Parkinson's Disease Brain Tissue Research Articles

Discovery and Evaluation of Imidazo[2,1-b][1,3,4]thiadiazole Derivatives as New Candidates for α-Synuclein PET Imaging.

α-synuclein (α-syn) pathologies are central to the development of synucleinopathies including Parkinson's disease (PD). Positron emission tomography (PET) imaging of α-syn pathologies is one strategy to facilitate the diagnosis, understanding, and treatment of synucleinopathies, but has been restricted by the lack of specific α-syn PET probes. In this work, we identified 2,6-disubstituted imidazo[2,1-b][1,3,4]thiadiazole (ITA) as a new α-syn-binding scaffold. Through autoradiography studies, we discovered an iodinated lead compound [125I]ITA-3, with moderate binding affinity (IC50 = 55 nM) to α-syn pathologies in human PD brain sections. Modified from [125I]ITA-3, we developed a potential PET tracer, [18F]FITA-2 (radiochemical yield >25%, molar activity >110 GBq/μmol), which demonstrated clear signals in α-syn-rich regions in human PD brain tissues (IC50 = 245 nM), good brain uptake (SUVpeak = 2.80 ± 0.45), and fast clearance rate in rats. Overall, [18F]FITA-2 appears to be a promising candidate for α-syn PET imaging and merits further development.

EEF1A1 regulates the expression and alternative splicing of genes associated with Parkinson's disease in U251 cells.

Eukaryotic elongation factor 1A1 (eEF1A1) is an RNA-binding protein that is associated with PARK2 activity in cells, suggesting a possible role in Parkinson's disease (PD). To clear whether eEF1A1 plays a role in PD through transcriptional or posttranscriptional regulation. The GSE68719 dataset was downloaded from the GEO database, and the RNA-seq data of all brain tissue autopsies were obtained from 29 PD patients and 44 neurologically normal control subjects. To inhibit eEF1A1 from being expressed in U251 cells, siRNA was transfected into those cells, and RNA-seq high-throughput sequencing was used to determine the differentially expressed genes (DEGs) and differentially alternative splicing events (ASEs) resulting from eEF1A1 knockdown. eEF1A1 was significantly overexpressed in PD brain tissue in the BA9 area. GO and KEGG enrichment analyses revealed that eEF1A1 knockdown significantly upregulated the expression of the genes CXCL10, NGF, PTX3, IL6, ST6GALNAC3, NUPR1, TNFRSF21, and CXCL2 and upregulated the alternative splicing of the genes ACOT7, DDX10, SHMT2, MYEF2, and NDUFAF5. These genes were enriched in pathways related to PD pathogenesis, such as apoptosis, inflammatory response, and mitochondrial dysfunction. The results suggesting that eEF1A1 involved in the development of PD by regulating the differential expression and alternative splicing of genes, providing a theoretical basis for subsequent research.

Molecular insights into PCB neurotoxicity: Comparing transcriptomic responses across dopaminergic neurons, population blood cells, and Parkinson's disease pathology

Parkinson's disease (PD) is a complex neurodegenerative disorder influenced by genetic factors and environmental exposures. Polychlorinated biphenyls (PCBs), a group of synthetic organic compounds, have been identified as potential environmental risk factors for neurodegenerative diseases, including PD.We explored PCB-induced neurotoxicity mechanisms using iPSC-derived dopaminergic neurons and assessed their transcriptomic responses to varying PCB concentrations (0.01 μM, 0.5 μM, and 10 μM). Specifically, we focused on PCB-180, a congener known for its accumulation in human brains. The exposure durations were 24 h and 74 h, allowing us to capture both short-term and more prolonged effects on gene expression patterns. We observed that PCB exposure led to the suppression of oxidative phosphorylation, synaptic function, and neurotransmitter release, implicating these pathways in PCB-induced neurotoxicity.In our comparative analysis, we noted similarities in PCB-induced changes with other PD-related compounds like MPP+ and rotenone. Our findings also aligned with gene expression changes in human blood derived from a population exposed to PCBs, highlighting broader inflammatory responses. Additionally, molecular patterns seen in iPSC-derived neurons were confirmed in postmortem PD brain tissues, validating our in vitro results.In conclusion, our study offers novel insights into the multifaceted impacts of PCB-induced perturbations on various cellular contexts relevant to PD. The use of iPSC-derived dopaminergic neurons allowed us to decipher intricate transcriptomic alterations, bridging the gap between in vitro and in vivo findings. This work underscores the potential role of PCB exposure in neurodegenerative diseases like PD, emphasizing the need to consider both systemic and cell specific effects.

