Brain proteome profiling implicates the complement and coagulation cascade in multiple system atrophy brain pathology.

BACKGROUND: Multiple system atrophy (MSA) is a rare, progressive, neurodegenerative disorder presenting glia pathology. Still, disease etiology and pathophysiology are unknown, but neuro-inflammation and vascular disruption may be contributing factors to the disease progression. Here, we performed an ex vivo deep proteome profiling of the prefrontal cortex of MSA patients to reveal disease-relevant molecular neuropathological processes. Observations were validated in plasma and cerebrospinal fluid (CSF) of novel cross-sectional patient cohorts. METHODS: Brains from 45 MSA patients and 30 normal controls (CTRLs) were included. Brain samples were homogenized and trypsinized for peptide formation and analyzed by high-performance liquid chromatography tandem mass spectrometry (LC-MS/MS). Results were supplemented by western blotting, immuno-capture, tissue clearing and 3D imaging, immunohistochemistry and immunofluorescence. Subsequent measurements of glial fibrillary acid protein (GFAP) and neuro-filament light chain (NFL) levels were performed by immunoblotting in plasma of 20 MSA patients and 20 CTRLs. Finally, we performed a proteome profiling of 144 CSF samples from MSA and CTRLs, as well as other parkinsonian disorders. Data were analyzed using relevant parametric and non-parametric two-sample tests or linear regression tests followed by post hoc tests corrected for multiple testing. Additionally, high-throughput bioinformatic analyses were applied. RESULTS: We quantified more than 4,000 proteins across samples and identified 49 differentially expressed proteins with significantly different abundances in MSA patients compared with CTRLs. Pathway analyses showed enrichment of processes related to fibrinolysis and complement cascade activation. Increased fibrinogen subunit ß (FGB) protein levels were further verified, and we identified an enriched recognition of FGB by IgGs as well as intra-parenchymal accumulation around blood vessels. We corroborated blood-brain barrier leakage by a significant increase in GFAP and NFL plasma levels in MSA patients that correlated to disease severity and/or duration. Proteome profiling of CSF samples acquired during the disease course, confirmed increased total fibrinogen levels and immune-related components in the soluble fraction of MSA patients. This was also true for the other atypical parkinsonian disorders, dementia with Lewy bodies and progressive supra-nuclear palsy, but not for Parkinson's disease patients. CONCLUSION: Our results implicate activation of the fibrinolytic cascade and immune system in the brain as contributing factors in MSA associated with a more severe disease course.

Cerebrospinal fluid and plasma distribution of anti-α-synuclein IgMs and IgGs in multiple system atrophy and Parkinson's disease.

INTRODUCTION: Ubiquitous naturally occurring autoantibodies (nAbs) against alpha-synuclein (α-syn) may play important roles in the pathogenesis of Multiple System Atrophy (MSA) and Parkinson's disease (PD). Recently, we reported reduced high-affinity/avidity anti-α-syn nAbs levels in plasma from MSA and PD patients, along with distinct inter-group immunoglobulin (Ig)G subclass distributions. The extent to which these observations in plasma may reflect corresponding levels in the cerebrospinal fluid (CSF) is unknown. METHODS: Using competitive and indirect ELISAs, we investigated the affinity/avidity of CSF anti-α-syn nAbs as well as the CSF and plasma distribution of IgG subclasses and IgM nAbs in a cross-sectional cohort of MSA and PD patients. RESULTS: Repertoires of high-affinity/avidity anti-α-syn IgG nAbs were reduced in CSF samples from MSA and PD patients compared to controls. Furthermore, anti-α-syn IgM nAb levels were relatively lower in CSF and plasma from MSA patients but were reduced only in plasma from PD patients. Interestingly, anti-α-syn IgG subclasses presented disease-specific profiles both in CSF and plasma. Anti-α-syn IgG1, IgG2 and IgG3 levels were relatively increased in CSF of MSA patients, whereas PD patients showed increased anti-α-syn IgG2 and reduced anti-α-syn IgG4 levels. CONCLUSIONS: Differences in the plasma/CSF distribution of anti-α-syn nAbs seem to be a common feature of synucleinopathies. Our data add further support to the notion that MSA and PD patients may have compromised immune reactivity towards α-syn. The differing α-syn-specific systemic immunological responses may reflect their specific disease pathophysiologies. These results are encouraging for further investigation of these immunological mechanisms in neurodegenerative diseases.

Risk of Multiple System Atrophy and the Use of Anti-Inflammatory Drugs: A Danish Register-Based Case-Control Study.

