Identification of diagnostic and prognostic biomarkers of PD using a multiplex proteomics approach.

Given the complexity of Parkinson's disease (PD), achieving acceptable diagnostic and prognostic accuracy will require the support of a panel of diverse biomarkers. We used Proximity extension assays to measure a panel of 92 proteins in CSF of 120 newly diagnosed PD patients and 45 control subjects without neurological disease. From 75 proteins detectable in the CSF of >90% of the subjects, regularized regression analysis identified four proteins (ß-NGF, CD38, tau and NCAN) as downregulated in newly diagnosed PD patients (age at diagnosis 67.2 ± 9.4 years) compared to controls (age 65.4 ± 10.9 years). Higher tau (ß -0.82 transformed MMSE points/year, 95% CI -1.37 to -0.27, P = 0.005) was also linked to faster cognitive decline over the first ten years after PD diagnosis. These findings provide insights into multiple aspects of PD pathophysiology and may serve as the foundation for identifying new biomarkers and therapeutic targets.

Seed Amplification Assay as a Diagnostic Tool in Newly-Diagnosed Parkinson's Disease.

Seed amplification assays (SAA) are the first credible molecular assay for Parkinson's disease (PD). However, the value of SAA to support the clinicians' initial diagnosis of PD is not clear. In our study, we analyzed cerebrospinal fluid samples from 121 PD patients recruited through population screening methods and taken within a median delay of 38 days from diagnosis and 51 normal controls without neurodegenerative disease. SAA yielded a sensitivity of 82.6% (95% CI, 74.7% - 88.9%) and specificity of 88.2% (95% CI, 76.1% - 95.6%). These results highlight the potential of SAA to support the initial diagnosis of PD in clinical practice and research.

Cognitive and Motor Decline in Dementia with Lewy Bodies and Parkinson's Disease Dementia.

Background: There is a need to better understand the rate of cognitive and motor decline of Dementia with Lewy bodies (DLB) and Parkinson's disease Dementia (PDD). Objectives: To compare the rate of cognitive and motor decline in patients with DLB and PDD from the E-DLB Consortium and the Parkinson's Incidence Cohorts Collaboration (PICC) Cohorts. Methods: The annual change in MMSE and MDS-UPDRS part III was estimated using linear mixed regression models in patients with at least one follow-up (DLB n = 837 and PDD n = 157). Results: When adjusting for confounders, we found no difference in the annual change in MMSE between DLB and PDD (-1.8 [95% CI -2.3, -1.3] vs. -1.9 [95% CI -2.6, -1.2] [P = 0.74]). MDS-UPDRS part III showed nearly identical annual changes (DLB 4.8 [95% CI 2.1, 7.5]) (PDD 4.8 [95% CI 2.7, 6.9], [P = 0.98]). Conclusions: DLB and PDD showed similar rates of cognitive and motor decline. This is relevant for future clinical trial designs.

Inflammatory Biomarkers in Newly Diagnosed Patients With Parkinson Disease and Related Neurodegenerative Disorders.

