Overexpression of the calpain-specific inhibitor calpastatin reduces human alpha-Synuclein processing, aggregation and synaptic impairment in [A30P]αSyn transgenic mice.

Lewy bodies, a pathological hallmark of Parkinson's disease (PD), contain aggregated alpha-synuclein (αSyn), which is found in several modified forms and can be discovered phosphorylated, ubiquitinated and truncated. Aggregation-prone truncated species of αSyn caused by aberrant cleavage of this fibrillogenic protein are hypothesized to participate in its sequestration into inclusions subsequently leading to synaptic dysfunction and neuronal death. Here, we investigated the role of calpain cleavage of αSyn in vivo by generating two opposing mouse models. We crossed into human [A30P]αSyn transgenic (i) mice deficient for calpastatin, a calpain-specific inhibitor, thus enhancing calpain activity (SynCAST(-)) and (ii) mice overexpressing human calpastatin leading to reduced calpain activity (SynCAST(+)). As anticipated, a reduced calpain activity led to a decreased number of αSyn-positive aggregates, whereas loss of calpastatin led to increased truncation of αSyn in SynCAST(-). Furthermore, overexpression of calpastatin decreased astrogliosis and the calpain-dependent degradation of synaptic proteins, potentially ameliorating the observed neuropathology in [A30P]αSyn and SynCAST(+) mice. Overall, our data further support a crucial role of calpains, particularly of calpain 1, in the pathogenesis of PD and in disease-associated aggregation of αSyn, indicating a therapeutic potential of calpain inhibition in PD.

Multifactorial assessment of Parkinson's disease course and outcomes using trajectory modeling in a multiethnic, multisite cohort - extension of the LONG-PD study.

Background: The severity, progression, and outcomes of motor and non-motor symptoms in Parkinson's disease (PD) are quite variable. Following PD cohorts holds promise for identifying predictors of disease severity and progression. Methods: PD patients (N = 871) were enrolled at five sites. Enrollment occurred within 5 years of initial motor symptom onset. Disease progression was assessed annually for 2-to-10 years after onset. Group-based trajectory modeling was used to identify groups differing in disease progression. Models were developed for UPDRS-III scores, UPDRS-III tremor and bradykinesia-rigidity subscores, Hoehn & Yahr (H&Y) stage, Mini-Mental Status Exam (MMSE) scores, and UPDRS-III, H&Y and MMSE scores considered together. Predictors of trajectory-group membership were modeled simultaneously with the trajectories. Kaplan-Meier survival analysis evaluated survival free of PD outcomes. Results: The best fitting models identified three groups. One showed a relatively benign, slowly progressing trajectory (Group 1), a second showed a moderate, intermediately progressing trajectory (Group 2), and a third showed a more severe, rapidly progressing trajectory (Group 3). Stable trajectory-group membership occurred relatively early in the disease course, 5 years after initial motor symptom. Predictors of intermediate and more severe trajectory-group membership varied across the single variable models and the multivariable model jointly considering UPDRS-III, H&Y and MMSE scores. In the multivariable model, membership in Group 2 (28.4% of patients), relative to Group 1 (50.5%), was associated with male sex, younger age-at-onset, fewer education-years, pesticide exposure, absence of reported head injury, and akinetic/rigid subtype at initial presentation. Membership in Group 3 (21.3%), relative to Group 1, was associated with older age-at-onset, fewer education-years, pesticide exposure, and the absence of a tremor-predominant subtype at initial presentation. Persistent freezing, persistent falls, and cognitive impairment occurred earliest and more frequently in Group 3, later and less frequently in Group 2, and latest and least frequently in Group 1. Furthermore, autonomic complications, dysphagia, and psychosis occurred more frequently in Groups 2 and 3 than in Group 1. Conclusion: Modeling disease course using multiple objective assessments over an extended follow-up duration identified groups that more accurately reflect differences in PD course, prognosis, and outcomes than assessing single parameters over shorter intervals.

Non-replication of association for six polymorphisms from meta-analysis of genome-wide association studies of Parkinson's disease: large-scale collaborative study.

Early genome-wide association (GWA) studies on Parkinson's disease (PD) have not been able to yield conclusive, replicable signals of association, perhaps due to limited sample size. We aimed to investigate whether association signals derived from the meta-analysis of the first two GWA investigations might be replicable in different populations. We examined six single-nucleotide polymorphisms (SNPs) (rs1000291, rs1865997, rs2241743, rs2282048, rs2313982, and rs3018626) that had reached nominal significance with at least two of three different strategies proposed in a previous analysis of the original GWA studies. Investigators from the "Genetic Epidemiology of Parkinson's Disease" (GEOPD) consortium were invited to join in this study. Ten teams contributed replication data from 3,458 PD cases and 3,719 controls. The data from the two previously published GWAs (599 PD cases, 592 controls and 443 sibling pairs) were considered as well. All data were synthesized using both fixed and random effects models. The summary allelic odds ratios were ranging from 0.97 to 1.09 by random effects, when all data were included. The summary estimates of the replication data sets (excluding the original GWA data) were very close to 1.00 (range 0.98-1.09) and none of the effects were nominally statistically significant. The replication data sets had significantly different results than the GWA data. Our data do not support evidence that any of these six SNPs reflect susceptibility markers for PD. Much stronger signals of statistical significance in GWA platforms are needed to have substantial chances of replication. Specifically in PD genetics, this would require much larger GWA studies and perhaps novel analytical techniques.

