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Background: Peripheral inflammation is an important feature of Parkinson's disease (PD). However, if and how CNS pathology is involved in the peripheral inflammation in PD remains to be fully investigated. Recently, the existence of meningeal lymphatics and its involvement in draining cerebral spinal fluid (CSF) to the cervical lymph node has been discovered. It is known that meningeal lymphatic dysfunction exists in idiopathic PD. The deep cervical lymph node (dCLN) substantially contributes to the drainage of the meningeal lymphatics. In addition, one of the lymphatics draining components, CSF, contains abundant α-synuclein (α-syn), a protein critically involved in PD pathogenesis and neuroinflammation. Thus, we began with exploring the possible structural and functional alterations of the dCLN in a PD mouse model (A53T mice) and investigated the role of pathological α-syn in peripheral inflammation and its potential underlying molecular mechanisms. Methods: In this study, the transgenic mice (prnp-SNCA*A53T) which specifically overexpressed A53T mutant α-syn in CNS were employed as the PD animal model. Immunofluorescent and Hematoxylin and eosin staining were used to evaluate structure of dCLN. Inflammation in dCLNs as well as in bone-marrow-derived macrophages (BMDMs) was assessed quantitatively by measuring the mRNA and protein levels of typical inflammatory cytokines (including IL-1β, IL-6 and TNF-α). Intra-cisterna magna injection, flow cytometric sorting and electrochemiluminescence immunoassays were applied to investigate the lymphatic drainage of α-syn from the CNS. RNA-seq and Western blot were used to explore how pathological α-syn mediated the inflammation in PD mice. Results: The results unequivocally revealed substantially enlarged dCLNs, along with slow lymphatic flow, and increased inflammation in the dCLNs of A53T mice. Oligomeric α-syn drained from CSF potently activated macrophages in the dCLN via endoplasmic reticulum (ER) stress. Notably, inhibition of ER stress effectively suppressed peripheral inflammation in PD mice. Conclusions: Our findings indicate that lymph node enlargement is closely related to macrophage activation, induced by meningeal lymphatics draining oligomeric α-syn, and contributes to the peripheral inflammation in PD. In addition, ER stress is a potential therapeutic target to ameliorate PD pathogenesis. [ABSTRACT FROM AUTHOR]

Objective: Accumulation of α‐synuclein (α‐syn) in neurons is a prominent feature of Parkinson's disease (PD). Recently, researchers have considered that extracellular vesicles (EVs) may play an important role in protein exportation and propagation, and α‐syn‐containing EVs derived from the central nervous system (CNS) have been detected in peripheral blood. However, mechanistic insights into CNS‐derived EVs have not been well‐described. Methods: Likely neurogenic EVs were purified from the plasma of PD patients and healthy controls using a well‐established immunoprecipitation assay with anti‐L1CAM‐coated beads. A Prnp‐SNCAA53T transgenic PD mouse model was used to evaluate the neuronal pathology induced by PD‐derived L1CAM‐purified EVs. EV‐associated microRNA (miRNA) profiling was used to screen for altered miRNAs in PD‐derived L1CAM‐purified EVs. Results: PD patient‐derived L1CAM‐purified (likely neurogenic) EVs facilitated α‐syn pathology and neuron loss in Prnp‐SNCAA53T transgenic PD mice. The miRNA, novel_miR_44438, was significantly increased in the PD group, which promoted α‐syn accumulation and neuronal degeneration in a dose‐dependent manner. Novel _miR_44438 directly targets NDST1 mRNA and inhibits the function of heparan sulfate, thus preventing exosome biogenesis and α‐syn release from exosomes. Interpretation: Novel_miR_44438 in PD‐derived L1CAM‐purified EVs inhibits the α‐syn efflux from neurons thereby promoting the pathological accumulation and aggregation of α‐syn. ANN NEUROL 2022;92:230–245 [ABSTRACT FROM AUTHOR]