BACKGROUND AND OBJECTIVES: Neuroinflammation contributes to Parkinson disease (PD) pathology, and inflammatory biomarkers may aid in PD diagnosis. Proximity extension assay (PEA) technology is a promising method for multiplex analysis of inflammatory markers. Neuroinflammation also plays a role in related neurodegenerative diseases, such as dementia with Lewy bodies (DLB) and Alzheimer disease (AD). The aim of this work was to assess the value of inflammatory biomarkers in newly diagnosed patients with PD and in patients with DLB and AD. METHODS: Patients from the Norwegian ParkWest and Dementia Study of Western Norway longitudinal cohorts (PD, n = 120; DLB, n = 15; AD, n = 27) and 44 normal controls were included in this study. A PEA inflammation panel of 92 biomarkers was measured in the CSF. Disease-associated biomarkers were identified using elastic net (EN) analysis. We assessed the discriminatory power of disease-associated biomarkers using receiver operating characteristic (ROC) curve analysis and estimated the optimism-adjusted area under the curve (AUC) using the bootstrapping method. RESULTS: EN analysis identified 9 PEA inflammatory biomarkers (ADA, CCL23, CD5, CD8A, CDCP1, FGF-19, IL-18R1, IL-6, and MCP-2) associated with PD. Seven of the 9 biomarkers were included in a diagnostic panel, which was able to discriminate between those with PD and controls (optimism-adjusted AUC 0.82). Our 7-biomarker PD panel was also able to distinguish PD from DLB and from AD. In addition, 4 inflammatory biomarkers were associated with AD and included in a panel, which could distinguish those with AD from controls (optimism-adjusted AUC 0.87). Our 4-biomarker AD panel was also able to distinguish AD from DLB and from PD. DISCUSSION: In our exploratory study, we identified a 7-biomarker panel for PD and a 4-biomarker panel for AD. Our findings indicate potential inflammation-related biomarker candidates that could contribute toward PD-specific and AD-specific diagnostic panels, which should be further explored in other larger cohorts.

Association of CSF Glucocerebrosidase Activity With the Risk of Incident Dementia in Patients With Parkinson Disease.

BACKGROUND AND OBJECTIVES: Variations in the glucocerebrosidase gene (GBA) are common risk factors for Parkinson disease (PD) and dementia in PD (PDD) and cause a reduction in the activity of the lysosomal enzyme glucocerebrosidase (GCase). It is anticipated that GCase dysfunction might contribute to a more malignant disease course and predict cognitive impairment in PD, although evidence is lacking. We aimed to discover whether CSF GCase activity is altered in newly diagnosed patients with PD and associated with future development of dementia. METHODS: Patients with PD were participants of the ongoing population-based longitudinal ParkWest study in Southwestern Norway and were followed prospectively for up to 10 years. CSF was collected at diagnosis, and GBA carrier status was obtained. Control samples were from persons without neurodegenerative disorders. GCase activity was measured using a validated assay. PD dementia diagnosis was set according to the Movement Disorder Society criteria, and parametric accelerated failure time models were applied to analyze the association of GCase activity with dementia-free survival. RESULTS: This study enrolled 117 patients with PD (mean age 67.2 years, including 12 GBA non-synonymous variant carriers) and 50 control participants (mean age 64 years). At the time of diagnosis, GCase activity was reduced in patients with PD with (mean ± SD, 0.92 ± 0.40 mU/mg, n = 12) or without GBA variations (1.00 ± 0.37 mU/mg, n = 105) compared with controls (1.20 ± 0.35, n = 50). GCase activity at the time of diagnosis was lower in patients with PD who developed dementia within 10 years (0.85 ± 0.27 mU/mg, n = 41) than in those who did not (1.07 ± 0.40 mU/mg, n = 76, p = 0.001). A 0.1-unit reduction in baseline GCase activity was associated with a faster development of PDD (hazard ratio 1.15, 95% CI 1.03-1.28, p = 0.014). DISCUSSION: The association of early CSF GCase activity with long-term progression to PD dementia will have important implications for the design of clinical trials for GCase targeting therapies and patient management. CLASSIFICATION OF EVIDENCE: This study provides Class III evidence that reduced CSF GCase activity at the time of PD diagnosis is associated with an increased risk for later development of PDD.

Mitochondrial haplogroups and cognitive progression in Parkinson's disease.

Mitochondria are a culprit in the onset of Parkinson's disease, but their role during disease progression is unclear. Here we used Cox proportional hazards models to exam the effect of variation in the mitochondrial genome on longitudinal cognitive and motor progression over time in 4064 patients with Parkinson's disease. Mitochondrial macro-haplogroup was associated with reduced risk of cognitive disease progression in the discovery and replication population. In the combined analysis, patients with the super macro-haplogroup J, T, U# had a 41% lower risk of cognitive progression with P = 2.42 × 10-6 compared to those with macro-haplogroup H. Exploratory analysis indicated that the common mitochondrial DNA variant, m.2706A>G, was associated with slower cognitive decline with a hazard ratio of 0.68 (95% confidence interval 0.56-0.81) and P = 2.46 × 10-5. Mitochondrial haplogroups were not appreciably linked to motor progression. This initial genetic survival study of the mitochondrial genome suggests that mitochondrial haplogroups may be associated with the pace of cognitive progression in Parkinson's disease over time.

