Single-cell sequencing of human midbrain reveals glial activation and a Parkinson-specific neuronal state.

Idiopathic Parkinson's disease is characterized by a progressive loss of dopaminergic neurons, but the exact disease aetiology remains largely unknown. To date, Parkinson's disease research has mainly focused on nigral dopaminergic neurons, although recent studies suggest disease-related changes also in non-neuronal cells and in midbrain regions beyond the substantia nigra. While there is some evidence for glial involvement in Parkinson's disease, the molecular mechanisms remain poorly understood. The aim of this study was to characterize the contribution of all cell types of the midbrain to Parkinson's disease pathology by single-nuclei RNA sequencing and to assess the cell type-specific risk for Parkinson's disease using the latest genome-wide association study. We profiled >41 000 single-nuclei transcriptomes of post-mortem midbrain from six idiopathic Parkinson's disease patients and five age-/sex-matched controls. To validate our findings in a spatial context, we utilized immunolabelling of the same tissues. Moreover, we analysed Parkinson's disease-associated risk enrichment in genes with cell type-specific expression patterns. We discovered a neuronal cell cluster characterized by CADPS2 overexpression and low TH levels, which was exclusively present in idiopathic Parkinson's disease midbrains. Validation analyses in laser-microdissected neurons suggest that this cluster represents dysfunctional dopaminergic neurons. With regard to glial cells, we observed an increase in nigral microglia in Parkinson's disease patients. Moreover, nigral idiopathic Parkinson's disease microglia were more amoeboid, indicating an activated state. We also discovered a reduction in idiopathic Parkinson's disease oligodendrocyte numbers with the remaining cells being characterized by a stress-induced upregulation of S100B. Parkinson's disease risk variants were associated with glia- and neuron-specific gene expression patterns in idiopathic Parkinson's disease cases. Furthermore, astrocytes and microglia presented idiopathic Parkinson's disease-specific cell proliferation and dysregulation of genes related to unfolded protein response and cytokine signalling. While reactive patient astrocytes showed CD44 overexpression, idiopathic Parkinson's disease microglia revealed a pro-inflammatory trajectory characterized by elevated levels of IL1B, GPNMB and HSP90AA1. Taken together, we generated the first single-nuclei RNA sequencing dataset from the idiopathic Parkinson's disease midbrain, which highlights a disease-specific neuronal cell cluster as well as 'pan-glial' activation as a central mechanism in the pathology of the movement disorder. This finding warrants further research into inflammatory signalling and immunomodulatory treatments in Parkinson's disease.

Genetic landscape of Parkinson's disease and related diseases in Luxembourg.

Objectives: To explore the genetic architecture of PD in the Luxembourg Parkinson's Study including cohorts of healthy people and patients with Parkinson's disease (PD) and atypical parkinsonism (AP). Methods: 809 healthy controls, 680 PD and 103 AP were genotyped using the Neurochip array. We screened and validated rare single nucleotide variants (SNVs) and copy number variants (CNVs) within seven PD-causing genes (LRRK2, SNCA, VPS35, PRKN, PARK7, PINK1 and ATP13A2). Polygenic risk scores (PRSs) were generated using the latest genome-wide association study for PD. We then estimated the role of common variants in PD risk by applying gene-set-specific PRSs. Results: We identified 60 rare SNVs in seven PD-causing genes, nine of which were pathogenic in LRRK2, PINK1 and PRKN. Eleven rare CNVs were detected in PRKN including seven duplications and four deletions. The majority of PRKN SNVs and CNVs carriers were heterozygous and not differentially distributed between cases and controls. The PRSs were significantly associated with PD and identified specific molecular pathways related to protein metabolism and signal transduction as drivers of PD risk. Conclusion: We performed a comprehensive genetic characterization of the deep-phenotyped individuals of the Luxembourgish Parkinson's Study. Heterozygous SNVs and CNVs in PRKN were not associated with higher PD risk. In particular, we reported novel digenic variants in PD related genes and rare LRRK2 SNVs in AP patients. Our findings will help future studies to unravel the genetic complexity of PD.

Accurate long-read sequencing identified GBA1 as major risk factor in the Luxembourgish Parkinson's study.

Heterozygous variants in the glucocerebrosidase GBA1 gene are an increasingly recognized risk factor for Parkinson's disease (PD). Due to the GBAP1 pseudogene, which shares 96% sequence homology with the GBA1 coding region, accurate variant calling by array-based or short-read sequencing methods remains a major challenge in understanding the genetic landscape of GBA1-associated PD. We analyzed 660 patients with PD, 100 patients with Parkinsonism and 808 healthy controls from the Luxembourg Parkinson's study, sequenced using amplicon-based long-read DNA sequencing technology. We found that 12.1% (77/637) of PD patients carried GBA1 variants, with 10.5% (67/637) of them carrying known pathogenic variants (including severe, mild, risk variants). In comparison, 5% (34/675) of the healthy controls carried GBA1 variants, and among them, 4.3% (29/675) were identified as pathogenic variant carriers. We found four GBA1 variants in patients with atypical parkinsonism. Pathogenic GBA1 variants were 2.6-fold more frequently observed in PD patients compared to controls (OR = 2.6; CI = [1.6,4.1]). Three novel variants of unknown significance (VUS) were identified. Using a structure-based approach, we defined a potential risk prediction method for VUS. This study describes the full landscape of GBA1-related parkinsonism in Luxembourg, showing a high prevalence of GBA1 variants as the major genetic risk for PD. Although the long-read DNA sequencing technique used in our study may be limited in its effectiveness to detect potential structural variants, our approach provides an important advancement for highly accurate GBA1 variant calling, which is essential for providing access to emerging causative therapies for GBA1 carriers.

Body-First Subtype of Parkinson's Disease with Probable REM-Sleep Behavior Disorder Is Associated with Non-Motor Dominant Phenotype.

BACKGROUND: The hypothesis of body-first vs. brain-first subtype of PD has been proposed with REM-Sleep behavior disorder (RBD) defining the former. The body-first PD presumes an involvement of the brainstem in the pathogenic process with higher burden of autonomic dysfunction. OBJECTIVE: To identify distinctive clinical subtypes of idiopathic Parkinson's disease (iPD) in line with the formerly proposed concept of body-first vs. brain-first subtypes in PD, we analyzed the presence of probable RBD (pRBD), sex, and the APOEɛ4 carrier status as potential sub-group stratifiers. METHODS: A total of 400 iPD patients were included in the cross-sectional analysis from the baseline dataset with a completed RBD Screening Questionnaire (RBDSQ) for classifying as pRBD by using the cut-off RBDSQ≥6. Multiple regression models were applied to explore (i) the effect of pRBD on clinical outcomes adjusted for disease duration and age, (ii) the effect of sex on pRBD, and (iii) the association of APOEɛ4 and pRBD. RESULTS: iPD-pRBD was significantly associated with autonomic dysfunction (SCOPA-AUT), level of depressive symptoms (BDI-I), MDS-UPDRS I, hallucinations, and constipation, whereas significantly negatively associated with quality of life (PDQ-39) and sleep (PDSS). No significant association between sex and pRBD or APOE ɛ4 and pRBD in iPD was found nor did we determine a significant effect of APOE ɛ4 on the PD phenotype. CONCLUSION: We identified an RBD-specific PD endophenotype, characterized by predominant autonomic dysfunction, hallucinations, and depression, corroborating the concept of a distinctive body-first subtype of PD. We did not observe a significant association between APOE ɛ4 and pRBD suggesting both factors having an independent effect on cognitive decline in iPD.