Revealing Novel Genes Related to Parkinson's Disease Pathogenesis and Establishing an associated Model

Parkinson's disease (PD) represents a multifaceted neurological disorder whose genetic underpinnings warrant comprehensive investigation. This study focuses on identifying genes integral to PD pathogenesis and evaluating their diagnostic potential. Initially, we screened for differentially expressed genes (DEGs) between PD and control brain tissues within a dataset comprising larger number of specimens. Subsequently, these DEGs were subjected to weighted gene co-expression network analysis (WGCNA) to discern relevant gene modules. Notably, the yellow module exhibited a significant correlation with PD pathogenesis. Hence, we conducted a detailed examination of the yellow module genes using a cytoscope-based approach to construct a protein–protein interaction (PPI) network, which facilitated the identification of central hub genes implicated in PD pathogenesis. Employing two machine learning techniques, including XGBoost and LASSO algorithms, along with logistic regression analysis, we refined our search to three pertinent hub genes: FOXO3, HIST2H2BE, and HDAC1, all of which demonstrated a substantial association with PD pathogenesis. To corroborate our findings, we analyzed two PD blood datasets and clinical plasma samples, confirming the elevated expression levels of these genes in PD patients. The association of the genes with PD, as reflected by the area under the curve (AUC) values for FOXO3, HIST2H2BE, and HDAC1, were moderate for each gene. Collectively, this research substantiates the heightened expression of FOXO3, HIST2H2BE, and HDAC1 in both PD brain and blood samples, underscoring their pivotal contribution to the pathogenesis of PD.

Identification of SV2C and DENR as Key Biomarkers for Parkinson's Disease Based on Bioinformatics, Machine Learning, and Experimental Verification.

The objective of this study is to investigate the potential biomarkers and therapeutic target genes for Parkinson's disease (PD). We analyzed four datasets (GSE8397, GSE20292, GSE20163, GSE20164) from the Gene Expression Omnibus database. We employed weighted gene co-expression network analysis and differential expression analysis to select genes and perform functional analysis. We applied three algorithms, namely, random forest, support vector machine recursive feature elimination, and least absolute shrinkage and selection operator, to identify hub genes, perform functional analysis, and assess their clinical diagnostic potential using receiver operating characteristic (ROC) curve analysis. We employed the xCell website to evaluate differences in the composition patterns of immune cells in the GEO datasets. We also collected serum samples from PD patients and established PD cell model to validate the expression of hub genes using enzyme-linked immunosorbent assay and quantitative real-time polymerase chain reaction. Our findings identified SV2C and DENR as two hub genes for PD and decreased in PD brain tissue compared with controls. ROC analysis showed effectively value of SV2C and DENR to diagnose PD, and they were downregulated in the serum of PD patients and cell model. Functional analysis revealed that dopamine vesicle transport and synaptic vesicle recycling are crucial pathways in PD. Besides, the differences in the composition of immune cells, especially basophils and T cells, were discovered between PD and controls. In summary, our study identifies SV2C and DENR as potential biomarkers for diagnosing PD and provides a new perspective for exploring the molecular mechanisms of PD.

Endonuclease VIII-like 1 deficiency potentiates nigrostriatal dopaminergic neuron degeneration in a male mouse model of Parkinson's disease.

Parkinson's disease (PD) is a common movement disorder caused by a characteristic loss of dopaminergic neurons in the substantia nigra and degeneration of dopamine terminals in the dorsal striatum. Previous studies have suggested that oxidative stress-induced DNA damage may be involved in PD pathogenesis, as steady-state levels of several types of oxidized nucleobases were shown to be elevated in PD brain tissues. These DNA lesions are normally removed from the genome by base excision repair, which is initiated by DNA glycosylase enzymes such as endonuclease VIII-like 1 (Neil1). In this study, we show that Neil1 plays an important role in limiting oxidative stress-induced degeneration of dopaminergic neurons. In particular, Neil1-deficient male mice exhibited enhanced sensitivity to nigrostriatal degeneration after 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) administration, and Neil1-deficient animals had higher levels of γH2AX-marked DNA damage than wild-type (WT) controls, regardless of treatment status. Moreover, MPTP-treated Neil1-/- male mice had slightly elevated expression of genes related to the nuclear factor erythroid 2-related factor 2 (Nrf2)-dependent antioxidant pathway. Treatment with the Nrf2 activator, monomethyl fumarate, reduced PD-like behaviors and pathology in Neil1-/- male mice, suggesting that Neil1 is an important defense molecule in an oxidative cellular environment. Taken together, these results suggest that Neil1 DNA glycosylase may play an important role in limiting oxidative stress-mediated PD pathogenesis.