Parkinson's disease clinical milestones and mortality.

Identification of factors predicting and driving mortality in PD is important for patient information, disease management, and design of future clinical trials. This study included newly diagnosed PD patients and normal controls (NC) from a population-based study with repeated assessments over a 10-year period. We used the Kaplan-Meier method to estimate survival, Cox proportional hazards regression models to identify baseline risk factors of mortality, and Cox regression models with time-dependent covariates to evaluate the impact of four clinical milestones of advanced PD (visual hallucinations, recurrent falls, dementia, and nursing home placement) on mortality risk. During the 10-year study, 65 (34.2%) of 190 patients and 25 (12.3%) of 203 NC died, with an unadjusted hazard ratio (HR) of 2.85 (95% CI 1.80-4.52) and a HR of 2.48 (95% CI 1.55-3.95) when adjusted for confounders, including comorbidities. Higher age, more severe motor impairment, and postural instability-gait difficulty (PIGD) phenotype were independent baseline predictors of mortality. Each clinical milestone alone more than doubled the risk of death and had a cumulative effect on mortality, with a HR of 10.83 (95% CI 4.39-26.73) in those experiencing all four milestones. PD patients have an increased mortality risk that is disease-related and becomes evident early during the course of the disease. While motor severity and PIGD phenotype were early risk factors of mortality, clinical milestones signaled a substantially increased risk of death later during the disease course, highlighting their potential significance in clinical disease staging and prognosis.

The incidence, baseline predictors, and outcomes of dementia in an incident cohort of Parkinson's disease and controls.

BACKGROUND: There are few long-term data on the incidence, baseline predictors, and outcomes of dementia in Parkinson's disease (PD) from prospective community-based incident cohorts. METHODS: The PINE study prospectively identified all incident PD patients in Aberdeen along with age-sex-matched, community-based controls who consented to standardized annual life-long follow-up. Each year, a clinical expert reviewed the diagnosis of PD and the presence of dementia according to DSM-IV-based criteria. Age-sex stratified incidence rates for dementia in PD and controls were calculated and compared with hazard ratios (HR) adjusted for age, sex, education, and socioeconomic status. Cox proportional-hazard modelling was used to assess baseline predictors for PD dementia and the influence of dementia on survival and institutionalization. RESULTS: 201 patients (mean age 72.6yrs) and 260 controls (mean age 75.4yrs) were followed for median 9.5 years. The incidence of dementia was 7.4 (PD) versus 2.1 (controls) per 100 person-years (adjusted HR 6.0, 95%CI 4.1-8.7), with a sixfold increase from under 60 to over 80 years in PD but no sex difference. Independent baseline predictors of PD dementia were older age at diagnosis, self-reported cognitive symptoms, dream enactment, lower MMSE scores, worse motor UPDRS scores, and the ApoE genotype. PD dementia increased the rates of subsequent death and institutionalization (32.0 and 26.9 per 100 person-years, respectively). CONCLUSION: The incidence of dementia in PD is high, increases markedly with age, is increased in those with baseline subjective cognitive symptoms as well as other established risk factors, and is associated with high rates of death and institutionalization.

GBA and APOE Impact Cognitive Decline in Parkinson's Disease: A 10-Year Population-Based Study.

