Α-synuclein Concentrations Research Articles

Unfolded protein response markers Grp78 and eIF2alpha are upregulated with increasing alpha-synuclein levels in Lewy body disease.

Endoplasmic reticulum stress followed by the unfolded protein response is one of the cellular mechanisms contributing to the progression of α-synuclein pathology in Parkinson's disease and other Lewy body diseases. We aimed to investigate the activation of endoplasmic reticulum stress and its correlation with α-synuclein pathology in human post-mortem brain tissue. We analysed brain tissue from 45 subjects-14 symptomatic patients with Lewy body disease, 19 subjects with incidental Lewy body disease, and 12 healthy controls. The analysed brain regions included the medulla, pons, midbrain, striatum, amygdala and entorhinal, temporal, frontal and occipital cortex. We analysed activation of endoplasmic reticulum stress via levels of the unfolded protein response-related proteins (Grp78, eIF2α) and endoplasmic reticulum stress-regulating neurotrophic factors (MANF, CDNF). We showed that regional levels of two endoplasmic reticulum-localised neurotrophic factors, MANF and CDNF, did not change in response to accumulating α-synuclein pathology. The concentration of MANF negatively correlated with age in specific regions. eIF2α was upregulated in the striatum of Lewy body disease patients and correlated with increased α-synuclein levels. We found the upregulation of chaperone Grp78 in the amygdala and nigral dopaminergic neurons of Lewy body disease patients. Grp78 levels in the amygdala strongly correlated with soluble α-synuclein levels. Our data suggest a strong but regionally specific change in Grp78 and eIF2α levels, which positively correlates with soluble α-synuclein levels. Additionally, MANF levels decreased in dopaminergic neurons in the substantia nigra. Our research suggests that endoplasmic reticulum stress activation is not associated with Lewy pathology but rather with soluble α-synuclein concentration and disease progression.

Salivary α-Synuclein as a Candidate Biomarker of Parkinsonism in 22q11.2 Deletion Syndrome.

22q11.2 deletion syndrome (22q11.2DS) has been linked to an increased risk of early-onset Parkinson's disease. However, the pathophysiological mechanisms underlying parkinsonism remain poorly understood. The objective is to investigate salivary total α-synuclein levels in 22q11.2DS patients with and without parkinsonian motor signs. This cross-sectional study included 10 patients with 22q11.2DS with parkinsonism (Park+), ten 22q11.2DS patients without parkinsonism (Park-), and 10 age and sex-comparable healthy subjects (HS). Salivary and serum α-synuclein levels were measured using enzyme-linked immunosorbent assay. Salivary total α-synuclein concentration was significantly lower in Park (+) patients than in Park (-) patients and HS (P = 0.007). In addition, salivary α-synuclein showed good accuracy in discriminating Park (+) from Park (-) patients (area under the curve = 0.86) and correlated with motor severity and cognitive impairment. This exploratory study suggests that the parkinsonian phenotype of 22q11.2DS is associated with a reduced concentration of monomeric α-synuclein in biological fluids.

An innovative transportable immune device for the recognition of α-synuclein using KCC-1-nPr-CS2 modified silver nano-ink: integration of pen-on-paper technology with biosensing toward early-stage diagnosis of Parkinson's disease.

Parkinson's disease (PD), the second most frequent neurodegenerative illness, is a neurological ailment that produces unintentional or uncontrolled body movements, which should be diagnosed in its early stages to hinder the progression. Monitoring the concentration of α-synuclein (α-Syn) in body fluids can be one of the most efficient ways for PD early detection. In this work, a paper-based electrochemical immunosensor was designed for α-Syn bio-assay in human plasma samples based on encapsulation of the biotinylated antibody on novel dendritic fibrous nanosilica ((KCC-1-nPr-CS2)-Ab). For this purpose, a three-electrode system was prepared using stabilization of silver nano-ink on photographic paper. Then, the (KCC-1-NH-CS2)-Ab was immobilized on its surface and used to detect the target antigen (α-Syn). After characterization of the prepared substrate by FE-SEM and EDS, the redox behavior of the biosensor was evaluated using chronoamperometry techniques. Under optimal experimental conditions and using a label-free strategy, the engineered immunosensor showed a linear relationship between peak current and antigen concentration in the linear range from 0.002 to 128 ng mL-1 with the lower limit of quantification of 0.002 ng mL-1. Moreover, this work involves unprecedented use of conductive nano-inks for the manufacture of α-Syn immunosensor, which is aided by the use of a mesoporous silicate dendrimer in encapsulating the α-Syn antibody, thus offering a robust and simple point-of-care device for early PD diagnosis. The ability of the proposed platform to detect small amounts of α-Syn offers a promising approach to developing low-cost, sensitive, and transportable biosensors for Parkinson's disease screening in its early stages.