INTRODUCTION: Multiple system atrophy (MSA) is a rare rapidly progressive atypical Parkinson disorder presenting clinically with parkinsonism and/or a cerebellar syndrome in combination with dysautonomia. Severe neuroinflammation develops along with hallmark neuropathological changes, and as in Parkinson's disease, intake of anti-inflammatory medication has been hypothesized to be protective for development of disease. We aimed to investigate if use of non-aspirin nonsteroidal anti-inflammatory drugs (NSAIDs), low-dose aspirin, or statins were associated with a reduced risk of MSA. METHODS: We performed a register-based case-control study in MSA (n = 155) cases and population controls (n= 7,750) matched on age, gender, and place of residency by risk-set sampling. Pharmacological exposure prior to diagnosis was assessed in 2 categories (user vs. nonuser, cumulated dose in tertiles [T1-T3]). In an unconditional logistic regression model, adjusted for age, gender, residency, and chronic obstructive pulmonary disease (COPD), we estimated ORs and 95% CIs. RESULTS: Data suggested a trend towards non-aspirin NSAID use to be associated with a decreased risk of MSA (OR 0.72 [95% CI 0.49-1.06]) compared to nonusers, decreasing dose-dependently (T2 OR 0.77 [95% CI 0.43-1.38]; T3 OR 0.55 [95% CI 0.29-1.06]). However, data were based on small numbers. Use of statins and low-dose aspirin was not associated with a decreased risk of MSA. Results were lagged 5 years from index date to address reverse causation. CONCLUSION: A trend toward use of non-aspirin NSAID and an associated reduced risk of MSA was observed in this study. However, our analyses had limited statistical precision, and further studies including larger sample sizes and longer exposure periods are needed.

Distinct Autoimmune Anti-α-Synuclein Antibody Patterns in Multiple System Atrophy and Parkinson's Disease.

Aggregation of alpha-synuclein (α-syn) is considered to be the major pathological hallmark and driving force of Multiple System Atrophy (MSA) and Parkinson's disease (PD). Immune dysfunctions have been associated with both MSA and PD and recently we reported that the levels of natural occurring autoantibodies (NAbs) with high-affinity/avidity toward α-synuclein are reduced in MSA and PD patients. Here, we aimed to evaluate the plasma immunoglobulin (Ig) composition binding α-syn and other amyloidogenic neuropathological proteins, and to correlate them with disease severity and duration in MSA and PD patients. All participants were recruited from a single neurological unit and the plasma samples were stored for later research at the Bispebjerg Movement Disorder Biobank. All patients were diagnosed according to current consensus criteria. Using multiple variable linear regression analyses, we observed higher levels of anti-α-syn IgG1 and IgG3 NAbs in MSA vs. PD, higher levels of anti-α-syn IgG2 NAbs in PD compared to controls, whereas anti-α-syn IgG4 NAbs were reduced in PD compared to MSA and controls. Anti-α-syn IgM levels were decreased in both MSA and PD. Further our data supported that MSA patients' immune system was affected with reduced IgG1 and IgM global levels compared to PD and controls, with further reduced global IgG2 levels compared to PD. These results suggest distinct autoimmune patterns in MSA and PD. These findings suggest a specific autoimmune physiological mechanism involving responses toward α-syn, differing in neurodegenerative disease with overlapping α-syn pathology.

The biomarker potential of cell-free microRNA from cerebrospinal fluid in Parkinsonian Syndromes.

BACKGROUND: MicroRNAs are small noncoding RNAs involved in the post-transcriptional regulation of protein synthesis. Extracellular microRNAs are accessible in a stable form in biofluids. OBJECTIVES: The aim was to identify individual microRNAs and/or subsets of microRNAs in CSF with biomarker potential and thus identify specific putative pathophysiological pathways. METHODS: In a two-step exploratory study design of PD, MSA, PSP, and controls, we initially profiled CSF microRNAs in a pilot cohort (n = 40) by screening for 372 microRNAs. Subsequently, we attempted to validate findings in an independent study cohort in CSF (n = 118) and ethylenediaminetetraacetic acid plasma (n = 114). This study cohort encompassed 46 microRNAs, of which 26 were singled out from the pilot cohort, and an additional 20 microRNAs were added based on previous publications. The most accurate diagnostic microRNA classifiers were identified in a multivariable logistic regression model adjusted for age and sex. RESULTS: A set of three microRNAs in CSF discriminated PD and MSA from controls with good diagnostic accuracy by receiver operating characteristics curve evaluation. The microRNAs were for PD versus controls: miR-7-5p, miR-331-5p, and miR-145-5p (area under the curve = 0.88) and MSA versus controls: miR-7-5p, miR-34c-3p, and miR-let-7b-5p (area under the curve = 0.87). The classifier that best distinguished MSA and PD consisted of two microRNAs: miR-9-3p and miR-106b-5p (area under the curve = 0.73). A single microRNA, miR-106b-5p, provided the best discrimination between PD and PSP (area under the curve = 0.85) in the CSF. CONCLUSIONS: Levels of specific trios of CSF-microRNAs discriminate well between α-synucleinopathies (PD and MSA) and controls. The results need to be validated in larger, independent cohorts. © 2018 International Parkinson and Movement Disorder Society.