BACKGROUND: Common genetic variance in apolipoprotein E (APOE), ß-glucocerebrosidase (GBA), microtubule-associated protein tau (MAPT), and α-synuclein (SNCA) has been linked to cognitive decline in Parkinson's disease (PD), although studies have yielded mixed results. OBJECTIVES: To evaluate the effect of genetic variants in APOE, GBA, MAPT, and SNCA on cognitive decline and risk of dementia in a pooled analysis of six longitudinal, non-selective, population-based cohorts of newly diagnosed PD patients. METHODS: 1002 PD patients, followed for up to 10 years (median 7.2 years), were genotyped for at least one of APOE-ε4, GBA mutations, MAPT H1/H2, or SNCA rs356219. We evaluated the effect of genotype on the rate of cognitive decline (Mini-Mental State Examanation, MMSE) using linear mixed models and the development of dementia (diagnosed using standardized criteria) using Cox regression; multiple comparisons were accounted for using Benjamini-Hochberg corrections. RESULTS: Carriers of APOE-ε4 (n = 281, 29.7%) and GBA mutations (n = 100, 10.3%) had faster cognitive decline and were at higher risk of progression to dementia (APOE-ε4, HR 3.57, P < 0.001; GBA mutations, HR 1.76, P = 0.001) than non-carriers. The risk of cognitive decline and dementia (HR 5.19, P < 0.001) was further increased in carriers of both risk genotypes (n = 23). No significant effects were observed for MAPT or SNCA rs356219. CONCLUSIONS: GBA and APOE genotyping could improve the prediction of cognitive decline in PD, which is important to inform the clinical trial selection and potentially to enable personalized treatment © 2022 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.

Cannabis use in Parkinson's disease-A nationwide online survey study.

OBJECTIVES: The aim of this study was to investigate the frequency of use, attitudes toward, and experiences with cannabis and cannabis-related products among people with Parkinson's disease (PwP) living in Norway. METHODS: Between February and August 2021, PwP and their caregivers were invited to participate in an anonymous online survey study on cannabis use. N = 530 PwP completed the 24-item survey collecting data on the participants' history of cannabis use, perceived benefits and adverse effects of cannabis use, and expectations toward health care professionals. N = 108 caregivers completed a brief survey detailing their experience with cannabis use. RESULTS: A total of 59 (11.3%) of PwP reported previous or current use of cannabis, compared to 7 (6.6%) of caregivers. Cannabis use was associated with increased disease duration, but not age or gender. Improvement in motor function (69.5%), sleep (52.5%), and pain (37.3%) was the most frequently perceived benefits of cannabis use, with benefits more frequently reported by current than previous users. While half (50.8%) of cannabis users had sought advice from a health care professional regarding cannabis use, only 55 (19.9%) of non-users with an interest in cannabis use had discussed the topic with health care professionals. Principal barriers for discussing cannabis use with health care professionals are discussed. CONCLUSIONS: One in 20 PwP reports cannabis use, and non-users report widespread interest in cannabis. The use of cannabis is often not reported and unknown for health care professionals, arguing for a vigilant approach to non-prescribed cannabis use in clinical follow-up of PwP.

First delirium episode in Parkinson's disease and parkinsonism: incidence, predictors, and outcomes.

To define the incidence, predictors and prognosis of the first hospital delirium episode in Parkinson's disease (PD) and atypical parkinsonism (AP), we identified the first hospital episode of delirium after diagnosis in the Parkinsonism Incidence in North-East Scotland (PINE) study, a prospective community-based incidence cohort of parkinsonism, using chart-based criteria to define delirium. Of 296 patients (189=PD, 107=AP [dementia with Lewy bodies, progressive supranuclear palsy, multiple system atrophy, vascular parkinsonism]), 152 developed delirium (PD = 98, AP = 54). Incidence of first hospital delirium episode per 100 person years was 8.1 (95% confidence interval [CI] 6.6-9.9) in PD and 18.5 (95% CI 13.9-24.7) in AP. Independent predictors of delirium were atypical parkinsonism (Hazard ratio [HR] vs PD = 2.83 [95% CI 1.60-5.03], age in PD but not in AP (HR for 10-year increase 2.29 [95% CI 1.74-3.02]), baseline MMSE (HR = 0.94 [95% CI 0.89-0.99]), APOE ε4 in PD (HR 2.16 [95% CI 1.15-4.08]), and MAPT H1/H1 in PD (HR 2.08 [95% CI 1.08-4.00]). Hazards of dementia and death after delirium vs before delirium were increased (dementia: HR = 6.93 [95% CI 4.18-11.48] in parkinsonism; death: HR = 3.76 [95% CI 2.65-5.35] in PD, 1.59 [95% CI 1.04-2.42] in AP). Delirium is a common non-motor feature of PD and AP and is associated with increased hazards of dementia and mortality. Whether interventions for early identification and treatment improve outcomes requires investigation.