Lab-made disposable screen-printed electrochemical sensors and immunosensors modified with Pd nanoparticles for Parkinson's disease diagnostics.

Anew conductive ink based on the addition of carbon black to a poly(vinyl alcohol) matrix is developed and investigated for electrochemical sensing and biosensing applications. The produced devices were characterized using morphological and electrochemical techniques and modified with Pd nanoparticles to enhance electrical conductivity and reaction kinetics. With the aid of chemometrics, the parameters for metal deposition were investigated and the sensor was applied tothe determination of Parkinson's disease biomarkers, specifically epinephrine and α-synuclein. A linear behavior was obtained in the range 0.75 to 100μmol L-1 of the neurotransmitter, and the device displayed a limit of detection (LOD) of 0.051μmol L-1. The three-electrode system was then tested using samples of synthetic cerebrospinal fluid. Afterward, the device was modified with specific antibodies to quantify α-synuclein using electrochemical impedance spectroscopy. In phosphate buffer, a linear range was obtained for α-synuclein concentrations from 1.5 to 15μgmL-1, with a calculated LOD of 0.13μgmL-1. The proposed immunosensor was also applied to blood serum samples, and, in this case, the linear range was observed from 6.0 to 100.5μgmL-1 of α-synuclein, with a LOD = 1.3µgmL-1. Both linear curves attend the range for the real diagnosis, demonstrating its potential application to complex matrices.

CSF Synaptic Biomarkers in AT(N)-Based Subgroups of Lewy Body Disease.

Patients with Lewy body disease (LBD) often show a co-occurring Alzheimer disease (AD) pathology. CSF biomarkers allow the detection in vivo of AD-related pathologic hallmarks included in the amyloid-tau-neurodegeneration (AT(N)) classification system. Here, we aimed to investigate whether CSF biomarkers of synaptic and neuroaxonal damage are correlated with the presence of AD copathology in LBD and can be useful to differentiate patients with LBD with different AT(N) profiles. We retrospectively measured CSF levels of AD core biomarkers (Aβ42/40 ratio, phosphorylated tau protein, and total tau protein) and of synaptic (β-synuclein, α-synuclein, synaptosomal-associated protein 25 [SNAP-25], and neurogranin) and neuroaxonal proteins (neurofilament light chain [NfL]) in 28 cognitively unimpaired participants with nondegenerative neurologic conditions and 161 participants with a diagnosis of either LBD or AD (at both mild cognitive impairment, AD-MCI, and dementia stages, AD-dem). We compared CSF biomarker levels in clinical and AT(N)-based subgroups. CSF β-synuclein, α-synuclein, SNAP-25, neurogranin, and NfL levels did not differ between LBD (n = 101, age 67.2 ± 7.8 years, 27.7% females) and controls (age 64.8 ± 8.6 years, 39.3% females) and were increased in AD (AD-MCI: n = 30, AD-dem: n = 30, age 72.3 ± 6.0 years, 63.3% females) compared with both groups (p < 0.001 for all comparisons). In LBD, we found increased levels of synaptic and neuroaxonal degeneration biomarkers in patients with A+T+ (LBD/A+T+) than with A-T- profiles (LBD/A-T-) (p < 0.01 for all), and β-synuclein showed the highest discriminative accuracy between the 2 groups (area under the curve 0.938, 95% CI 0.884-0.991). CSF β-synuclein (p = 0.0021), α-synuclein (p = 0.0099), and SNAP-25 concentrations (p = 0.013) were also higher in LBD/A+T+ than in LBD/A+T- cases, which had synaptic biomarker levels within the normal range. CSF α-synuclein was significantly decreased only in patients with LBD with T- profiles compared with controls (p = 0.0448). Moreover, LBD/A+T+ and AD cases did not differ in any biomarker level. LBD/A+T+ and AD cases showed significantly increased CSF levels of synaptic and neuroaxonal biomarkers compared with LBD/A-T- and control subjects. Patients with LBD and AT(N)-based AD copathology showed, thus, a distinct signature of synaptic dysfunction from other LBD cases. This study provides Class II evidence that CSF levels of β-synuclein, α-synuclein, SNAP-25, neurogranin, and NfL are higher in patients with AD than in patients with LBD.