De Novo Design of a Robust Fluorescent Probe for Basal HClO Imaging in a Mouse Parkinson's Disease Model.

Elevated HClO gets involved in the pathogenesis of Parkinson's disease (PD). Herein, a novel fluorescent probe NUU-1 was designed and synthesized. Distinct from the general strategies, NUU-1 features two distinct HClO reactive sites, a HClO-specific reaction site and a HClO-nonspecific reactive site, which in turn endows NUU-1 with the "0 + 1 > 1" amplification effect, that thus dramatically promotes the selectivity. NUU-1 displayed a fast response rate (within 15 s), remarkable fluorescence enhancement (about 538-fold), and excellent sensitivity (LOD = 25.8 nM) in response to HClO while the remaining fluorescence silence toward other common ROS (H2O2, •OH, ONOO-, O2•-, and 1O2) even at high concentrations (up to 0.5 mM). NUU-1 allows for the imaging of both exogenous and endogenous HClO in living dopaminergic cells (SH-SY5Y). Moreover, by employing NUU-1 as the probe, the image of HClO in C. elegans and zebrafish was successfully achieved. Significantly, in the first trial, NUU-1 was successfully utilized for the brain basal HClO imaging in PD mice models and distinguished PD brain tissues from normal control, thereby holding great potential for in-depth biological applications.

Analysis of neuronal iron deposits in Parkinson's disease brain tissue by synchrotron x-ray spectromicroscopy

BackgroundNeuromelanin-pigmented neurons, which are highly susceptible to neurodegeneration in the Parkinson’s disease substantia nigra, harbour elevated iron levels in the diseased state. Whilst it is widely believed that neuronal iron is stored in an inert, ferric form, perturbations to normal metal homeostasis could potentially generate more reactive forms of iron capable of stimulating toxicity and cell death. However, non-disruptive analysis of brain metals is inherently challenging, since use of stains or chemical fixatives, for example, can significantly influence metal ion distributions and/or concentrations in tissues. AimsThe aim of this study was to apply synchrotron soft x-ray spectromicroscopy to the characterisation of iron deposits and their local environment within neuromelanin-containing neurons of Parkinson’s disease substantia nigra. MethodsSoft x-ray spectromicroscopy was applied in the form of Scanning Transmission X-ray Microscopy (STXM) to analyse resin-embedded tissue, without requirement for chemically disruptive processing or staining. Measurements were performed at the oxygen and iron K-edges in order to characterise both organic and inorganic components of anatomical tissue using a single label-free method. ResultsSTXM revealed evidence for mixed oxidation states of neuronal iron deposits associated with neuromelanin clusters in Parkinson’s disease substantia nigra. The excellent sensitivity, specificity and spatial resolution of these STXM measurements showed that the iron oxidation state varies across sub-micron length scales. ConclusionsThe label-free STXM approach is highly suited to characterising the distributions of both inorganic and organic components of anatomical tissue, and provides a proof-of-concept for investigating trace metal speciation within Parkinson’s disease neuromelanin-containing neurons.

Simultaneous structural and elemental nano-imaging of human brain tissue.

Examining chemical and structural characteristics of micro-features in complex tissue matrices is essential for understanding biological systems. Advances in multimodal chemical and structural imaging using synchrotron radiation have overcome many issues in correlative imaging, enabling the characterization of distinct microfeatures at nanoscale resolution in ex vivo tissues. We present a nanoscale imaging method that pairs X-ray ptychography and X-ray fluorescence microscopy (XFM) to simultaneously examine structural features and quantify elemental content of microfeatures in complex ex vivo tissues. We examined the neuropathological microfeatures Lewy bodies, aggregations of superoxide dismutase 1 (SOD1) and neuromelanin in human post-mortem Parkinson's disease tissue. Although biometals play essential roles in normal neuronal biochemistry, their dyshomeostasis is implicated in Parkinson's disease aetiology. Here we show that Lewy bodies and SOD1 aggregates have distinct elemental fingerprints yet are similar in structure, whilst neuromelanin exhibits different elemental composition and a distinct, disordered structure. The unique approach we describe is applicable to the structural and chemical characterization of a wide range of complex biological tissues at previously unprecedented levels of detail.

