Project description:In Parkinson's disease (PD) there is a selective degeneration of neuromelanin-containing neurons, especially substantia nigra dopaminergic neurons. In humans, neuromelanin accumulates with age, the latter being the main risk factor for PD. The contribution of neuromelanin to PD pathogenesis remains unknown because, unlike humans, common laboratory animals lack neuromelanin. Synthesis of peripheral melanins is mediated by tyrosinase, an enzyme also present at low levels in the brain. Here we report that overexpression of human tyrosinase in rat substantia nigra results in age-dependent production of human-like neuromelanin within nigral dopaminergic neurons, up to levels reached in elderly humans. In these animals, intracellular neuromelanin accumulation above a specific threshold is associated to an age-dependent PD phenotype, including hypokinesia, Lewy body-like formation and nigrostriatal neurodegeneration. Enhancing lysosomal proteostasis reduces intracellular neuromelanin and prevents neurodegeneration in tyrosinase-overexpressing animals. Our results suggest that intracellular neuromelanin levels may set the threshold for the initiation of PD.
Project description:Neuromelanin is present in the cathecolaminergic neuron cells of the substantia nigra and locus coeruleus of the midbrain of primates. Neuromelanin plays a role in Parkinson's disease (PD). Literature reports that neuromelanin features, among others, antioxidant properties by metal ion chelation and free radical scavenging. The pigment has been reported to have prooxidant properties too, in certain experimental conditions. We propose an explorative electrochemical study of the effect of the presence of metal ions and reactive oxygen species (ROS) on the cyclic voltammograms of a synthetic model of neuromelanin. Our work improves the current understanding on experimental conditions where neuromelanin plays an antioxidant or prooxidant behavior, thus possibly contributing to shed light on factors promoting the appearance of PD.
Project description:Background:Recent developments in magnetic resonance imaging (MRI) techniques have offered new research opportunities to visualize in vivo substantia nigra pathology in Parkinson's disease (PD). This paper summarizes the main findings of nigrosome imaging and neuromelanin sensitive MRI studies in patients with PD and other parkinsonisms. Methods:The PubMed database was searched from 2005 to 2017 using the following keywords: Parkinson's disease and parkinsonism, in combination with MRI, nigrosome, neuromelanin, and iron. Only publications in English were included. Results:Nigrosome or dorsal nigral hyperintensity abnormalities are studied using T2* and susceptibility weighted imaging MRI sequences in most studies, whereas Neuromelanin imaging is usually performed using T1-weighted fast spin echo sequence. Nigrosome abnormalities have been consistently demonstrated in PD patients, and nigrosome imaging has high sensitivity and specificity in distinguishing PD from healthy controls, though it is unable to reliably separate PD from atypical parkinsonisms. Reduced neuromelanin-related signals and/or volume loss in neuromelanin containing structures have been found in PD patients, and neuromelanin sensitive MRI imaging can also discriminate PD patients from healthy controls with high accuracy, though there is a degree of heterogeneity in the imaging findings. Preliminary findings suggested that longitudinal change of neuromelanin signal could be detected in PD, raising the possibility of using it as a marker of disease progression. Conclusion:Nigrosome imaging and neuromelanin sensitive MRI are promising tools to study nigral pathology and to improve the diagnosis of PD. However, further studies are required to standardize analysis approaches, confirm longitudinal changes, and assess their generalizability.