Toward a Biomarker Panel measured in CNS-originating Extracellular Vesicles for Differential Diagnosis of Parkinson’s Disease and Multiple System Atrophy (S37.005)

Toward a Biomarker Panel measured in CNS-originating Extracellular Vesicles for Differential Diagnosis of Parkinson’s Disease and Multiple System Atrophy (S37.005)

Decreased serum levels of α-synuclein in patients with schizophrenia and their unaffected siblings.

The final common pathway in the etiopathogenesis of schizophrenia is suggested that there is a defect in the presynaptic terminal in dopaminergic transmission, in which α-synuclein has an important role. Peripheral biomarker studies in schizophrenia have become crucial for better diagnoses, early interventions, and personalized therapies. This study aims to compare α-synuclein levels in patients with schizophrenia and their unaffected siblings with healthy controls, as a potential peripheral biomarker for schizophrenia. The quantifications of α-synuclein serum concentrations were conducted by the ELISA method. PANSS and CGI-S were used to analyse the severity of the symptoms of the subjects. Data were analysed by nonparametric tests and the Receiver Operating Curve (ROC) analysis. Sixty-two patients with schizophrenia (mean age: 34,8± 9,9, %64,5 male), their 56 unaffected siblings (mean age: 39,4± 11,5, %55,4 male) and 56 healthy controls (mean age: 36,2± 9,8, %64,3 male) were included. α-synuclein levels were significantly lower in the patient (27,65 (12,61-46,09) pg/ml) and the unaffected sibling groups (24,62 (15,60-57,87) pg/ml) compared with healthy controls (45,58 (11,25-108,30) pg/ml) (p< .001). According to the ROC analysis, the optimal cut-off value for α-synuclein levels in distinguishing the schizophrenia group from the control group was 42.20. The sensitivity of the measurement of serum α-synuclein at this point was 93.5%, and the specificity was 60.7%. Our study demonstrates that decreased levels of serum α-synuclein may be utilized as a possible peripheral biomarker of familial risk for schizophrenia in both patients and their siblings.

Oligomeric α-synuclein and tau aggregates in NDEVs differentiate Parkinson's disease from atypical parkinsonisms

The early differential diagnosis of Parkinson's disease (PD) and atypical Parkinsonian syndromes (APS), including corticobasal degeneration (CBD) and progressive supranuclear palsy (PSP), is challenging because of an overlap of clinical features and the lack of reliable biomarkers. Neural-derived extracellular vesicles (NDEVs) isolated from blood provide a window into the brain's biochemistry and may assist in distinguishing between PD and APS. We verified in a case-control study whether oligomeric α-Synuclein and Tau aggregates isolated from NDEVs could allow the differential diagnosis of these conditions.Blood sampling and clinical data, including disease duration, motor severity, global cognition, and levodopa equivalent daily dose (LEDD), were collected from patients with a diagnosis of either PD (n = 70), PSP (n = 21), or CBD (n = 19). NDEVs were isolated from serum by immunocapture using an antibody against the neuronal surface marker L1CAM; oligomeric α-Synuclein and aggregated Tau were measured by ELISA.NDEVs analyses showed that oligomeric α-Synuclein is significantly augmented in PD compared to APS, whereas Tau aggregates are significantly increased in APS compared to PD (p < 0.0001). ROC analyses showed that these two biomarkers have a “good” power of classification (p < 0.0001 for both proteins), with high sensitivity and specificity, with NDEVs concentration of Tau aggregates and oligomeric α-Synuclein being respectively the best biomarker for PD/PSP and PD/CBD diagnostic differentiation.Logistic and multiple regression analysis confirmed that NDEVs-derived oligomeric α-Synuclein and Tau aggregates differentiate PD from CBD and PSP (p < 0.001). Notably, a positive correlation between NDEVs oligomeric α-Synuclein and disease severity (disease duration, p = 0.023; Modified H&Y, p = 0.015; UPDRS motor scores, p = 0.004) was found in PD patients and, in these same patients, NDEVs Tau aggregates concentration inversely correlated with global cognitive scores (p = 0.043).A minimally invasive blood test measuring the concentration of α-synuclein and Tau aggregates in NDEVs can represent a promising tool to distinguish with high sensitivity and specificity PD from CBD or PSP patients. Optimization and validation of these data will be needed to confirm the diagnostic value of these biomarkers in distinguishing synucleinopathies from taupathies.