Corrigendum: Contributing Factors and Evolution of Impulse Control Disorder in the Luxembourg Parkinson Cohort.

[This corrects the article DOI: 10.3389/fneur.2020.578924.].

Contributing Factors and Evolution of Impulse Control Disorder in the Luxembourg Parkinson Cohort.

Background: To establish the frequency of impulse control disorder (ICD) in Parkinson's disease (PD). Methods: Within the Luxembourg Parkinson's Study, PD patients were evaluated for ICD presence (score ≥ 1 on MDS-UPDRS I item 1.6), use of dopamine agonists (DA) and other medications. Results: 470 patients were enrolled. Among 217 patients without DA use, 6.9% scored positive for ICD, vs. 15.4% among 253 patients with DA use (p = 0.005). The regression analysis showed that age at PD diagnosis had only a minor impact on ICD occurrence, while there was no influence by gender or co-medications. The longitudinal study over 2 years in 156 patients demonstrated increasing ICD frequency in DA users (p = 0.005). Conclusion: This large and non-interventional study confirms that PD patients with DA treatment show higher frequency of ICD than patients without DA use. It newly demonstrates that ICD can develop independently from age, gender, or co-medications.

Machine learning-assisted neurotoxicity prediction in human midbrain organoids.

INTRODUCTION: Brain organoids are highly complex multi-cellular tissue proxies, which have recently risen as novel tools to study neurodegenerative diseases such as Parkinson's disease (PD). However, with increasing complexity of the system, usage of quantitative tools becomes challenging. OBJECTIVES: The primary objective of this study was to develop a neurotoxin-induced PD organoid model and to assess the neurotoxic effect on dopaminergic neurons using microscopy-based phenotyping in a high-content fashion. METHODS: We describe a pipeline for a machine learning-based analytical method, allowing for detailed image-based cell profiling and toxicity prediction in brain organoids treated with the neurotoxic compound 6-hydroxydopamine (6-OHDA). RESULTS: We quantified features such as dopaminergic neuron count and neuronal complexity and built a machine learning classifier with the data to optimize data processing strategies and to discriminate between different treatment conditions. We validated the approach with high content imaging data from PD patient derived midbrain organoids. CONCLUSIONS: The here described model is a valuable tool for advanced in vitro PD modeling and to test putative neurotoxic compounds.

Unraveling Molecular Mechanisms of THAP1 Missense Mutations in DYT6 Dystonia.

Mutations in THAP1 (THAP domain-containing apoptosis-associated protein 1) are responsible for DYT6 dystonia. Until now, more than eighty different mutations in THAP1 gene have been found in patients with primary dystonia, and two third of them are missense mutations. The potential pathogeneses of these missense mutations in human are largely elusive. In the present study, we generated stable transfected human neuronal cell lines expressing wild-type or mutated THAP1 proteins found in DYT6 patients. Transcriptional profiling using microarrays revealed a set of 28 common genes dysregulated in two mutated THAP1 (S21T and F81L) overexpression cell lines suggesting a common mechanism of these mutations. ChIP-seq showed that THAP1 can bind to the promoter of one of these genes, superoxide dismutase 2 (SOD2). Overexpression of THAP1 in SK-N-AS cells resulted in increased SOD2 protein expression, whereas fibroblasts from THAP1 patients have less SOD2 expression, which indicates that SOD2 is a direct target gene of THAP1. In addition, we show that some THAP1 mutations (C54Y and F81L) decrease the protein stability which might also be responsible for altered transcription regulation due to dosage insufficiency. Taking together, the current study showed different potential pathogenic mechanisms of THAP1 mutations which lead to the same consequence of DYT6 dystonia.

Automated high-throughput high-content autophagy and mitophagy analysis platform.

Autophagic processes play a central role in cellular homeostasis. In pathological conditions, the flow of autophagy can be affected at multiple and distinct steps of the pathway. Current analyses tools do not deliver the required detail for dissecting pathway intermediates. The development of new tools to analyze autophagic processes qualitatively and quantitatively in a more straightforward manner is required. Defining all autophagy pathway intermediates in a high-throughput manner is technologically challenging and has not been addressed yet. Here, we overcome those requirements and limitations by the developed of stable autophagy and mitophagy reporter-iPSC and the establishment of a novel high-throughput phenotyping platform utilizing automated high-content image analysis to assess autophagy and mitophagy pathway intermediates.

Alpha-synuclein and Parkinson's disease: implications from the screening of more than 1,900 patients.

Data on the frequency of alpha-synuclein mutations in Parkinson's disease (PD) are limited. Screening the entire coding region in 1,921 PD patients with denaturing high performance liquid chromatography and subsequent sequencing we only detected silent mutations (g.2654A>G, g.10151G>A, and g.15986A>T) and the c.209G>A substitution corresponding to the p.A53T mutation. These results demonstrate that mutations in the alpha-synuclein gene are rare and suggest that other factors contribute to alpha-synuclein aggregation in the majority of PD patients.