Assessing serum anti-nuclear antibodies HEp-2 patterns in synucleinopathies.

This study investigates the presence of antinuclear antibodies (ANA) in three primary synucleinopathies - Parkinson's disease (PD), multiple system atrophy (MSA), and dementia with Lewy bodies (DLB), compared to healthy controls. Autoinflammatory disorders typically involve the immune system mistakenly attacking the body's own cells and start producing ANA. There is an increasing body of evidence that immune-mediated inflammation is a pathological feature linked to synucleinopathies. To investigate whether this could be autoimmune mediated we analyzed for ANA in the plasma of 25 MSA, 25 PD, and 17 DLB patients, along with 25 healthy controls, using the ANA HEp-2 indirect immunofluorescence antibody assay (ANA HEp-2 IFA). Contrary to initial expectations, results showed ANA HEp-2 positivity in 12% of PD, 8% of MSA patients, 18% of DLB patients, and 17% of healthy controls, indicating no increased prevalence of ANA in synucleinopathies compared to age-matched healthy individuals. Various ANA HEp-2 patterns were identified, but no specific pattern was associated with individual synucleinopathies. We conclude hereby that synucleinopathies are not associated with detectable presence of ANA in plasma.

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.

Paroxysmal Cranial Dyskinesia and Nail-Patella Syndrome Caused by a Novel Variant in the LMX1B Gene.

BACKGROUND: In a Danish family, multiple individuals in five generations present with early-onset paroxysmal cranial dyskinesia, musculoskeletal abnormalities, and kidney dysfunction. OBJECTIVE: To demonstrate linkage and to identify the underlying genetic cause of disease. METHODS: Genome-wide single-nucleotide polymorphisms analysis, Sequence-Tagged-Site marker analyses, exome sequencing, and Sanger sequencing were performed. RESULTS: Linkage analyses identified a candidate locus on chromosome 9. Exome sequencing revealed a novel variant in LMX1B present in all affected individuals, logarithm of the odds (LOD) score of z = 6.54, predicted to be damaging. Nail-patella syndrome (NPS) is caused by pathogenic variants in LMX1B encoding a transcription factor essential to cytoskeletal and kidney growth and dopaminergic and serotonergic network development. NPS is characterized by abnormal musculoskeletal features and kidney dysfunction. Movement disorders have not previously been associated with NPS. CONCLUSIONS: Paroxysmal dyskinesia is a heretofore unrecognized feature of the NPS spectrum. The pathogenic mechanism might relate to aberrant dopaminergic circuits. © 2020 International Parkinson and Movement Disorder Society.

Familial aggregation of Parkinson's disease in the Faroe Islands.

The Faroe Islands are a geographic population isolated in the North Atlantic with a high prevalence of Parkinson's disease (PD). Although environmental risk factors are well described, the familial aggregation of PD on the Islands has yet to be explored. Complete ascertainment of all patients with PD was performed, including 217 cases and 251 control subjects. All patients were neurologically assessed and diagnosed using UK Brain Bank criteria and Hohn and Yahr staging. Comprehensive genealogical and detailed cartographic analyses were performed. Relative risk and risk ratios were calculated with respect to the general population. Patients with PD in the Faroes have a higher age at symptom onset and diagnosis than for neighboring countries. Clinically, patients are similar; however, they are more likely to have affected relatives than randomly selected control subjects, matched by sex and age. Disease is most prevalent within two geographic regions. Overall, the relative risk for PD was 2.3 (95% confidence interval [CI], 1.2-4.3; P = 0.008) for siblings and 1.4 (95% CI, 1.01-1.99, P = 0.04) for first cousins. The etiology and excess prevalence of PD on the Faroes is complicated. Regional and familial clustering, and subsequent segregation analysis, suggests the disease best fits a genetic etiology with limited support for an environmental contribution. Pedigree-based analysis of PD on the Faroe Islands which has few founders and a relatively homogeneous background may elaborate on these possibilities and their joint contribution.

Severe and rapidly progressing cognitive phenotype in a SCA17-family with only marginally expanded CAG/CAA repeats in the TATA-box binding protein gene: a case report.