The Role of Amyloid-β, Tau, and α-Synuclein Proteins as Putative Blood Biomarkers in Patients with Cerebral Amyloid Angiopathy.

Cerebral amyloid angiopathy (CAA) is a cerebrovascular disorder characterized by the deposition of amyloid-β protein (Aβ) within brain blood vessels that develops in elderly people and Alzheimer's disease (AD) patients. Therefore, the investigation of biomarkers able to differentiate CAA patients from AD patients and healthy controls (HC) is of great interest, in particular in peripheral fluids. The current study aimed to detect the neurodegenerative disease (ND)-related protein (i.e., Aβ1-40, Aβ1-42, tau, and α-synuclein) levels in both red blood cells (RBCs) and plasma of CAA patients and HC, evaluating their role as putative peripheral biomarkers for CAA. For this purpose, the proteins' concentration was quantified in RBCs and plasma by homemade immunoenzymatic assays in an exploratory cohort of 20 CAA patients and 20 HC. The results highlighted a significant increase of Aβ1-40 and α-synuclein concentrations in both RBCs and plasma of CAA patients, while higher Aβ1-42 and t-tau levels were detected only in RBCs of CAA individuals compared to HC. Moreover, Aβ1-42/Aβ1-40 ratio increased in RBCs and decreased in plasma of CAA patients. The role of these proteins as candidate peripheral biomarkers easily measurable with a blood sample in CAA needs to be confirmed in larger studies. In conclusion, we provide evidence concerning the possible use of blood biomarkers for contributing to CAA diagnosis and differentiation from other NDs.

Diagnostic Utility of Cerebrospinal Fluid α-Synuclein in Creutzfeldt-Jakob Disease: A Systematic Review and Meta-Analysis.

Creutzfeldt-Jakob disease (CJD) can be difficult to distinguish clinically from some non-prion neurological diseases. Previous studies have reported markedly increased levels of α-synuclein in cerebrospinal fluid (CSF) of CJD patients, indicating that it is a potential diagnostic biomarker. The aim of this study was to assess the diagnostic power of CSF α-synuclein in discriminating CJD from non-prion disorders. The Ovid MEDLINE, Cochrane, and Embase databases were searched for articles published on or before February 25, 2022, using the search term (prion diseases OR Creutzfeldt-Jakob syndrome) AND (synuclein OR α-synuclein). The difference in CSF α-synuclein levels between CJD and non-prion diseases was calculated using random-effects models (I2 &gt; 50%) or fixed-effects models (I2 &lt; 50%) in terms of standardized mean difference (SMD) and 95% confidence interval (CI). The publication bias was estimated using funnel plots and the Egger's test. Ten studies were included in this study. The concentrations of CSF α-synuclein were significantly higher in CJD patients compared to total non-prion controls (SMD = 1.98, 95% CI 1.60 to 2.36, p &lt; 0.00001), tauopathies (SMD = 1.34, 95% CI 0.99 to 1.68, p &lt; 0.00001), synucleinopathies (SMD = 1.78, 95% CI 1.11 to 2.44, p &lt; 0.00001), or Alzheimer's (SMD = 1.14, 95% CI 0.95 to 1.33, p &lt; 0.00001). CSF α-synuclein could distinguish CJD from non-prion diseases with overall sensitivity of 89% (95% CI 80-95%), specificity of 92% (95% CI 86-95%), and AUC of 0.96 (95% CI: 0.94-0.97). CSF α-synuclein has excellent diagnostic value in discriminating CJD from non-prion neurological diseases. Given the high heterogeneity among the included studies, further studies are needed to confirm its clinical utility.

Measurement of α-Synuclein Dynamics In Vivo Using Microdialysis with a Novel Homogeneous Immunoassay.