Project description:During aging, neuronal organelles filled with neuromelanin (a dark-brown pigment) and lipid bodies accumulate in the brain, particularly in the substantia nigra, a region targeted in Parkinson's disease. We have investigated protein and lipid systems involved in the formation of these organelles and in the synthesis of the neuromelanin of human substantia nigra. Membrane and matrix proteins characteristic of lysosomes were found in neuromelanin-containing organelles at a lower number than in typical lysosomes, indicating a reduced enzymatic activity and likely impaired capacity for lysosomal and autophagosomal fusion. The presence of proteins involved in lipid transport may explain the accumulation of lipid bodies in the organelle and the lipid component in neuromelanin structure. The major lipids observed in lipid bodies of the organelle are dolichols with lower amounts of other lipids. Proteins of aggregation and degradation pathways were present, suggesting a role for accumulation by this organelle when the ubiquitin-proteasome system is inadequate. The presence of proteins associated with aging and storage diseases may reflect impaired autophagic degradation or impaired function of lysosomal enzymes. The identification of typical autophagy proteins and double membranes demonstrates the organelle's autophagic nature and indicates that it has engulfed neuromelanin precursors from the cytosol. Based on these data, it appears that the neuromelanin-containing organelle has a very slow turnover during the life of a neuron and represents an intracellular compartment of final destination for numerous molecules not degraded by other systems.
Project description:This article gives a detailed description of a protocol using density gradient centrifugation for the enrichment of neuromelanin granules and synaptosomes from low amounts (?0.15g) of human substantia nigra pars compacta tissue. This has a great advantage compared to already existing methods as it allows for the first time (i) a combined enrichment of neuromelanin granules and synaptosomes and (ii) just minimal amounts of tissue necessary to enable donor specific analysis. Individual specimens were classified as control or diseased according to clinical evaluation and neuropathological examination. For the enrichment of synaptosomes and neuromelanin granules from the same tissue sample density gradient centrifugations using Percoll® and Iodixanol were performed. The purity of resulting fractions was checked by transmission electron microscopy. We were able to establish a reproducible and easy to handle protocol combining two different density gradient centrifugations: using an Iodixanol gradient neuromelanin granules were enriched and in parallel, from the same sample, a fraction of synaptosomes with high purity using a Percoll® gradient was obtained. Our subfractionation strategy will enable a subsequent in depth proteomic characterization of neurodegenerative processes in the substantia nigra pars compacta in patients with Parkinson's disease and dementia with Lewy bodies compared to appropriate controls.Key features of Parkinson's disease are the degeneration of dopaminergic neurons in the substantia nigra pars compacta, an associated loss of the brain pigment neuromelanin and a resulting impairment of the neuronal network. The accumulation of iron binding neuromelanin granules is age- and disease-dependent and disease specific alterations could affect the neuronal iron homeostasis leading to oxidative stress induced cell death. The focus of the described method is the analysis of neuromelanin granules as well as axonal cell-endings of nerve cells (synaptosomes) of individual donors (control and diseased). It is the basis for the identification of disease-relevant changes in the iron homeostasis and the generation of new insight into altered protein compositions or regulations which might lead to disturbed communications between nerve cells resulting in pathogenic processes.
Project description:Neuromelanin sensitive magnetic resonance imaging (NMS-MRI) has been crucial in identifying abnormalities in the substantia nigra pars compacta (SNc) in Parkinson's disease (PD) as PD is characterized by loss of dopaminergic neurons in the SNc. Current techniques employ estimation of contrast ratios of the SNc, visualized on NMS-MRI, to discern PD patients from the healthy controls. However, the extraction of these features is time-consuming and laborious and moreover provides lower prediction accuracies. Furthermore, these do not account for patterns of subtle changes in PD in the SNc. To mitigate this, our work establishes a computer-based analysis technique that uses convolutional neural networks (CNNs) to create prognostic and diagnostic biomarkers of PD from NMS-MRI. Our technique not only performs with a superior testing accuracy (80%) as compared to contrast ratio-based classification (56.5% testing accuracy) and radiomics classifier (60.3% testing accuracy), but also supports discriminating PD from atypical parkinsonian syndromes (85.7% test accuracy). Moreover, it has the capability to locate the most discriminative regions on the neuromelanin contrast images. These discriminative activations demonstrate that the left SNc plays a key role in the classification in comparison to the right SNc, and are in agreement with the concept of asymmetry in PD. Overall, the proposed technique has the potential to support radiological diagnosis of PD while facilitating deeper understanding into the abnormalities in SNc.
