Objective Evidence of abnormal alpha-synuclein (alpha-Syn) deposition in the brain is required for definitive diagnosis of synucleinopathies, which remains challenging. The seed amplification assay (SAA) is an innovative technique that can detect the seeding activity of misfolded alpha-Syn, enabling the amplification and detection of minute quantities of pathogenic alpha-Syn aggregates. This study aimed to evaluate oral mucosa alpha-Syn SAA as possible diagnostic and prodromal biomarkers for synucleinopathies. Methods A total of 107 Parkinson's disease (PD) patients, 99 multiple system atrophy (MSA) patients, 33 patients with isolated rapid eye movement sleep behavior disorder (iRBD) and 103 healthy controls (HC) were included. The SAA was applied to detect the seeding activity of alpha-Syn from oral mucosa. A combination of morphological, biochemical, and biophysical methods was also used to analyze the fibrils generated from the oral mucosa alpha-Syn SAA. Results Structured illumination microscopy images revealed the increased alpha-Syn species in oral mucosa of PD, MSA, and iRBD patients than in HCs. Oral mucosa alpha-Syn SAA distinguished patients with PD from HC with 67.3% sensitivity and 90.3% specificity. Oral mucosa was alpha-Syn SAA positive in 53.5% MSA patients and 63.6% iRBD patients. Furthermore, the alpha-Syn fibrils generated from MSA demonstrated greater resistance to proteinase K digestion and exhibited stronger cytotoxicity compared to those from PD patients. Conclusion Oral mucosa alpha-Syn seeding activity may serve as novel non-invasive diagnostic and prodromal biomarkers for synucleinopathies. The alpha-Syn aggregates amplified from the oral mucosa of PD and MSA exhibited distinct biochemical and biophysical properties. (c) 2024 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.

AbstractObjectiveIsolated REM sleep behavior disorder (iRBD) is considered as the strongest predictor of Parkinson's disease (PD). Reliable and accurate biomarkers for iRBD detection and the prediction of phenoconversion are in urgent need. This study aimed to investigate whether α‐Synuclein (α‐Syn) species in plasma neuron‐derived extracellular vesicles (NDEVs) could differentiate between iRBD patients and healthy controls (HCs).MethodsNanoscale flow cytometry was used to detect α‐Syn‐containing NDEVs in plasma.ResultsA total of 54 iRBD patients and 53 HCs were recruited. The concentrations of total α‐Syn, α‐Syn aggregates, and phosphorylated α‐Syn at Ser129 (pS129)‐containing NDEVs in plasma of iRBD individuals were significantly higher than those in HCs (p < 0.0001 for all). In distinguishing between iRBD and HCs, the area under the receiver operating characteristic (ROC) curve (AUC) for an integrative model incorporating the levels of α‐Syn, pS129, and α‐Syn aggregate‐containing NDEVs in plasma was 0.965. This model achieved a sensitivity of 94.3% and a specificity of 88.9%. In iRBD group, the concentrations of α‐Syn aggregate‐containing NDEVs exhibited a negative correlation with Sniffin’ Sticks olfactory scores (r = −0.351, p = 0.039). Smokers with iRBD exhibited lower levels of α‐Syn aggregates and pS129‐containing NDEVs in plasma compared to nonsmokers (pα‐Syn aggregates = 0.014; ppS129 = 0.003).InterpretationThe current study demonstrated that the levels of total α‐Syn, α‐Syn aggregates, and pS129‐containing NDEVs in the plasma of individuals with iRBD were significantly higher compared to HCs. The levels of α‐Syn species‐containing NDEVs in plasma may serve as biomarkers for iRBD.

BackgroundAdaptive immune dysfunction may play a crucial role in Parkinson's disease (PD) development. Isolated rapid eye movement sleep behavior disorder (iRBD) represents the prodromal stage of synucleinopathies, including PD. Elucidating the peripheral adaptive immune system is crucial in iRBD, but current knowledge remains limited.ObjectiveThis study aimed to characterize peripheral lymphocyte profiles in iRBD patients compared with healthy control subjects (HCs).MethodsThis cross-sectional study recruited polysomnography-confirmed iRBD patients and age- and sex-matched HCs. Venous blood was collected from each participant. Flow cytometry was used to evaluate surface markers and intracellular cytokine production in peripheral blood mononuclear cells.ResultsForty-four iRBD patients and 36 HCs were included. Compared with HCs, patients with iRBD exhibited significant decreases in absolute counts of total lymphocytes and CD3+ T cells. In terms of T cell subsets, iRBD patients showed higher frequencies and counts of proinflammatory T helper 1 cells and INF-gamma+ CD8+ T cells, along with lower frequencies and counts of anti-inflammatory T helper 2 cells. A significant increase in the frequency of central memory T cells in CD8+ T cells was also observed in iRBD. Regarding B cells, iRBD patients demonstrated reduced frequencies and counts of double-negative memory B cells compared with control subjects.ConclusionsThis study demonstrated alterations in the peripheral adaptive immune system in iRBD, specifically in CD4+ and INF-gamma+ CD8+ T cell subsets. An overall shift toward a proinflammatory state of adaptive immunity was already evident in iRBD. These observations might provide insights into the optimal timing for initiating immune interventions in PD. (c) 2024 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.