Background: Immune system dysfunction, including higher levels of peripheral monocytes and inflammatory cytokines, is an important feature of Parkinson's disease (PD) pathogenesis, although the mechanism underlying the process remains to be investigated. In the central nervous system, it is well-known that α-synuclein (α-syn), a key protein involved in PD, activates microglia potently, and it is also reported that α-syn exists in the peripheral system, especially in erythrocytes or red blood cells (RBC) at exceedingly high concentration. The current study focused on the possibility that RBC-derived α-syn mediates the sensitization of peripheral monocytes in PD patients.Methods: The hyperactivation of monocytes was assessed quantitatively by measuring mRNA levels of typical inflammatory cytokines (including IL-1β, IL-6 and TNF-α) and protein levels of secreted inflammatory cytokines (including pro-inflammatory cytokines: IL-1β, IL-6, TNF-α, IL-8, IFN-γ, IL-2, and IL-12p70 and anti-inflammatory cytokines: IL-4, IL-10, and IL-13). Western blot, nanoparticle tracking analysis and electron microscopy were used to characterize RBC-derived extracellular vesicles (RBC-EVs). Inhibitors of endocytosis and leucine-rich repeat kinase 2 (LRRK2), another key protein involved in PD, were used to investigate how these two factors mediated the process of monocyte sensitization by RBC-EVs.Results: Increased inflammatory sensitization of monocytes was observed in PD patients and PD model mice. We found that α-syn-containing RBC-EVs isolated from PD model mice or free form oligomeric α-syn induced the inflammatory sensitization of THP-1 cells, and demonstrated that endocytosis was a requirement for this pathophysiological pathway. Furthermore, the hyperactivation of THP-1 cells induced by RBC-EVs was associated with increased LRRK2 production and kinase activity. The phenomenon of inflammatory sensitization of human monocytes and increased LRRK2 were also observed by the treatment of RBC-EVs isolated from PD patients.Conclusions: Our data provided new insight into how hyperactivation of monocytes occurs in PD patients, and identified the central role played by α-syn-containing RBC-EVs in this process. [ABSTRACT FROM AUTHOR]

The authors describe the fabrication of a stationary phase for use in enantioseparation in capillary electrochromatography (CEC). The stationary phase was obtained by covalent immobilization of graphene oxide (GO) on the inner walls of a fused-silica capillary. Methyl-β-cyclodextrin was used as the chiral selector, and the enantiomers were detected by UV absorptiometry. Compared with uncoated fused-silica capillary, obviously improved separation of three tested drug enantiomers was obtained in the GO-polymer coated capillary column (the resolution of Naproxen: 0 → 1.56, Warfarin: 0 → 1.89 and Pranoprofen:0 → 1.51). The effects of buffer pH, type and proportion of organic modifier, concentration of chiral selector and applied voltage were optimized. Molecular modeling was applied to demonstrate the mechanism of enantioseparation, which showed a good agreement with the experimental results. [ABSTRACT FROM AUTHOR]

Rationale: Synucleinopathies, including Parkinson's disease (PD), multiple system atrophy (MSA), and dementia with Lewy bodies (DLB), share a distinct pathological feature, that is, a widespread accumulation of α‐synuclein (α‐syn) in the brain. There is a significant clinical unmet need for disease‐modifying treatments for synucleinopathies. Recently, a seaweed‐derived mixture of oligosaccharides sodium oligomannate, GV‐971, was approved for Phase 2 clinical trials for PD. This study aimed to further evaluate the therapeutic effects of GV‐971 on synucleinopathies using cellular and animal models and explore its associated molecular mechanisms. Methods: α‐Syn aggregation was assessed, in vitro and ex vivo, by ThT assay. A dopaminergic neuron cell line, Prnp‐SNCAA53T mice, and brain slices from PD and DLB patients were used to determine the efficacy of GV‐971 in ameliorating α‐syn pathology. Measurements of motor functions, including pole, cylinder, and rotarod tests, were conducted on Prnp‐SNCAA53T mice 4 weeks after intragastric administration of GV‐971 (200 mg day−1 kg−1). Results: GV‐971 effectively prevented α‐syn aggregation and even disassembled pre‐aggregated α‐syn fibrils, in vitro and ex vivo. In addition, GV‐971 was able to rescue α‐syn‐induced neuronal damage and reduced release of extracellular vesicles (EVs), likely via modulating Alix expression. In the Prnp‐SNCAA53T mouse model, when treated at the age of 5 months, GV‐971 significantly decreased α‐syn deposition in the cortex, midbrain, and cerebellum regions, along with ameliorating the motor dysfunctions. Conclusions: Our results indicate that GV‐971, when administered at a relatively early stage of the disease process, significantly reduced α‐syn accumulation and aggregation in Prnp‐SNCAA53T mice. Furthermore, GV‐971 corrected α‐syn‐induced inhibition of EVs release in neurons, contributing to neuronal protection. Future studies are needed to further assess GV‐971 as a promising disease‐modifying therapy for PD and other synucleinopathies. [ABSTRACT FROM AUTHOR]

