic chemistry of PD.

Protein Misfolding and Aggregation: The Relatedness between Parkinson's Disease and Hepatic Endoplasmic Reticulum Storage Disorders.

Dysfunction of cellular homeostasis can lead to misfolding of proteins thus acquiring conformations prone to polymerization into pathological aggregates. This process is associated with several disorders, including neurodegenerative diseases, such as Parkinson's disease (PD), and endoplasmic reticulum storage disorders (ERSDs), like alpha-1-antitrypsin deficiency (AATD) and hereditary hypofibrinogenemia with hepatic storage (HHHS). Given the shared pathophysiological mechanisms involved in such conditions, it is necessary to deepen our understanding of the basic principles of misfolding and aggregation akin to these diseases which, although heterogeneous in symptomatology, present similarities that could lead to potential mutual treatments. Here, we review: (i) the pathological bases leading to misfolding and aggregation of proteins involved in PD, AATD, and HHHS: alpha-synuclein, alpha-1-antitrypsin, and fibrinogen, respectively, (ii) the evidence linking each protein aggregation to the stress mechanisms occurring in the endoplasmic reticulum (ER) of each pathology, (iii) a comparison of the mechanisms related to dysfunction of proteostasis and regulation of homeostasis between the diseases (such as the unfolded protein response and/or autophagy), (iv) and clinical perspectives regarding possible common treatments focused on improving the defensive responses to protein aggregation for diseases as different as PD, and ERSDs.