Understanding the regulation of α-synuclein release could be important in better understanding Parkinson's disease development, progression, and treatment. Advances in such studies are hindered by technical challenges that limit the ability to monitor α-synuclein concentration in vivo. We developed a novel α-synuclein microdialysis method coupled with a specific and sensitive immunoassay that requires a small sample volume (1 μL). Using this method, basal α-synuclein level was estimated at 254 ± 78 pM in the striatum of freely moving mice. Additionally, we observed that potassium (75 mM) and nicotine (0.5 mg/kg) administration significantly increased α-synuclein in dialysates. These results provide evidence that the methods we report here can be useful to investigate the physiological roles of α-synuclein and support the idea that α-synuclein is secreted to the extracellular space in a neuronal activity-dependent manner.

Dysregulation of a Heme Oxygenase-Synuclein Axis in Parkinson Disease.

α-Synuclein is a key driver of the pathogenesis of Parkinson disease (PD). Heme oxygenase-1 (HO-1), a stress protein that catalyzes the conversion of heme to biliverdin, carbon monoxide and free ferrous iron, is elevated in PD-affected neural tissues and promotes iron deposition and mitochondrial dysfunction in models of the disease, pathways also impacted by α-synuclein. Elevated expression of human HO-1 in astrocytes of GFAP.HMOX1 transgenic mice between 8.5 and 19 months of age elicits a parkinsonian phenotype characterized by nigrostriatal hypodopaminergia, locomotor incoordination and overproduction of neurotoxic native S129-phospho-α-synuclein. Two microRNAs (miRNA) known to regulate α-synuclein, miR-153 and miR-223, are significantly decreased in the basal ganglia of GFAP.HMOX1 mice. Serum concentrations of both miRNAs progressively decline in wild-type (WT) and GFAP.HMOX1 mice between 11 and 18 months of age. Moreover, circulating levels of miR-153 and miR-223 are significantly lower, and erythrocyte α-synuclein concentrations are increased, in GFAP.HMOX1 mice relative to WT values. MiR-153 and miR-223 are similarly decreased in the saliva of PD patients compared to healthy controls. Upregulation of glial HO-1 may promote parkinsonism by suppressing miR-153 and miR-223, which, in turn, enhance production of neurotoxic α-synuclein. The aim of the current review is to explore the link between HO-1, α-synuclein and PD, evaluating evidence derived from our laboratory and others. HO-1, miR-153 and miR-223 and α-synuclein may serve as potential biomarkers and targets for disease-modifying therapy in idiopathic PD.

Asthma amplifies dementia risk: Evidence from CSF biomarkers and cognitive decline.

IntroductionEvidence from epidemiology, neuroimaging, and animal models indicates that asthma adversely affects the brain, but the nature and extent of neuropathophysiological impact remain unclear.MethodsWe tested the hypothesis that asthma is a risk factor for dementia by comparing cognitive performance and cerebrospinal fluid biomarkers of glial activation/neuroinflammation, neurodegeneration, and Alzheimer's disease (AD) pathology in 60 participants with asthma to 315 non‐asthma age‐matched control participants (45–93 years), in a sample enriched for AD risk.ResultsParticipants with severe asthma had higher neurogranin concentrations compared to controls and those with mild asthma. Positive relationships between cardiovascular risk and concentrations of neurogranin and α‐synuclein were amplified in severe asthma. Severe asthma also amplified the deleterious associations that apolipoprotein E ε4 carrier status, cardiovascular risk, and phosphorylated tau181/amyloid beta42 have with rate of cognitive decline.DiscussionOur data suggest that severe asthma is associated with synaptic degeneration and may compound risk for dementia posed by cardiovascular disease and genetic predisposition.HighlightsThose with severe asthma showed evidence of higher dementia risk than controls evidenced by: higher levels of the synaptic degeneration biomarker neurogranin regardless of cognitive status, cardiovascular or genetic risk, and controlling for demographics.steeper increase in levels of synaptic degeneration biomarkers neurogranin and α‐synuclein with increasing cardiovascular risk.accelerated cognitive decline with higher cardiovascular risk, genetic predisposition, or pathological tau.

α-Synuclein in Plasma-Derived Extracellular Vesicles Is a Potential Biomarker of Parkinson's Disease.