Project description:BACKGROUND AND PURPOSE:Whether the neuromelanin-positive substantia nigra pars compacta area (NM-SNc) on neuromelanin magnetic resonance imaging (NM-MRI) and the specific binding ratio (SBR) on 123 I-N-v-fluoropropyl-2b-carbomethoxy3b-(4-iodophenyl)nortropane single photon emission computed tomography (DaT-SPECT) can be correlated with motor fluctuations (MFs) in advanced Parkinson's disease (PD) was investigated. METHODS:Thirty-five PD patients (60 ± 13 years) and 23 healthy individuals as controls (59 ± 19 years) were enrolled. The relationships between NM-MRI and DaT-SPECT were prospectively examined in two subgroups divided according to the presence or absence of MFs. Multivariate analysis was performed using the Cox proportional hazard model to screen for association factors. RESULTS:The NM-SNc size was correlated with the SBR (Spearman's ? = 0.43, P < 0.05). The NM-SNc size was significantly reduced in PD with MFs compared with the subgroup without (P < 0.001), whereas the SBR did not significantly differ between the groups. NM-SNc size was a significant association factor for MFs (hazard ratio 0.94, P = 0.04). In receiver operating characteristic analysis of the factors for MF occurrence, the area under the receiver operating characteristic curve of the NM-SNc size showed a significant difference of 0.89 (P < 0.05) but no significant difference was found in the SBR. CONCLUSIONS:NM-SNc size was significantly correlated with the SBR in PD, but several factors in advanced PD were more closely associated with NM-SNc size than the SBR. NM-MRI might reflect the status of advanced PD more accurately than DaT-SPECT. Therefore, NM-MRI appears to provide a better marker for discriminating advanced PD than DaT-SPECT.
Project description:Neuromelanin is a complex polymer pigment found primarily in the dopaminergic neurons of human substantia nigra. Neuromelanin pigment is stored in granules including a protein matrix and lipid droplets. Neuromelanin granules are yet only partially characterised regarding their structure and function. To clarify the exact function of neuromelanin granules in humans, their enrichment and in-depth characterization from human substantia nigra is necessary. Previously published global proteome studies of neuromelanin granules in human substantia nigra required high tissue amounts. Due to the limited availability of human brain tissue we established a new method based on laser microdissection combined with mass spectrometry for the isolation and analysis of neuromelanin granules. With this method it is possible for the first time to isolate a sufficient amount of neuromelanin granules for global proteomics analysis from ten 10 µm tissue sections. In total 1,000 proteins were identified associated with neuromelanin granules. More than 68% of those proteins were also identified in previously performed studies. Our results confirm and further extend previously described findings, supporting the connection of neuromelanin granules to iron homeostasis and lysosomes, respectively endosomes. Hence, this method is suitable for the donor specific enrichment and proteomic analysis of neuromelanin granules.
Project description:In Parkinson's disease (PD), there is a reduction of neuromelanin (NM) in the substantia nigra (SN). Manual quantification of the NM volume in the SN is unpractical and time-consuming; therefore, we aimed to quantify NM in the SN with a novel semi-automatic segmentation method. Twenty patients with PD and twelve healthy subjects (HC) were included in this study. T1-weighted spectral pre-saturation with inversion recovery (SPIR) images were acquired on a 3T scanner. Manual and semi-automatic atlas-free local statistics signature-based segmentations measured the surface and volume of SN, respectively. Midbrain volume (MV) was calculated to normalize the data. Receiver operating characteristic (ROC) analysis was performed to determine the sensitivity and specificity of both methods. PD patients had significantly lower SN mean surface (37.7 ± 8.0 vs. 56.9 ± 6.6 mm2) and volume (235.1 ± 45.4 vs. 382.9 ± 100.5 mm3) than HC. After normalization with MV, the difference remained significant. For surface, sensitivity and specificity were 91.7 and 95 percent, respectively. For volume, sensitivity and specificity were 91.7 and 90 percent, respectively. Manual and semi-automatic segmentation methods of the SN reliably distinguished between PD patients and HC. ROC analysis shows the high sensitivity and specificity of both methods.