Background: Progressive spreading of α‐synuclein via gut‐brain axis has been hypothesized in the pathogenesis of Parkinson's disease (PD). However, the source of seeding‐capable α‐synuclein in the gastrointestinal tract (GIT) has not been fully investigated. Additionally, the mechanism by which the GIT microbiome contributes to PD pathogenesis remains to be characterized. Objectives: We aimed to investigate whether blood‐derived α‐synuclein might contribute to PD pathology via a gut‐driven pathway and involve GIT microbiota. Methods: The GIT expression of α‐synuclein and the transmission of extracellular vesicles (EVs) derived from erythrocytes/red blood cells (RBCs), with their cargo α‐synuclein, to the GIT were explored with various methods, including radioactive labeling of RBC‐EVs and direct analysis of the transfer of α‐synuclein protein. The potential role of microbiota on the EVs transmission was further investigated by administering butyrate, the short‐chain fatty acids produced by gut microbiota and studying mice with different α‐synuclein genotypes. Results: This study demonstrated that RBC‐EVs can effectively transport α‐synuclein to the GIT in a region‐dependent manner, along with variations closely associated with regional differences in the expression of gut‐vascular barrier markers. The investigation further revealed that the infiltration of α‐synuclein into the GIT was influenced significantly by butyrate and α‐synuclein genotypes, which may also affect the GIT microbiome directly. Conclusion: By demonstrating the transportation of α‐synuclein through RBC‐EVs to the GIT, and its potential association with gut‐vascular barrier markers and gut microbiome, this work highlights a potential mechanism by which RBC α‐synuclein may impact PD initiation and/or progression. © 2023 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society. [ABSTRACT FROM AUTHOR]

Peripheral biomarkers indicative of brain pathology are critically needed for early detection of Parkinson's disease (PD). In this study, using NanoString and digital PCR technologies, we began by screening for alterations in genes associated with PD or atypical Parkinsonism in erythrocytes of PD patients, in which PD-related changes have been reported, and which contain ~ 99% of blood α-synuclein. Erythrocytic CHCHD2 mRNA was significantly reduced even at the early stages of the disease. A significant reduction in protein and/or mRNA expression of CHCHD2 was confirmed in PD brains collected at autopsy as well as in the brains of a PD animal model overexpressing α-synuclein, in addition to seeing a reduction of CHCHD2 in erythrocytes of the same animals. Overexpression of α-synuclein in cellular models of PD also resulted in reduced CHCHD2, via mechanisms likely involving altered subcellular localization of p300 histone acetyltransferase. Finally, the utility of reduced CHCHD2 mRNA as a biomarker for detecting PD, including early-stage PD, was validated in a larger cohort of 205 PD patients and 135 normal controls, with a receiver operating characteristic analysis demonstrating > 80% sensitivity and specificity. [ABSTRACT FROM AUTHOR]

Parkinson's disease is a neurodegenerative disorder characterized by the transmission and accumulation of toxic species of α-synuclein (α-syn). Extracellular vesicles (EVs) are believed to play a vital role in the spread of toxic α-syn species. Recently, peripheral α-syn pathology has been investigated, but little attention has been devoted to erythrocytes, which contain abundant α-syn. In this study, we first demonstrated that erythrocyte-derived EVs isolated from Parkinson's disease patients carried elevated levels of oligomeric α-syn, compared to those from healthy controls. Moreover, human erythrocyte-derived EVs, when injected into peripheral blood in a mouse model of Parkinson's disease, were found to readily cross the blood-brain barrier (BBB). These EVs accumulated in astrocyte endfeet, a component of the BBB, where they impaired glutamate uptake, likely via interaction between excitatory amino acid transporter 2 (EAAT2) and oligomeric α-syn. These data suggest that erythrocyte-derived EVs and the oligomeric α-syn carried in them may play critical roles in the progression or even initiation of Parkinson's disease. Additionally, the mechanisms involved are attributable at least in part to dysfunction of astrocytes induced by these EVs. These observations provide new insight into the understanding of the mechanisms involved in Parkinson's disease. [ABSTRACT FROM AUTHOR]