Occupational and recreational physical activity and Parkinson's disease in Denmark.

Objectives This study aimed to examine whether occupational and physical activity (PA) at different ages contribute to Parkinson's disease (PD) risk in a large population-based case-control study in Denmark. Methods We identified 1828 PD patients from the Danish National Hospital Register and recruited 1909 gender and year of birth matched controls from the Danish Central Population Register. Occupational and leisure-time PA were determined from a job exposure matrix based on occupational history and self-reported leisure-time information. Results No association was found for occupational PA alone in men, but higher leisure-time PA (≥5 hours/week of strenuous activities) in young adulthood (15-25 years) was associated with a lower PD risk (adjusted odds ratio (OR adj) 0.75, 95% confidence interval (95% CI) 0.62-0.90); men who engaged in high occupational and high leisure-time PA in young adulthood had the lowest PD risk (OR adj0.58, 95% CI 0.41-0.81). Among women, inverse associations were found for occupation PA before age 50 (highest vs lowest, OR adj0.75, 95% CI 0.55-1.06) and strenuous leisure-time PA after age 50 (OR adj0.65, 95% CI 0.87-0.99); no clear pattern was seen for leisure and occupational PA combined. Conclusions We observed gender-specific inverse associations between occupational and leisure-time PA and PD risk; however, we cannot preclude reverse causation especially in older ages since PD has a long prodromal stage that might lead to a reduction of PA years before motor symptom onset and PD diagnosis.

Clinical Features in a Danish Population-Based Cohort of Probable Multiple System Atrophy Patients.

BACKGROUND: Multiple system atrophy (MSA) is a rare, sporadic and progressive neurodegenerative disorder. We aimed to describe the clinical features of Danish probable MSA patients, evaluate their initial response to dopaminergic therapy and examine mortality. METHODS: From the Danish National Patient Registry, we identified 782 patients diagnosed with conditions potentially compatible with probable MSA (International Classification of Diseases, version 10 (ICD-10) codes G23.2, G23.8 and G23.9) during 1994-2009. Through medical record review, we narrowed our sample to 115 patients who fulfilled the criteria for probable MSA. We recorded clinical features, examined differences by MSA subtype and used Kaplan-Meier survival analysis to examine mortality. RESULTS: The mean age at onset of patients with probable MSA was 60.2 years (range 36-75 years) and mean time to wheelchair dependency was 4.7 years (range 0-15 years). One-third of patients experienced a transient improvement in motor symptoms with use of levodopa. Median survival from disease onset was 6.9 years (range 1-16 years, 95% CI 6.3-7.5) with no apparent variation according to gender or subtype. CONCLUSIONS: Our nationwide approach corroborated that MSA is associated with diverse and grave symptoms, only limited response to levodopa, and poor prognosis.

Dependency and transfer incomes in idiopathic Parkinson's disease.

INTRODUCTION: Idiopathic Parkinson's disease (IPD) is a progressive neurodegenerative disorder affecting approximately 1% of the population above 65 years of age. The aim of this study was to define the estimated Danish IPD population and to elucidate source of income and labour market affiliation for working-age IPD patients. MATERIAL AND METHODS: IPD cases were included through the Danish Register of Medicinal Product Statistics. The participants had to be alive by the end of 2010 and at least twice have cashed in prescriptions on IPD medication in the 2009-2010 period. Information on employment status and transfer income was retrieved through the DREAM database under the Danish Ministry of Employment. RESULTS: A total of 7,033 estimated IPD patients were identified. The mean age at time of registration (2010, week 50) was 72 years. Overall, 7% of the IPD patients were employed and 5% were self-supportive. In the working age range (18-64 years), 25% were employed and 10% enrolled in supported employment. Compared with the age-adjusted general population, twice as many IPD patients were outside the ordinary labour market and, furthermore, the proportion receiving anticipatory pension was increased threefold. The majority (89%) of the patients were living at home with a spouse (59%). 11% were nursing home residents. CONCLUSION: The working age IPD population was more prone to be outside employment and to receive public transfer income than an age-adjusted population sample. FUNDING: The study was funded by the Danish Parkinson Association. TRIAL REGISTRATION: not relevant.