Kinetic Modeling of pH-Dependent Oxidation of Dopamine by Iron and Its Relevance to Parkinson's Disease.

Parkinson's disease is the second most common neurodegenerative disease. While age is the most significant risk factor, the exact cause of this disease and the most effective approaches to mitigation remain unclear. It has long been proposed that dopamine may play a role in the pathology of Parkinson's disease in view of its ability to generate both protein-modifying quinones such as aminochrome and reactive oxygen species, especially in the presence of pathological iron accumulation in the primary site of neuron loss. Given the clinically measured acidosis of post-mortem Parkinson's disease brain tissue, the interaction between dopamine and iron was investigated over a pH range of 7.4 to 6.5 with emphasis on the accumulation of toxic quinones and generation of reactive oxygen species. Our results show that the presence of iron accelerates the formation of aminochrome with ferrous iron (Fe[II]) being more efficient in this regard than ferric iron (Fe[III]). Our results further suggest that a reduced aminochrome rearrangement rate coupled with an enhanced turnover rate of Fe[II] as a result of brain tissue acidosis could result in aminochrome accumulation within cells. Additionally, under these conditions, the enhanced redox cycling of iron in the presence of dopamine aggravates oxidative stress as a result of the production of damaging reactive species, including hydroxyl radicals.

Development of an aggregate-selective, human-derived α-synuclein antibody BIIB054 that ameliorates disease phenotypes in Parkinson's disease models

Aggregation of α-synuclein (α-syn) is neuropathologically and genetically linked to Parkinson's disease (PD). Since stereotypic cell-to-cell spreading of α-syn pathology is believed to contribute to disease progression, immunotherapy with antibodies directed against α-syn is considered a promising therapeutic approach for slowing disease progression. Here we report the identification, binding characteristics, and efficacy in PD mouse models of the human-derived α-syn antibody BIIB054, which is currently under investigation in a Phase 2 clinical trial for PD. BIIB054 was generated by screening human memory B-cell libraries from healthy elderly individuals. Epitope mapping studies conducted using peptide scanning, X-ray crystallography, and mutagenesis show that BIIB054 binds to α-syn residues 1–10. BIIB054 is highly selective for aggregated forms of α-syn with at least an 800-fold higher apparent affinity for fibrillar versus monomeric recombinant α-syn and a strong preference for human PD brain tissue. BIIB054 discriminates between monomers and oligomeric/fibrillar forms of α-syn based on high avidity for aggregates, driven by weak monovalent affinity and fast binding kinetics. In efficacy studies in three different mouse models with intracerebrally inoculated preformed α-syn fibrils, BIIB054 treatment attenuated the spreading of α-syn pathology, rescued motor impairments, and reduced the loss of dopamine transporter density in dopaminergic terminals in striatum. The preclinical data reported here provide a compelling rationale for clinical development of BIIB054 for the treatment and prevention of PD.

Path mediation analysis reveals GBA impacts Lewy body disease status by increasing α-synuclein levels