BACKGROUND: The autosomal dominant spinocerebellar ataxias (SCAs) confine a group of rare and heterogeneous disorders, which present with progressive ataxia and numerous other features e.g. peripheral neuropathy, macular degeneration and cognitive impairment, and a subset of these disorders is caused by CAG-repeat expansions in their respective genes. The diagnosing of the SCAs is often difficult due to the phenotypic overlap among several of the subtypes and with other neurodegenerative disorders e.g. Huntington's disease. CASE PRESENTATION: We report a family in which the proband had rapidly progressing cognitive decline and only subtle cerebellar symptoms from age 42. Sequencing of the TATA-box binding protein gene revealed a modest elongation of the CAG/CAA-repeat of only two repeats above the non-pathogenic threshold of 41, confirming a diagnosis of SCA17. Normally, repeats within this range show reduced penetrance and result in a milder disease course with slower progression and later age of onset. Thus, this case presented with an unusual phenotype. CONCLUSIONS: The current case highlights the diagnostic challenge of neurodegenerative disorders and the need for a thorough clinical and paraclinical examination of patients presenting with rapid cognitive decline to make a precise diagnosis on which further genetic counseling and initiation of treatment modalities can be based.

Autonomic dysfunction in Parkinson's disease: Results from the Faroese Parkinson's disease cohort.

The presence of autonomic symptoms are a common part of the symptomatology of Parkinsons disease (PD), with the potential to impact the quality of life of patients. The aim of this study was to assess the frequency of autonomic symptoms among Faroese PD patients compared to a control group, using the Scales for Outcome in Parkinson's Disease - Autonomic (SCOPA-AUT), and to determine the relationship between autonomic and motor symptoms in PD patients using the Unified Parkinsons Disease Rating Scale - Part III (UPDRS) and Hoehn and Yahr Scale (H&Y). The study included 54 PD patients and 190 control individuals which were unaffected relatives. The mean SCOPA-AUT scores were significantly higher for PD patients in gastrointestinal (OR = 1.62), urinary (OR = 1.38), cardiovascular (OR = 1.65), thermoregulatory (OR = 1.54) and sexual dysfunction (OR = 1.71) scores, as well as the total score (OR = 1.26). UPDRS scores were significant correlated with SCOPA-AUT scores (p = 0.015), while H&Y scores were not (p = 0.103). In conclusion, PD patients experience an increased frequency of autonomic symptoms compared with controls and the frequency is associated with the motor symptoms assessed with UPDRS. Our findings are consistent with similar studies and our current understanding of PD pathology.

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.

Impact of dietary exposure to food contaminants on the risk of Parkinson's disease.

This study aimed to investigate the association of Parkinson's disease (PD) with dietary exposure to polychlorinated biphenyls (PCBs) and methylmercury (MeHg) in a community with increased exposure levels. A total of 79 clinically verified idiopathic PD cases and 154 controls matched by sex and age were examined in this case-control study in the Faroe Islands. Blood and hair samples were collected and a questionnaire recorded lifetime information on residence, dietary habits, smoking history, and occupational exposure to solvents, pesticides, and metals. Both unconditional and conditional logistic regression analyses were used to estimate the odds ratio (OR) and 95% confidence interval (CI) in regard to relevant exposure variables. Increased ORs for dietary intakes of whale meat and blubber during adult life were statistically significant. The ORs for occupational exposure to solvents, pesticides and metals also suggested an increased risk for PD. Current serum concentrations of summation operator PCB and related contaminants suggested slightly increased ORs, although only beta-hexachlorocyclohexane (beta-HCH) was statistically significant. Increased intake of whale meat and blubber in adult life was significantly associated with PD, thus suggesting a positive association between previous exposure to marine food contaminants and development of PD.

Carnitine levels and mutations in the SLC22A5 gene in Faroes patients with Parkinson's disease.

INTRODUCTION: Mitochondrial dysfunction, oxidative stress and energy production have been implicated in the etiology of Parkinson's disease (PD). Several agents are under investigation for potential neuroprotective effects including acetyl-l-carnitine (ALC). OBJECTIVE: To investigate whether low carnitine levels and mutations in the SLC22A5 gene encoding the carnitine transporter are associates with PD risk in the Faroe Islands where the prevalence of both PD and carnitine transporter deficiency (CTD) is high. METHODS: We conducted a case-control study with 121 cases and 235 randomly selected controls, matched by gender and age. Blood spots were analyzed for free and total carnitine levels by QUATTRO LC triple quadrupole mass spectrometry (MS/MS) and sequencing performed for five genetic mutations in the SLC22A5 gene with ABI PRISM 3130. RESULTS: PD cases had significantly lower levels of free and total carnitine levels compared with controls (P < .001). However, stratifying according to mutation status, the lower levels of carnitine was only evident among the non-mutation carriers. Specifically, no difference was found in c.95A > G mutation frequency in the SLC22A5 gene among cases and controls (p = .70). CONCLUSION: Low carnitine levels seem to be associated with PD, but only in individuals without the c.95A > G mutation rendering the carnitine transporter less efficient. Thus, the difference in carnitine levels is not caused by a higher frequency of c.95A > G mutation carriers in cases. The cause may be dietary or due to different gut microbiota among cases.