Extracellular vesicles are small vesicles that are released from many cells, including neurons. α-Synuclein has recently been described in extracellular vesicles derived from the central nervous system and may contribute to the spreading of disease pathology in α-synuclein-related neurodegeneration. We aimed to examine the potential diagnostic value of α-synuclein in plasma extracellular vesicles from patients with Parkinson's disease (PD). Preanalytical variables were studied to establish an optimized assay for preparation of plasma extracellular vesicles and detection of extracellular vesicle-derived α-synuclein. Plasma samples were obtained from 2 independent cohorts. The Tübingen cohort contained 96 patients with PD, 50 patients with dementia with Lewy bodies, 50 patients with progressive supranuclear palsy (PSP), and 42 healthy controls; the Kassel cohort included 47 patients with PD, 43 patients with dementia with Lewy bodies, and 36 controls with secondary parkinsonian syndromes. Extracellular vesicles were prepared from total plasma by size exclusion chromatography and quantified by nanoparticle tracking analysis, α-synuclein content was measured by an electrochemiluminescence assay. α-Synuclein concentration in plasma extracellular vesicles provided the best discrimination between PD, dementia with Lewy bodies, PSP, and healthy controls, with an area under the curve of 0.804 (PD vs dementia with Lewy bodies), 0.815 (PD vs. PSP), and 0.769 (PD vs healthy controls) in the Tübingen cohort. Results were validated in the Kassel cohort. The concentration of α-synuclein in plasma extracellular vesicles may serve as a potential diagnostic biomarker for PD. © 2021 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.

Interplay between tau and α-synuclein liquid-liquid phase separation.

In Parkinson's disease with dementia, up to 50% of patients develop a high number of tau‐containing neurofibrillary tangles. Tau‐based pathologies may thus act synergistically with the α‐synuclein pathology to confer a worse prognosis. A better understanding of the relationship between the two distinct pathologies is therefore required. Liquid–liquid phase separation (LLPS) of proteins has recently been shown to be important for protein aggregation involved in amyotrophic lateral sclerosis, whereas tau phase separation has been linked to Alzheimer's disease. We therefore investigated the interaction of α‐synuclein with tau and its consequences on tau LLPS. We find α‐synuclein to have a low propensity for both, self‐coacervation and RNA‐mediated LLPS at pH 7.4. However, full‐length but not carboxy‐terminally truncated α‐synuclein efficiently partitions into tau/RNA droplets. We further demonstrate that Cdk2‐phosphorylation promotes the concentration of tau into RNA‐induced droplets, but at the same time decreases the amount of α‐synuclein inside the droplets. NMR spectroscopy reveals that the interaction of the carboxy‐terminal domain of α‐synuclein with the proline‐rich region P2 of tau is required for the recruitment of α‐synuclein into tau droplets. The combined data suggest that the concentration of α‐synuclein into tau‐associated condensates can contribute to synergistic aSyn/tau pathologies.

Molecular crowding accelerates aggregation of α-synuclein by altering its folding pathway.

Intracellular macromolecular crowding can lead to increased aggregation of proteins, especially those that lack a natively folded conformation. Crowding may also be mimicked by the addition of polymers like polyethylene glycol (PEG) in vitro. α-Synuclein is an intrinsically disordered protein that exhibits increased aggregation and amyloid fibril formation in a crowded environment. Two hypotheses have been proposed to explain this observation. One is the excluded volume effect positing that reduced water activity in a crowded environment leads to increased effective protein concentration, promoting aggregation. An alternate explanation is that increased crowding facilitates conversion to a non-native form increasing the rate of aggregation. In this work, we have segregated these two hypotheses to investigate which one is operating. We show that mere increase in concentration of α-synuclein is not enough to induce aggregation and consequent fibrillation. In vitro, we find a complex relationship between PEG concentrations and aggregation, in which smaller PEGs delay fibrillation; while, larger ones promote fibril nucleation. In turn, while PEG600 did not increase the rate of aggregation, PEG1000 did and PEG4000 and PEG12000 slowed it but led to a higher overall fibril burden in the latter to cases. In cells, PEG4000 reduces the aggregation of α-synuclein but in a way specific to the cellular environment/due to cellular factors. The aggregation of the similarly sized, globular lysozyme does not increase in vitro when at the same concentrations with either PEG8000 or PEG12000. Thus, natively disordered α-synuclein undergoes a conformational transition in specific types of crowded environment, forming an aggregation-prone conformer.