Synucleinopathies including Parkinson's disease (PD) and Dementia with Lewy bodies (DLB) are characterized by the accumulation of abnormal α-synuclein in intraneuronal inclusions, named Lewy bodies. Mutations in GBA1, the gene encoding the lysosomal hydrolase glucocerebrosidase, have been identified as the most common genetic risk factor for PD and DLB. However, despite extensive research, the mechanism by which glucocerebrosidase dysfunction increases the risk for PD or DLB still remains elusive.In our study we expand the toolbox for PD-DLB post-mortem studies by introducing new quantitative biochemical assays for glucocerebrosidase and α-synuclein. Applying causal modelling, we determine how these parameters are interrelated and ultimately impact disease manifestation.We developed quantitative immuno-based assays for glucocerebrosidase and α-synuclein (total and phosphorylated at Serine 129) protein levels, as well as a liquid chromatography–mass spectrometry method for the detection of the glucocerebrosidase lipid substrate glucosylsphingosine. These assays were applied on tissue samples from frontal cortex, putamen and substantia nigra of PD (n = 15) and DLB (n = 15) patients and age-matched non-demented controls (n = 15).Our results confirm elevated p-129 over total α-synuclein levels in the insoluble fraction of PD and DLB post-mortem brain tissue and we found significantly increased α-synuclein levels in the soluble fractions in PD and DLB. Furthermore, we identified an inverse correlation between reduced glucocerebrosidase enzyme activity and protein levels with increased glucosylsphingosine levels. In the substantia nigra, a brain region particularly vulnerable in Parkinson's disease, we found a significant correlation between glucocerebrosidase protein reduction and increased p129/total α-synuclein ratios.We assessed the direction and strength of the interrelation between all measured parameters by confirmatory path analysis. Interestingly, we found that glucocerebrosidase dysfunction impacts the PD-DLB status by increasing α-synuclein ratios in the substantia nigra, which was partly mediated by increasing glucosylsphingosine levels.In conclusion, we show that the introduced immuno-based assays enable the quantitative assessment of glucocerebrosidase and α-synuclein parameters in post-mortem brain. In the substantia nigra, reduced glucocerebrosidase levels contribute to the increase in α-synuclein levels and to PD-DLB disease manifestation partly by increasing its glycolipid substrate glucosylsphingosine. This interrelation between glucocerebrosidase, glucosylsphingosine and α-synuclein parameters supports the hypothesis that glucocerebrosidase acts as a modulator of PD-DLB.

Th17 Lymphocytes Induce Neuronal Cell Death in a Human iPSC-Based Model of Parkinson's Disease.

Parkinson's disease (PD) is a neurodegenerative disorder characterized by the progressive degeneration of midbrain neurons (MBNs). Recent evidence suggests contribution of the adaptive immune system in PD. Here, we show a role for human T lymphocytes as cell death inducers of induced pluripotent stem cell (iPSC)-derived MBNs in sporadic PD. Higher Th17 frequencies were found in the blood of PD patients and increased numbers of T lymphocytes were detected in postmortem PD brain tissues. We modeled this finding using autologous co-cultures of activated T lymphocytes and iPSC-derived MBNs of sporadic PD patients and controls. After co-culture with T lymphocytes or the addition of IL-17, PD iPSC-derived MBNs underwent increased neuronal death driven by upregulation of IL-17 receptor (IL-17R) and NFκB activation. Blockage of IL-17 or IL-17R, or the addition of the FDA-approved anti-IL-17 antibody, secukinumab, rescued the neuronal death. Our findings indicate a critical role for IL-17-producing T lymphocytes in sporadic PD.

The lysosomal enzyme alpha-Galactosidase A is deficient in Parkinson's disease brain in association with the pathologic accumulation of alpha-synuclein

The aberrant accumulation of alpha-synuclein (α-syn) is believed to contribute to the onset and pathogenesis of Parkinson's disease (PD). The autophagy-lysosome pathway (ALP) is responsible for the high capacity clearance of α-syn. ALP dysfunction is documented in PD and pre-clinical evidence suggests that inhibiting the ALP promotes the pathological accumulation of α-syn. We previously identified the pathological accumulation of α-syn in the brains of mice deficient for the soluble lysosomal enzyme alpha-Galactosidase A (α-Gal A), a member of the glycosphingolipid metabolism pathway. In the present study, we quantified α-Gal A activity and levels of its glycosphingolipid metabolites in postmortem temporal cortex specimens from control individuals and in PD individuals staged with respect to α-syn containing Lewy body pathology. In late-state PD temporal cortex we observed significant decreases in α-Gal A activity and the 46kDa “active” species of α-Gal A as determined respectively by fluorometric activity assay and western blot analysis. These decreases in α-Gal A activity/levels correlated significantly with increased α-syn phosphorylated at serine 129 (p129S-α-syn) that was maximal in late-stage PD temporal cortex. Mass spectrometric analysis of 29 different isoforms of globotriaosylceramide (Gb3), a substrate of α-Gal A indicated no significant differences with respect to different stages of PD temporal cortex. However, significant correlations were observed between increased levels of several Gb3 isoforms and with decreased α-Gal A activity and/or increased p129S-α-syn. Deacylated Gb3 (globotriaosylsphingosine or lyso-Gb3) was also analyzed in PD brain tissue but was below the limit of detection of 20pmol/g. Analysis of other lysosomal enzymes revealed a significant decrease in activity for the lysosomal aspartic acid protease cathepsin D but not for glucocerebrosidase (GCase) or cathepsin B in late-stage PD temporal cortex. However, a significant correlation was observed between decreasing GCase activity and increasing p129S-α-syn. Together our findings indicate α-Gal A deficiency in late-stage PD brain that correlates significantly with the pathological accumulation of α-syn, and further suggest the potential for α-Gal A and its glycosphingolipid substrates as putative biomarkers for PD.