Early and precise diagnosis of α-synucleinopathies is challenging but critical. In this study, we developed a molecular beacon-based assay to evaluate microRNA-containing extracellular vesicles (EVs) in plasma. We recruited 1203 participants including healthy controls (HCs) and patients with isolated REM sleep behavior disorder (iRBD), α-synucleinopathies, or non-α-synucleinopathies from eight centers across China. Plasma miR-44438-containing EV levels were significantly increased in α-synucleinopathies, including those in the prodromal stage (e.g., iRBD), compared to both non-α-synucleinopathy patients and HCs. However, there are no significant differences between Parkinson's disease (PD) and multiple system atrophy. The miR-44438-containing EV levels negatively correlated with age and the Hoehn and Yahr stage of PD patients, suggesting a potential association with disease progression. Furthermore, a longitudinal analysis over 16.3 months demonstrated a significant decline in miR-44438-containing EV levels in patients with PD. These results highlight the potential of plasma miR-44438-containing EV as a biomarker for early detection and progress monitoring of α-synucleinopathies.

BACKGROUND AND PURPOSE:Alpha-synuclein seed amplification assays (α-syn SAAs) are promising diagnostic methods for Parkinson's disease (PD) and other synucleinopathies. However, there is limited consensus regarding the diagnostic and differential diagnostic performance of α-syn SAAs on biofluids and peripheral tissues. METHODS:A comprehensive research was performed in PubMed, Web of Science, Embase and Cochrane Library. Meta-analysis was performed using a random-effects model. A network meta-analysis based on an ANOVA model was conducted to compare the relative accuracy of α-syn SAAs with different specimens. RESULTS:The pooled sensitivity and specificity of α-syn SAAs in distinguishing PD from healthy controls or non-neurodegenerative neurological controls were 0.91 (95% confidence interval [CI] 0.89-0.92) and 0.95 (95% CI 0.94-0.96) for cerebrospinal fluid (CSF); 0.91 (95% CI 0.86-0.94) and 0.92 (95% CI 0.87-0.95) for skin; 0.80 (95% CI 0.66-0.89) and 0.87 (95% CI 0.69-0.96) for submandibular gland; 0.44 (95% CI 0.30-0.59) and 0.92 (95% CI 0.79-0.98) for gastrointestinal tract; 0.79 (95% CI 0.70-0.86) and 0.88 (95% CI 0.77-0.95) for saliva; and 0.51 (95% CI 0.39-0.62) and 0.91 (95% CI 0.84-0.96) for olfactory mucosa (OM). The pooled sensitivity and specificity were 0.91 (95% CI 0.89-0.93) and 0.50 (95% CI 0.44-0.55) for CSF, 0.92 (95% CI 0.83-0.97) and 0.22 (95% CI 0.06-0.48) for skin, and 0.55 (95% CI 0.42-0.68) and 0.50 (95% CI 0.35-0.65) for OM in distinguishing PD from multiple system atrophy. The pooled sensitivity and specificity were 0.92 (95% CI 0.89-0.94) and 0.84 (95% CI 0.73-0.91) for CSF, 0.92 (95% CI 0.83-0.97) and 0.88 (95% CI 0.64-0.99) for skin and 0.63 (95% CI 0.52-0.73) and 0.86 (95% CI 0.64-0.97) for OM in distinguishing PD from progressive supranuclear palsy. The pooled sensitivity and specificity were 0.94 (95% CI 0.90-0.97) and 0.95 (95% CI 0.77-1.00) for CSF and 0.94 (95% CI 0.84-0.99) and 0.86 (95% CI 0.42-1.00) for skin in distinguishing PD from corticobasal degeneration. CONCLUSIONS:α-Synuclein SAAs of CSF, skin, saliva, submandibular gland, gastrointestinal tract and OM are promising diagnostic assays for PD, with CSF and skin α-syn SAAs demonstrating higher diagnostic performance.

BackgroundCortical amyloid deposition is a common observation in Parkinson’s disease dementia (PDD) patients. Aβ1-42 is linked to a more rapid progression of dementia. Platelets, which degranulate upon activation, are a primary source of Aβ. It has been repeatedly reported that peripheral extracellular vesicles (EVs) can partially reach the central nervous system. Thus, we speculate that activated platelet-derived Aβ1-42-containing EVs (PEV-Aβ1-42) play a crucial role in the cognitive decline of PD patients.MethodsThe study included 189 participants: 66 with non-dementia PD, 73 with PDD, and 50 healthy controls. All participants underwent blood collection and clinical assessments. Twenty PD patients underwent re-examination and repeated blood collection 14 months later. A nano-scale flow cytometry assay was used to detect PEVs and PEV-Aβ1-42 using fluorescence-labeled CD62P and Aβ1-42 antibodies.ResultsParkinson’s disease dementia patients had higher PEV-Aβ1-42 concentrations than healthy controls (p = 0.028). The ratio of PEV-Aβ1-42 to PEV was significantly higher in PDD patients compared to those in non-dementia PD and healthy controls (pPD-ND < 0.001, pHC = 0.041). The PEV-Aβ1-42/PEV ratio appears to influence the odds of developing dementia (OR = 1.76, p < 0.001). The change in the PEV-Aβ1-42/PEV ratio was also correlated with cognitive decline over 14 months (r = −0.447, p < 0.05).ConclusionThe plasma PEV-Aβ1-42/PEV ratio may serve as a diagnostic and prognostic biomarker for PDD patients.