RESUMO
AIMS: Endoplasmic reticulum stress followed by the unfolded protein response is one of the cellular mechanisms contributing to the progression of α-synuclein pathology in Parkinson's disease and other Lewy body diseases. We aimed to investigate the activation of endoplasmic reticulum stress and its correlation with α-synuclein pathology in human post-mortem brain tissue. METHODS: We analysed brain tissue from 45 subjects-14 symptomatic patients with Lewy body disease, 19 subjects with incidental Lewy body disease, and 12 healthy controls. The analysed brain regions included the medulla, pons, midbrain, striatum, amygdala and entorhinal, temporal, frontal and occipital cortex. We analysed activation of endoplasmic reticulum stress via levels of the unfolded protein response-related proteins (Grp78, eIF2α) and endoplasmic reticulum stress-regulating neurotrophic factors (MANF, CDNF). RESULTS: We showed that regional levels of two endoplasmic reticulum-localised neurotrophic factors, MANF and CDNF, did not change in response to accumulating α-synuclein pathology. The concentration of MANF negatively correlated with age in specific regions. eIF2α was upregulated in the striatum of Lewy body disease patients and correlated with increased α-synuclein levels. We found the upregulation of chaperone Grp78 in the amygdala and nigral dopaminergic neurons of Lewy body disease patients. Grp78 levels in the amygdala strongly correlated with soluble α-synuclein levels. CONCLUSIONS: Our data suggest a strong but regionally specific change in Grp78 and eIF2α levels, which positively correlates with soluble α-synuclein levels. Additionally, MANF levels decreased in dopaminergic neurons in the substantia nigra. Our research suggests that endoplasmic reticulum stress activation is not associated with Lewy pathology but rather with soluble α-synuclein concentration and disease progression.
Assuntos
Chaperona BiP do Retículo Endoplasmático , Fator de Iniciação 2 em Eucariotos , Proteínas de Choque Térmico , Doença por Corpos de Lewy , Resposta a Proteínas não Dobradas , Regulação para Cima , alfa-Sinucleína , Humanos , Doença por Corpos de Lewy/patologia , Doença por Corpos de Lewy/metabolismo , Chaperona BiP do Retículo Endoplasmático/metabolismo , alfa-Sinucleína/metabolismo , Masculino , Idoso , Feminino , Resposta a Proteínas não Dobradas/fisiologia , Idoso de 80 Anos ou mais , Proteínas de Choque Térmico/metabolismo , Fator de Iniciação 2 em Eucariotos/metabolismo , Encéfalo/metabolismo , Encéfalo/patologia , Pessoa de Meia-Idade , Fatores de Crescimento Neural/metabolismo , Estresse do Retículo Endoplasmático/fisiologia , Biomarcadores/metabolismoRESUMO
Several studies have confirmed the α-synuclein real-time quaking-induced conversion (RT-QuIC) assay to have high sensitivity and specificity for Parkinson's disease. However, whether the assay can be used as a robust, quantitative measure to monitor disease progression, stratify different synucleinopathies and predict disease conversion in patients with idiopathic REM sleep behaviour disorder remains undetermined. The aim of this study was to assess the diagnostic value of CSF α-synuclein RT-QuIC quantitative parameters in regard to disease progression, stratification and conversion in synucleinopathies. We performed α-synuclein RT-QuIC in the CSF samples from 74 Parkinson's disease, 24 multiple system atrophy and 45 idiopathic REM sleep behaviour disorder patients alongside 55 healthy controls, analysing quantitative assay parameters in relation to clinical data. α-Synuclein RT-QuIC showed 89% sensitivity and 96% specificity for Parkinson's disease. There was no correlation between RT-QuIC quantitative parameters and Parkinson's disease clinical scores (e.g. Unified Parkinson's Disease Rating Scale motor), but RT-QuIC positivity and some quantitative parameters (e.g. Vmax) differed across the different phenotype clusters. RT-QuIC parameters also added value alongside standard clinical data in diagnosing Parkinson's disease. The sensitivity in multiple system atrophy was 75%, and CSF samples showed longer T50 and lower Vmax compared to Parkinson's disease. All RT-QuIC parameters correlated with worse clinical progression of multiple system atrophy (e.g. change in Unified Multiple System Atrophy Rating Scale). The overall sensitivity in idiopathic REM sleep behaviour disorder was 64%. In three of the four longitudinally followed idiopathic REM sleep behaviour disorder cohorts, we found around 90% sensitivity, but in one sample (DeNoPa) diagnosing idiopathic REM sleep behaviour disorder earlier from the community cases, this was much lower at 39%. During follow-up, 14 of 45 (31%) idiopathic REM sleep behaviour disorder patients converted to synucleinopathy with 9/14 (64%) of convertors showing baseline RT-QuIC positivity. In summary, our results showed that α-synuclein RT-QuIC adds value in diagnosing Parkinson's disease and may provide a way to distinguish variations within Parkinson's disease phenotype. However, the quantitative parameters did not correlate with disease severity in Parkinson's disease. The assay distinguished multiple system atrophy patients from Parkinson's disease patients and in contrast to Parkinson's disease, the quantitative parameters correlated with disease progression of multiple system atrophy. Our results also provided further evidence for α-synuclein RT-QuIC having potential as an early biomarker detecting synucleinopathy in idiopathic REM sleep behaviour disorder patients prior to conversion. Further analysis of longitudinally followed idiopathic REM sleep behaviour disorder patients is needed to better understand the relationship between α-synuclein RT-QuIC signature and the progression from prodromal to different synucleinopathies.