Implications of DNA Methylation in Parkinson's Disease.

It has been 200 years since Parkinson’s disease (PD) was first described, yet many aspects of its etiopathogenesis remain unclear. PD is a progressive and complex neurodegenerative disorder caused by genetic and environmental factors including aging, nutrition, pesticides and exposure to heavy metals. DNA methylation may be altered in response to some of these factors; therefore, it is proposed that epigenetic mechanisms, particularly DNA methylation, can have a fundamental role in gene–environment interactions that are related with PD. Epigenetic changes in PD-associated genes are now widely studied in different populations, to discover the mechanisms that contribute to disease development and identify novel biomarkers for early diagnosis and future pharmacological treatment. While initial studies sought to find associations between promoter DNA methylation and the regulation of associated genes in PD brain tissue, more recent studies have described concordant DNA methylation patterns between blood and brain tissue DNA. These data justify the use of peripheral blood samples instead of brain tissue for epigenetic studies. Here, we summarize the current data about DNA methylation changes in PD and discuss the potential of DNA methylation as a potential biomarker for PD. Additionally, we discuss environmental and nutritional factors that have been implicated in DNA methylation. Although the search for significant DNA methylation changes and gene expression analyses of PD-associated genes have yielded inconsistent and contradictory results, epigenetic modifications remain under investigation for their potential to reveal the link between environmental risk factors and the development of PD.

MicroRNAs in Parkinson's disease and emerging therapeutic targets

Parkinson's disease (PD) is the second most common age-related neurodegenerative disorder, with the clinical main symptoms caused by a loss of dopaminergic neurons in the substantia nigra, corpus striatum and brain cortex. Over 90% of patients with PD have sporadic PD and occur in people with no known family history of the disorder. Currently there is no cure for PD. Treatment with medications to increase dopamine relieves the symptoms but does not slow down or reverse the damage to neurons in the brain. Increasing evidence points to inflammation as a chief mediator of PD with inflammatory response mechanisms, involving microglia and leukocytes, activated following loss of dopaminergic neurons. Oxidative stress is also recognized as one of the main causes of PD, and excessive reactive oxygen species (ROS) and reactive nitrogen species can lead to dopaminergic neuron vulnerability and eventual death. MicroRNAs control a range of physiological and pathological functions, and may serve as potential targets for intervention against PD to mitigate damage to the brain. Several studies have demonstrated that microRNAs can regulate oxidative stress and prevent ROS-mediated damage to dopaminergic neurons, suggesting that specific microRNAs may be putative targets for novel therapeutic strategies in PD. Recent human and animal studies have identified a large number of dysregulated microRNAs in PD brain tissue samples, many of which were downregulated. The dysregulated microRNAs affect downstream targets such as SNCA, PARK2, LRRK2, TNFSF13B, LTA, SLC5A3, PSMB2, GSR, GBA, LAMP-2A, HSC. Apart from one study, none of the studies reviewed had used agomirs or antagomirs to reverse the levels of downregulated or upregulated microRNAs, respectively, in mouse models of PD or with isolated human or mouse dopaminergic cells. Further large-scale studies of brain tissue samples collected with short postmortem interval from human PD patients are warranted to provide more information on the microRNA profiles in different brain regions and to test for gender differences.

Tandem Mass Spectrometry Multiplex Analysis of Glucosylceramide and Galactosylceramide Isoforms in Brain Tissues at Different Stages of Parkinson Disease.