The impact of common genetic variants in cognitive decline in the first seven years of Parkinson's disease: A longitudinal observational study.

INTRODUCTION: Cognitive impairment is a common feature of Parkinson's disease and is a significant determinant of patients' quality of life and dependence. The pattern and progression of cognitive symptoms vary greatly between individuals, and genetic biomarkers may help to predict the severity and trajectory of cognitive impairment in groups of patients. METHODS: The study included 171 patients from a longitudinal population-based incident Parkinson's disease study from South Western Norway. All participants were followed from the time of diagnosis for up to seven years, undertaking repeated batteries of clinical and neuropsychological tests, measuring global cognitive impairment, executive function, attention, verbal learning and memory, and visuospatial skills. We used linear mixed regression analyses to explore associations between the function in specific cognitive domains over time and common genetic variants in APOE, MAPT, COMT and BDNF. RESULTS: The COMT158Val/Val allele wasassociatedwith faster decline in executive function (p = 0.028), verbal learning and memory (p = 0.029), and visuospatial skills (p = 0.027). The BDNF, MAPT and APOE genotypes were not significantly associated with longitudinal changes in individual cognitive domains, however carriers of the APOE-ε4 allele were shown to be at increased risk of mild cognitive impairment and dementia within the study period (OR3.03; p = 0.006). CONCLUSIONS: This population-based study of newly diagnosed patients provides new evidence that COMTVal158Met effects cognitive outcomes limited to discrete domains and APOE-ε4 status predicts a poor overall cognitive prognosis. Together, these data contribute to our understanding of the biology underlying the heterogeneity observed in the progression of PD.

Lack of Association Between GBA Mutations and Motor Complications in European and American Parkinson's Disease Cohorts.

BACKGROUND: Motor complications are a consequence of the chronic dopaminergic treatment of Parkinson's disease (PD) and include levodopa-induced dyskinesia (LIDs) and motor fluctuations (MF). Currently, evidence is on lacking whether patients with GBA-associated PD differ in their risk of developing motor complications compared to the general PD population. OBJECTIVE: To evaluate the association of GBA carrier status with the development of LIDS and MFs from early PD. METHODS: Motor complications were recorded prospectively in 884 patients with PD from four longitudinal cohorts using part IV of the UPDRS or MDS-UPDRS. Subjects were followed for up to 11 years and the associations of GBA mutations with the development of motor complications were assessed using parametric accelerated failure time models. RESULTS: In 439 patients from Europe, GBA mutations were detected in 53 (12.1%) patients and a total of 168 cases of LIDs and 258 cases of MF were observed. GBA carrier status was not associated with the time to develop LIDs (HR 0.78, 95%CI 0.47 to 1.26, p = 0.30) or MF (HR 1.19, 95%CI 0.84 to 1.70, p = 0.33). In the American cohorts, GBA mutations were detected in 36 (8.1%) patients and GBA carrier status was also not associated with the progression to LIDs (HR 1.08, 95%CI 0.55 to 2.14, p = 0.82) or MF (HR 1.22, 95%CI 0.74 to 2.04, p = 0.43). CONCLUSION: This study does not provide evidence that GBA-carrier status is associated with a higher risk of developing motor complications. Publication of studies with null results is vital to develop an accurate summary of the clinical features that impact patients with GBA-associated PD.

A systematic review of associations between common SNCA variants and clinical heterogeneity in Parkinson's disease.