Plasma Total α-Synuclein and Neurofilament Light Chain: Clinical Validation for Discriminating Parkinson’s Disease from Normal Control

Background: A previously published paper (referred to as the original cohort) showed that using a cutoff value of 116.1 fg/mL for the plasma total α-synuclein concentrations could discriminate Parkinson’s disease (PD) patients from normal controls (NCs). In this study, another independent cohort (referred to as the validation cohort) was recruited to validate the agreement between the clinical diagnosis and the use of plasma total α-synuclein to identify PD patients. In addition to total α-synuclein, plasma neurofilament light chain (NfL) in the validation cohort was detected. Methods: Seventy PD patients and 33 NCs were enrolled in the validation cohort. A clinical diagnosis and the immunomagnetic reduction (IMR) assay for plasma total α-synuclein were performed for each participant. Thirty-three of 70 PD patients and 23 of 33 NCs were subjected to the plasma NfL assay via IMR. Results: The positive, negative, and overall percentages of agreement between the clinical diagnosis and plasma total α-synuclein diagnosis determined based on 116.1 fg/mL as the cutoff value were found to be 0.943, 0.818, and 0.903, respectively. The PD patients and NCs showed plasma NfL levels of 8.38 ± 4.19 pg/mL and 17.6 ± 7.95 pg/mL (p < 0.001), respectively. The cutoff value of the plasma NfL level used to differentiate PD patients from NCs was 12.8 pg/mL, with sensitivity and specificity values of 0.788 and 0.870, respectively. Conclusion: The results demonstrate the usefulness of the plasma total α-synuclein concentration to discriminate PD patients from NCs and reveal the elevation of the plasma NfL level in PD patients.

Extracellular clusterin limits the uptake of α-synuclein fibrils by murine and human astrocytes.

The progressive neuropathological damage seen in Parkinson's disease (PD) is thought to be related to the spreading of aggregated forms of α‐synuclein. Clearance of extracellular α‐synuclein released by degenerating neurons may be therefore a key mechanism to control the concentration of α‐synuclein in the extracellular space. Several molecular chaperones control misfolded protein accumulation in the extracellular compartment. Among these, clusterin, a glycoprotein associated with Alzheimer's disease, binds α‐synuclein aggregated species and is present in Lewy bodies, intraneuronal aggregates mainly composed by fibrillary α‐synuclein. In this study, using murine primary astrocytes with clusterin genetic deletion, human‐induced pluripotent stem cell (iPSC)‐derived astrocytes with clusterin silencing and two animal models relevant for PD we explore how clusterin affects the clearance of α‐synuclein aggregates by astrocytes. Our findings showed that astrocytes take up α‐synuclein preformed fibrils (pffs) through dynamin‐dependent endocytosis and that clusterin levels are modulated in the culture media of cells upon α‐synuclein pffs exposure. Specifically, we found that clusterin interacts with α‐synuclein pffs in the extracellular compartment and the clusterin/α‐synuclein complex can be internalized by astrocytes. Mechanistically, using clusterin knock‐out primary astrocytes and clusterin knock‐down hiPSC‐derived astrocytes we observed that clusterin limits the uptake of α‐synuclein pffs by cells. Interestingly, we detected increased levels of clusterin in the adeno‐associated virus‐ and the α‐synuclein pffs‐ injected mouse model, suggesting a crucial role of this chaperone in the pathogenesis of PD. Overall, our observations indicate that clusterin can limit the uptake of extracellular α‐synuclein aggregates by astrocytes and, hence, contribute to the spreading of Parkinson pathology.

Ambroxol for the Treatment of Patients With Parkinson Disease With and Without Glucocerebrosidase Gene Mutations