Previous studies demonstrated that Parkinson disease (PD) is associated with a decreased activity of the glucocerebrosidase (GCase) enzyme in brain tissues. The objective of this study was to determine if GCase deficiency is associated with the accumulation of its glucosylceramide (GluCer) substrate in PD brain tissues. An ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method was developed, optimized, and validated for the multiplex analysis of GluCer isoforms (C18:0, C20:0, C22:0, C24:1, and C24:0) in brain tissue samples. These molecules were chromatographically separated from their isobaric galactosylceramide (GalCer) counterparts using normal phase chromatography. The analysis was performed by tandem mass spectrometry in the multiple reaction monitoring (MRM) acquisition mode. Limits of detection ranging from 0.4 to 1.1 nmol/g brain tissue were established for the different GluCer isoforms analyzed. For the first time, GluCer isoform levels were analyzed in temporal cortex brain tissue samples from 26 PD patients who were divided into three PD disease stages (IIa, III, and IV) according to the Unified Staging System for Lewy Body Disorders. These specimens were compared with brain tissue samples from 12 controls and 6 patients with Incidental Lewy Body Disease. No significant GluCer concentration differences were observed between the 5 sample groups. The GluCer isoform levels were also normalized with their matching GalCer isoforms. The normalized results showed a trend for GluCer levels which increased with PD severity. However, the differences observed between the groups were not significant, owing likely to the high standard deviations measured.

Oligomeric α-synuclein and β-amyloid variants as potential biomarkers for Parkinson's and Alzheimer's diseases.

Oligomeric forms of α-synuclein and β-amyloid are toxic protein variants that are thought to contribute to the onset and progression of Parkinson's disease (PD) and Alzheimer's disease (AD), respectively. The detection of toxic variants in human cerebrospinal fluid (CSF) and blood has great promise for facilitating early and accurate diagnoses of these devastating diseases. Two hurdles that have impeded the use of these protein variants as biomarkers are the availability of reagents that can bind the different variants and a sensitive assay to detect their very low concentrations. We previously isolated antibody-based reagents that selectively bind two different oligomeric variants of α-synuclein and two of β-amyloid, and developed a phage-based capture enzyme-linked immunosorbent assay (ELISA) with subfemtomolar sensitivity to quantify their presence. Here, we used these reagents to show that these oligomeric α-synuclein variants are preferentially present in PD brain tissue, CSF and serum, and that the oligomeric β-amyloid variants are preferentially present in AD brain tissue, CSF, and serum. Some AD samples also had α-synuclein pathology and some PD samples also had β-amyloid pathology, and, very intriguingly, these PD cases also had a history of dementia. Detection of different oligomeric α-synuclein and β-amyloid species is an effective method for identifying tissue, CSF and sera from PD and AD samples, respectively, and samples that also contained early stages of other protein pathologies, indicating their potential value as blood-based biomarkers for neurodegenerative diseases.

Identifying allele specific expression in Parkinson's disease brain tissue

Identifying allele specific expression in Parkinson's disease brain tissue

Alpha-synuclein transcript isoforms in three different brain regions from Parkinson's disease and healthy subjects in relation to the SNCA rs356165/rs11931074 polymorphisms

Mutations in the alpha-synuclein (SNCA) gene cause autosomal dominant Parkinson's disease (PD). Common SNCA polymorphisms have been associated with the risk of developing PD. Abnormal expression and post-translational modification of SNCA has been found in PD-brains. In addition to a full length transcript (SNCA-140) there are three short isoforms (SNCA-98, -112, and -126) that could be prone to aggregation. The association between SNCA polymorphisms and PD could be explained through an increased expression of these alternative transcripts. Our aim was to measure the different SNCA transcripts in the substantia nigra (SN), cerebellum (CB), and occipital cortex (OC) from PD-patients (n=9) and healthy subjects (n=6). In addition, we determined whether two SNCA polymorphisms (SNPs rs356165 and rs11931074) were related to differences in transcript isoform expression. PD brain tissues showed higher levels of the three short transcripts in the SN, but only SNCA-112 and SNCA-98 were significantly increased in the CB of patients vs. controls (p=0.02, p=0.03). The genotyping of a large cohort of PD-patients and controls showed that haplotype rs356165-A+rs11931074-G had a protective effect (OR=0.71; CI=0.59–0.83), while the G-T haplotype increased the risk for PD (OR=1.44; CI=1.06–1.96). We did not find significant differences for the SNCA levels between the haplotypes. In conclusion, we found statistically significant higher levels of the SNCA-112 and SNCA-98 transcripts in the CB of PD brains, and a trend toward higher levels of the short transcript isoforms in the SN of PD brains.