There is great heterogeneity in both the clinical presentation and rate of disease progression among patients with Parkinson's disease (PD). This can pose prognostic difficulties in a clinical setting, and a greater understanding of the risk factors that contribute to modify disease course is of clear importance for optimizing patient care and clinical trial design. Genetic variants in SNCA are an established risk factor for PD and are candidates to modify disease presentation and progression. This systematic review aimed to summarize all available primary research reporting the association of SNCA polymorphisms with features of PD. We systematically searched PubMed and Web of Science, from inception to 1 June 2020, for studies evaluating the association of common SNCA variants with age at onset (AAO) or any clinical feature attributed to PD in patients with idiopathic PD. Fifty-eight studies were included in the review that investigated the association between SNCA polymorphisms and a broad range of outcomes, including motor and cognitive impairment, sleep disorders, mental health, hyposmia, or AAO. The most reproducible findings were with the REP1 polymorphism or rs356219 and an earlier AAO, but no clear associations were identified with an SNCA polymorphism and any individual clinical outcome. The results of this comprehensive summary suggest that, while there is evidence that genetic variance in the SNCA region may have a small impact on clinical outcomes in PD, the mechanisms underlying the association of SNCA polymorphisms with PD risk may not be a major factor driving clinical heterogeneity in PD.

Genome-wide survival study identifies a novel synaptic locus and polygenic score for cognitive progression in Parkinson's disease.

A key driver of patients' well-being and clinical trials for Parkinson's disease (PD) is the course that the disease takes over time (progression and prognosis). To assess how genetic variation influences the progression of PD over time to dementia, a major determinant for quality of life, we performed a longitudinal genome-wide survival study of 11.2 million variants in 3,821 patients with PD over 31,053 visits. We discover RIMS2 as a progression locus and confirm this in a replicate population (hazard ratio (HR) = 4.77, P = 2.78 × 10-11), identify suggestive evidence for TMEM108 (HR = 2.86, P = 2.09 × 10-8) and WWOX (HR = 2.12, P = 2.37 × 10-8) as progression loci, and confirm associations for GBA (HR = 1.93, P = 0.0002) and APOE (HR = 1.48, P = 0.001). Polygenic progression scores exhibit a substantial aggregate association with dementia risk, while polygenic susceptibility scores are not predictive. This study identifies a novel synaptic locus and polygenic score for cognitive disease progression in PD and proposes diverging genetic architectures of progression and susceptibility.

Association of GBA Genotype With Motor and Functional Decline in Patients With Newly Diagnosed Parkinson Disease.

OBJECTIVE: To establish the significance of glucocerebrosidase gene (GBA) carrier status on motor impairment in a large cohort of patients with incident Parkinson disease (PD). METHODS: Three European population-based studies followed 528 patients with PD from diagnosis. A total of 440 with genomic DNA from baseline were assessed for GBA variants. We evaluated motor and functional impairment annually using the Unified Parkinson's Disease Rating Scale (UPDRS) motor and activities of daily living (ADL) sections. Differential effects of classes of GBA variants on disease progression were evaluated using mixed random and fixed effects models. RESULTS: A total of 387 patients with idiopathic disease (age at baseline 70.3 ± 9.5 years; 60.2% male) and 53 GBA carriers (age at baseline 66.8 ± 10.1 years; 64.2% male) were included. The motor profile of the groups was clinically indistinguishable at diagnosis. GBA carriers showed faster annual increase in UPDRS scores measuring ADL (1.5 point per year, 95% confidence interval [CI] 1.1-2.0) and motor symptoms (2.2 points per year, 95% CI 1.3-3.1) compared to noncarriers (ADL, 1.0 point per year, 95% CI 0.9-1.1, p = 0.003; motor, 1.3 point per year, 95% CI 1.1-1.6, p = 0.007). Simulations of clinical trial designs showed that recruiting only GBA carriers can reduce trial size by up to 65% compared to a trial recruiting all patients with PD. CONCLUSION: GBA variants are linked to a more aggressive motor disease course over 7 years from diagnosis in patients with PD. A better understanding of PD progression in genetic subpopulations may improve disease management and has direct implications for improving the design of clinical trials.