[Challenges in the differential diagnoses of parkinsonism].

Parkinsonism (PD) is the clinical syndrome of bradykinesia combined with rigidity and/or tremor at rest. These are the defining characteristics of PD, but they are present in many other diseases of the brain. The most frequent differential diagnosis of PD are the atypical parkinsonian syndromes and the conditions presenting with mainly lower body parkinsonism. Discrimination between these can be challenging, especially at early stages of the disease, but nevertheless of utmost importance, because treatment and prognosis vary. Diagnoses are clinical, as disease-specific biomarkers are still lacking.

Parkinson's disease, genetic variability and the Faroe Islands.

INTRODUCTION: The Faroe Islands is a geographically isolated population in the North Atlantic with a high prevalence of Parkinson disease (PD). The disease etiology is still unknown, although dietary pollutants are considered a risk factor. The genetic risk underlying disease susceptibility has yet to be elucidated. METHODS: Sequence analysis was performed in genes previously linked with PD in 91 patients and 96 healthy control subjects. RESULTS: Fourteen missense mutations, of which one was novel, were identified in six genes. One patient (1%) did carry the known pathogenic mutation LRRK2 p.G2019S mutation, 19 patients (22%) did carry mutations of unknown significance while 70 patients (78.0%) did not have any identifiable genetic risk. A total of 14 controls (14.6%) carried mutations of unknown significance. CONCLUSION: This study suggests that rare variants in genes previously linked to PD are not major contributors to PD in the Faroe Islands. Further exome sequencing and comparative analyses within and among well-described pedigrees with multi-incident PD are now warranted.

The role of vitamin D levels and vitamin D receptor polymorphism on Parkinson's disease in the Faroe Islands.

The role of vitamin D in Parkinson's disease (PD) has been proposed and both low serum 25-hydroxyvitamin D levels (25(OH)D) and vitamin D receptor polymorphisms (VDR) have been linked to PD. The aim of this study is to investigate the associations among 25(OH)D and three VDR polymorphisms and PD in the Faroese population where the prevalence of PD is high. We conducted a case-control study where 121 cases were studied for 25(OH)D levels and VDR polymorphisms against 235 randomly selected controls, matched by gender and age. No significant difference was observed in 25(OH)D levels between PD cases and controls (P=0.49), although cases had slightly lower values than controls. As well, no differences were found in genotype frequencies between cases and controls in the VDR polymorphisms studied (ApaI, BsmI, TaqI) (P=0.70, P=0.56 and P=0.54, respectively). However, we found that VDR ApaI/AC genotype was significantly associated with 25(OH)D levels (P=0.01). Although our results indicate no association between PD and vitamin D polymorphisms and/or 25(OH)D levels, the study cannot exclude a weak association. However, the known doubling in PD prevalence in the Faroe Islands cannot be explained by the polymorphisms examined in the VDR gene or the 25(OH)D levels and has to be explored further.

The influence of preanalytical conditions on the DJ-1 concentration in human cerebrospinal fluid.

AIM: The purpose of this study was to establish the influence of centrifugation and protease activity on the cerebrospinal fluid (CSF) concentrations of DJ-1 and hemoglobin. MATERIALS & METHODS: The concentrations of DJ-1 and hemoglobin were determined in 12 (DJ-1) and six (hemoglobin) pairs of CSF samples, with one sample being stored without centrifugation and the other being centrifuged at 2000 × g before storage. The DJ-1 concentration was also determined in centrifuged and uncentrifuged CSF containing protease inhibitors and compared with values determined in centrifuged and uncentrifuged CSF samples without protease inhibitors. Furthermore, specific protein concentrations were determined in CSF from two groups, each comprising 23 patients with Parkinson's disease. In one group the CSF was centrifuged at 1300-1800 × g, 4°C, 10 min, and in the other at 2000 × g, 4°C, 10 min. RESULTS: Centrifugation at 2000 × g resulted in significantly lower CSF DJ-1 concentrations compared with no centrifugation and centrifugation at a lower g-force. There was a significant difference in the hemoglobin concentration between centrifuged and uncentrifuged CSF. In all centrifuged samples the hemoglobin concentration was <200 ng/ml including blood contaminated samples centrifuged at 2000 × g. When a protease inhibitor cocktail was added to the CSF prior to centrifugation, the DJ-1 concentration was significantly higher. CONCLUSION: Preanalytical factors such as centrifugation and protease inhibition must be carefully controlled when handling CSF for analysis of DJ-1 and other biomarkers, as DJ-1 was influenced by blood contamination, centrifugation and protease activity.