Several chiral reagents including cyclodextrins (CDs) and derivatives, crown ethers, proteins, chiral surfactants, and polymers have been involved in dual-selector systems for enantioseparation of a series of compounds by capillary electrophoresis (CE). In this paper, chondroitin sulfate D-based dual-selector system (CSD/CM-β-CD) was firstly established and investigated for the enantioseparation of six basic racemic drugs in capillary electrophoresis. Compared to the single-selector systems, synergistic effect and significantly improved separations for all tested analytes were observed in CSD/CM-β-CD system. The effect of several parameters, such as buffer pH, chiral selector concentration, and applied voltage, was systematically optimized. Meanwhile, to investigate the possible chiral recognition mechanisms in CSD/CM-β-CD synergistic system, we tried to apply the molecular docking method to simulate the host-guest binding procedures of the polysaccharide-based dual system for the first time. The difference in binding free energy was found to correspond to the chiral selectivity factor. The existence of CSD-CM-β-CD complex may give rise to a higher discriminatory ability against the enantiomers, indicating the synergistic effect in CSD/CM-β-CD system.ᅟ [ABSTRACT FROM AUTHOR]

Recently,ionic liquids (ILs) functionalized cyclodextrins (CDs) have attracted more and more attention in the fields of enantioseparation. In this study, a novel IL amino triazolium functionalized β-CD derivative, mono-6-deoxy-6-(4-amino-1,2,4-triazolium)-β-cyclodextrin chloride (4-ATMCDCl), was synthesized for the first time and managed to separate dansyl amino acids and naproxen by capillary electrophoresis (CE). Compared with native β-CD, the new selector exhibited good water solubility and enhanced enantioselectivity. Several crucial parameters such as selector concentration, buffer pH, and applied voltage were systematically investigated. The molecular docking program Autodock was applied to further demonstrate the mechanism of chiral recognition and the enhanced enantioselectivity of 4-ATMCDCl, which showed good agreement with our experimental results. [ABSTRACT FROM AUTHOR]

Medulloblastoma (MB), the most common malignant pediatric brain tumor, is composed of at least four molecular subgroups with distinct clinical characteristics. The sonic hedgehog (SHH) subgroup exhibits the most abundant tumor-associated microglia/macrophages (TAMs) infiltration. SHH-MB patients treated by anti-SHH drugs showed high drug resistance. However, the comprehensive role of TAMs in SHH-MB remains enigma. The aim of this study is to explore the mechanism of TAM activation/polarization in SHH-MB and discover a potential immunotherapeutic target to reduce drug resistance. We first analyzed expression profiles of immuno-microenvironment (IME) in four subgroups of 48 MB tumors using NanoString PanCancer IO360 panel and found TAMs were the major component of IME in SHH-MBs. We further distinguished M1/M2-like TAMs in tumors and found M2-like macrophages, rather than microglia, were enriched in SHH-MBs. In transgenic SHH-MB mice, these TAMs had close relationship with tumor progression. Polarization of the TAMs could be induced by MB-derived exosomes in vitro. We then screened SHH MB-derived exosomal miRNAs and their target genes using RNA sequencing and luciferase assay to clarify their roles in regulating TAM polarization. We found down-regulated let-7i-5p and miR-221-3p can induce M2-like polarization of TAMs via upregulating peroxisome proliferator activated receptor gamma (PPARγ). Finally, we demonstrated the PPARγ antagonist efficiently improved the antitumor activity of SMO inhibitor in vivo, which may be related to inhibition of M2-like TAMs. Our findings suggest a potential therapeutic strategy for SHH-MB by targeting tumor-supportive M2-like TAMs to enhance the therapeutic effect of SMO inhibitors. [ABSTRACT FROM AUTHOR]