Differences in the Presentation and Progression of Parkinson's Disease by Sex.

BACKGROUND: Previous studies reported various symptoms of Parkinson's disease (PD) associated with sex. Some were conflicting or confirmed in only one study. OBJECTIVES: We examined sex associations to PD phenotypes cross-sectionally and longitudinally in large-scale data. METHODS: We tested 40 clinical phenotypes, using longitudinal, clinic-based patient cohorts, consisting of 5946 patients, with a median follow-up of 3.1 years. For continuous outcomes, we used linear regressions at baseline to test sex-associated differences in presentation, and linear mixed-effects models to test sex-associated differences in progression. For binomial outcomes, we used logistic regression models at baseline and Cox regression models for survival analyses. We adjusted for age, disease duration, and medication use. In the secondary analyses, data from 17 719 PD patients and 7588 non-PD participants from an online-only, self-assessment PD cohort were cross-sectionally evaluated to determine whether the sex-associated differences identified in the primary analyses were consistent and unique to PD. RESULTS: Female PD patients had a higher risk of developing dyskinesia early during the follow-up period, with a slower progression in activities of daily living difficulties, and a lower risk of developing cognitive impairments compared with male patients. The findings in the longitudinal, clinic-based cohorts were mostly consistent with the results of the online-only cohort. CONCLUSIONS: We observed sex-associated contributions to PD heterogeneity. These results highlight the necessity of future research to determine the underlying mechanisms and importance of personalized clinical management. © 2020 International Parkinson and Movement Disorder Society.

Validation and assessment of preanalytical factors of a fluorometric in vitro assay for glucocerebrosidase activity in human cerebrospinal fluid.

Lysosomal dysfunction is an emerging feature in the pathology of Parkinson's disease and Dementia with Lewy bodies. Mutations in the GBA gene, encoding the enzyme Glucocerebrosidase (GCase), have been identified as a genetic risk factor for these synucleinopathies. As a result, there has been a growing interest in the involvement of GCase in these diseases. This GCase activity assay is based on the catalytic hydrolysis of 4-methylumbelliferyl ß-D-glucopyranoside that releases the highly fluorescent 4-methylumbelliferyl (4-MU). The final assay protocol was tested for the following parameters: Lower limit of quantification (LLOQ), precision, parallelism, linearity, spike recovery, number of freeze-thaw events, and sample handling stability. The GCase activity assay is within acceptable criteria for parallelism, precision and spike recovery. The LLOQ of this assay corresponds to an enzymatic activity of generating 0.26 pmol 4-MU/min/ml. The enzymatic activity was stable when samples were processed and frozen at - 80 °C within 4 h after the lumbar puncture procedure. Repetitive freeze-thaw events significantly decreased enzyme activity. We present the validation of an optimized in vitro GCase activity assay, based on commercially available components, to quantify its enzymatic activity in human cerebrospinal fluid and the assessment of preanalytical factors.

Plasma-derived exosome-like vesicles are enriched in lyso-phospholipids and pass the blood-brain barrier.

Exosomes are vesicles involved in intercellular communication. Their membrane structure and core content is largely dependent on the cell of origin. Exosomes have been investigated both for their biological roles and their possible use as disease biomarkers and drug carriers. These potential technological applications require the rigorous characterization of exosomal blood brain barrier permeability and a description of their lipid bilayer composition. To achieve these goals, we have established a 3D static blood brain barrier system based on existing systems for liposomes and a complementary LC-MS/MS and 31P nuclear magnetic resonance methodology for the analysis of purified human plasma-derived exosome-like vesicles. Results show that the isolated vesicles pass the blood brain barrier and are taken up in endothelial cells. The compositional analysis revealed that the isolated vesicles are enriched in lyso phospholipids and do not contain phosphatidylserine. These findings deviate significantly from the composition of exosomes originating from cell culture, and may reflect active removal by macrophages that respond to exposed phosphahtidylserine.