Amyloid-related biomarkers and axonal damage proteins in parkinsonian syndromes.

BACKGROUND: Clinical differentiation between parkinsonian syndromes (PS) remains a challenge despite well-established clinical diagnostic criteria. Specific diagnostic biomarkers have yet to be identified, though in recent years, studies have been published on the aid of certain brain related proteins (BRP) in the diagnosing of PS. We investigated the levels of the light subunit of neurofilament triplet protein (NF-L), total tau and phosphorylated tau, amyloid-ß(1-42), and the soluble α- and ß-cleaved fragments of amyloid precursor proteins in a cohort of patients with various PS. METHODS: Seventy-one patients with different PS and cerebellar disorders were included consecutively over 21 months. CSF was collected at inclusion. Clinical follow-up was performed after 16 months (median; range: 9-30). Statistical comparison was performed after follow-up on 53 patients in four subgroups of PS: multiple system atrophy (MSA)(n = 10), progressive supranuclear palsy (PSP)(n = 10), dementia with Lewy bodies (DLB)(n = 11), and Parkinson's disease (PD)(n = 22), using the non-parametric Kruskal-Wallis test. RESULTS: A statistically significant difference was found for NF-L (p < 0.0001, lowest values for PD), Aß(1-42,) (p = 0.002, lowest values for DLB), and sAPPα and sAPPß (p = 0.03 and 0.02, lower values observed for DLB and MSA). CONCLUSION: We demonstrate a potential role for sAPPα and sAPPß in distinguishing between PS, a finding that needs to be confirmed in future studies of larger cohorts. There is a tendency towards low levels of Aß(1-42) in DLB patients in our cohort. Further, our results support findings from previous studies, which indicate an ability to separate atypical PS from PD based on levels of NF-L.

Proteomic investigations of the ventriculo-lumbar gradient in human CSF.

Cerebrospinal fluid (CSF) is an ideal biological material in which to search for new biomarkers for improved diagnosis of neurological diseases. During a lumbar puncture between 5 and 15 mL of CSF are obtained. Previous studies have assessed the ventriculo-lumbar concentration gradient of a number of specific proteins. In the present study we took a proteomics approach to investigate the possible concentration gradient of a panel of proteins and peptides in the CSF of 16 patients with neurodegenerative diseases. Using two different mass spectrometry techniques, matrix assisted laser desorption ionization time of flight (MALDI-TOF) and surface enhanced laser desorption ionization time of flight (SELDI-TOF), we found that only one of the investigated proteins, apolipoprotein CI, was significantly decreased between the 1st and the 10th mL of CSF. Furthermore, we confirmed previous results showing a significant decrease in albumin concentration from the first to the last CSF aliquots. In conclusion, we found a significant gradient effect for only two of the measured proteins. However, a standardized procedure for CSF collection for diagnostic and research purposes is crucial to allow comparisons of results between patient groups and between laboratories. This is especially important since CSF is usually collected at several centres and variation in sampled CSF due to pre-analytical factors could complicate the interpretation of the results.

Huntington's disease-like and ataxia syndromes: identification of a family with a de novo SCA17/TBP mutation.

The autosomal dominant spinocerebellar ataxias, commonly referred to as SCAs, are clinically and genetically heterogeneous neurodegenerative disorders. Twenty-eight genetic subtypes have been identified, of which 7 are caused by expansion of a CAG trinucleotide repeat that encodes a polyglutamine tract in the respective proteins. SCA17 is caused by a CAG/CAA repeat expansion in the TATA box-binding protein-gene (TBP). In some cases the clinical phenotype of SCA17 overlaps that of Huntington's disease (HD), hence the use of the term Huntington's disease-like. We screened 89 patients with a Huntington's disease-like phenotype without the HD-gene mutation and 178 patients with genetically unclassified cerebellar ataxia for the mutation in TBP. A 33-year old woman presenting with an HD like phenotype with a de novo 54 CAG/CAA repeat expansion was identified. Her normal allele included 38 repeats. The patient's mother and father both carried normal range repeats, 38/38 and 33/39 respectively. Analysis of the repeat structures revealed that the expansion had occurred upon expansion of the longer paternal allele. We conclude that, however rare, SCA17 must be considered as a cause of Huntington's disease-like phenotypes and ataxia syndromes, also in isolated cases.