Mutations of the glucocerebrosidase gene, GBA1 (OMIM 606463), are the most important risk factor for Parkinson disease (PD). In vitro and in vivo studies have reported that ambroxol increases β-glucocerebrosidase (GCase) enzyme activity and reduces α-synuclein levels. These observations support a potential role for ambroxol therapy in modifying a relevant pathogenetic pathway in PD. To assess safety, tolerability, cerebrospinal fluid (CSF) penetration, and target engagement of ambroxol therapy with GCase in patients with PD with and without GBA1 mutations. An escalating dose of oral ambroxol to 1.26 g per day. This single-center open-label noncontrolled clinical trial was conducted between January 11, 2017, and April 25, 2018, at the Leonard Wolfson Experimental Neuroscience Centre, a dedicated clinical research facility and part of the University College London Queen Square Institute of Neurology in London, United Kingdom. Participants were recruited from established databases at the Royal Free London Hospital and National Hospital for Neurology and Neurosurgery in London. Twenty-four patients with moderate PD were evaluated for eligibility, and 23 entered the study. Of those, 18 patients completed the study; 1 patient was excluded (failed lumbar puncture), and 4 patients withdrew (predominantly lumbar puncture-related complications). All data analyses were performed from November 1 to December 14, 2018. Primary outcomes at 186 days were the detection of ambroxol in the CSF and a change in CSF GCase activity. Of the 18 participants (15 men [83.3%]; mean [SD] age, 60.2 [9.7] years) who completed the study, 17 (8 with GBA1 mutations and 9 without GBA1 mutations) were included in the primary analysis. Between days 0 and 186, a 156-ng/mL increase in the level of ambroxol in CSF (lower 95% confidence limit, 129 ng/mL; P < .001) was observed. The CSF GCase activity decreased by 19% (0.059 nmol/mL per hour; 95% CI, -0.115 to -0.002; P = .04). The ambroxol therapy was well tolerated, with no serious adverse events. An increase of 50 pg/mL (13%) in the CSF α-synuclein concentration (95% CI, 14-87; P = .01) and an increase of 88 ng/mol (35%) in the CSF GCase protein levels (95% CI, 40-137; P = .002) were observed. Mean (SD) scores on part 3 of the Movement Disorders Society Unified Parkinson Disease Rating Scale decreased (ie, improved) by 6.8 (7.1) points (95% CI, -10.4 to -3.1; P = .001). These changes were observed in patients with and without GBA1 mutations. The study results suggest that ambroxol therapy was safe and well tolerated; CSF penetration and target engagement of ambroxol were achieved, and CSF α-synuclein levels were increased. Placebo-controlled clinical trials are needed to examine whether ambroxol therapy is associated with changes in the natural progression of PD. ClinicalTrials.gov identifier: NCT02941822; EudraCT identifier: 2015-002571-24.

α-synuclein promotes progression of Parkinson's disease by upregulating autophagy signaling pathway to activate NLRP3 inflammasome.

Mechanism by which α-synuclein affects the progression of Parkinson's disease through Pyrin Domain Containing Protein 3 (NLRP3) was explored. Peripheral blood plasma of 40 Parkinson's disease patients and 40 normal healthy people attending the department of neurology of the Third Affiliated Hospital of Qiqihar Medical University were collected from March 2018 to January 2019. The expression levels of oligomers, phosphorylated α-synuclein, interleukin-1β (IL-1β), interleukin-6 (IL-6) and transforming growth factor-α (TGF-α) in plasma were detected by ELISA. Astrocytes in mouse brain tissues were extracted by primary culture method, the cells were divided into drug group and the drug + inhibitor group. After adding 0, 5, 10 and 20 µg oligomerized α-synuclein or 5 mM autophagy inhibitor 3-Methyladenine (3-MA), the expression level of NLRP3, caspase-1, IL-1β and Atg5 proteins in the cells was detected. The expression level of IL-1β in peripheral blood of PD patients was significantly increased (0.604±0.136 µmol/l vs. 1.876±0.327 µmol/l, P=0.002), while there was no significant difference between IL-6 and TGF-α. Both oligomers (0.171±0.045 µmol/l vs. 0.676±0.084 µmol/l, P<0.0001) and phosphorylated α-synuclein (0.128±0.041 µmol/l vs. 0.849±0.108 µmol/l, P<0.0001) in peripheral blood of PD patients were significantly elevated. The expression levels of NLRP3, caspase-1 and IL-1β in mouse astrocytes all increased with the increase of the concentration of oligomerized α-synuclein, and Atg5 protein expression also increased gradually with the concentration, and reached the highest level when the concentration was 10 µg/ml. The expression levels of NLRP3, caspase-1 and IL-1β were inhibited after the addition of autophagy inhibitor 3-MA. α-synuclein mediates the activation of NLRP3 inflammasome in PD patients by upregulating Atg5 protein expression.