Assuntos
Atrofia de Múltiplos Sistemas , Doença de Parkinson , Transtorno do Comportamento do Sono REM , Sinucleinopatias , Progressão da Doença , Humanos , Atrofia de Múltiplos Sistemas/diagnóstico , Doença de Parkinson/diagnóstico , Transtorno do Comportamento do Sono REM/diagnóstico , Sinucleinopatias/diagnóstico , alfa-Sinucleína/análiseRESUMO
Aggregation of alpha-synuclein into inclusion bodies, termed Lewy pathology, is a defining feature of Parkinson's disease (PD) and Dementia with Lewy bodies (DLB). In the majority of post mortem cases, the distribution of Lewy pathology seems to follow two overarching patterns: a caudo-rostral pattern with relatively more pathology in the brainstem than in the telencephalon, and an amygdala-centered pattern with the most abundant pathology in the "center of the brain", including the amygdala, entorhinal cortex, and substantia nigra, and relatively less pathology in the lower brainstem and spinal autonomic nuclei. The recent body-first versus brain-first model of Lewy Body Disorders proposes that the initial pathogenic alpha-synuclein in some patients originates in the enteric nervous system with secondary spreading to the brain; and in other patients originates inside the CNS with secondary spreading to the lower brainstem and peripheral autonomic nervous system. Here, we use two existing post mortem datasets to explore the possibility that clinical body-first and brain-first subtypes are equivalent to the caudo-rostral and amygdala-centered patterns of Lewy pathology seen at post mortem.
Assuntos
Doença por Corpos de Lewy , Doença de Parkinson , Encéfalo/metabolismo , Tronco Encefálico/metabolismo , Humanos , Doença por Corpos de Lewy/patologia , Doença de Parkinson/patologia , Substância Negra/metabolismo , alfa-Sinucleína/metabolismoRESUMO
OBJECTIVE: Neuronal loss in the substantia nigra pars compacta (SNpc) in Parkinson disease (PD) is not uniform, as dopamine neurons from the ventral tier are lost more rapidly than those of the dorsal tier. Identifying the intrinsic differences that account for this differential vulnerability may provide a key for developing new treatments for PD. METHODS: Here, we compared the RNA-sequenced transcriptomes of ~100 laser captured microdissected SNpc neurons from each tier from 7 healthy controls. RESULTS: Expression levels of dopaminergic markers were similar across the tiers, whereas markers specific to the neighboring ventral tegmental area were virtually undetected. After accounting for unwanted sources of variation, we identified 106 differentially expressed genes (DEGs) between the SNpc tiers. The genes higher in the dorsal/resistant SNpc tier neurons displayed coordinated patterns of expression across the human brain, their protein products had more interactions than expected by chance, and they demonstrated evidence of functional convergence. No significant shared functionality was found for genes higher in the ventral/vulnerable SNpc tier. Surprisingly but importantly, none of the identified DEGs was among the familial PD genes or genome-wide associated loci. Finally, we found some DEGs in opposite tier orientation between human and analogous mouse populations. INTERPRETATION: Our results highlight functional enrichments of vesicular trafficking, ion transport/homeostasis and oxidative stress genes showing higher expression in the resistant neurons of the SNpc dorsal tier. Furthermore, the comparison of gene expression variation in human and mouse SNpc populations strongly argues for the need of human-focused omics studies. ANN NEUROL 2020;87:853-868.
Assuntos
Neurônios Dopaminérgicos/patologia , Mesencéfalo/patologia , Doença de Parkinson/genética , Doença de Parkinson/patologia , Transcriptoma , Animais , Regulação da Expressão Gênica/genética , Estudo de Associação Genômica Ampla , Voluntários Saudáveis , Humanos , Camundongos , RNA/genética , Substância Negra/patologia , Área Tegmentar Ventral/patologiaRESUMO
Currently, the neuropathological diagnosis of Lewy body disease (LBD) may be stated according to several staging systems, which include the Braak Lewy body stages (Braak), the consensus criteria by McKeith and colleagues (McKeith), the modified McKeith system by Leverenz and colleagues (Leverenz), and the Unified Staging System by Beach and colleagues (Beach). All of these systems use semi-quantitative scoring (4- or 5-tier scales) of Lewy pathology (LP; i.e., Lewy bodies and Lewy neurites) in defined cortical and subcortical areas. While these systems are widely used, some suffer from low inter-rater reliability and/or an inability to unequivocally classify all cases with LP. To address these limitations, we devised a new system, the LP consensus criteria (LPC), which is based on the McKeith system, but applies a dichotomous approach for the scoring of LP (i.e., "absent" vs. "present") and includes amygdala-predominant and olfactory-only stages. α-Synuclein-stained slides from brainstem, limbic system, neocortex, and olfactory bulb from a total of 34 cases with LP provided by the Newcastle Brain Tissue Resource (NBTR) and the University of Pennsylvania brain bank (UPBB) were scanned and assessed by 16 raters, who provided diagnostic categories for each case according to Braak, McKeith, Leverenz, Beach, and LPC systems. In addition, using LP scores available from neuropathological reports of LP cases from UPBB (n = 202) and NBTR (n = 134), JT (UPBB) and JA (NBTR) assigned categories according to all staging systems to these cases. McKeith, Leverenz, and LPC systems reached good (Krippendorff's α ≈ 0.6), while both Braak and Beach systems had lower (Krippendorff's α ≈ 0.4) inter-rater reliability, respectively. Using the LPC system, all cases could be unequivocally classified by the majority of raters, which was also seen for 97.1% when the Beach system was used. However, a considerable proportion of cases could not be classified when using Leverenz (11.8%), McKeith (26.5%), or Braak (29.4%) systems. The category of neocortical LP according to the LPC system was associated with a 5.9 OR (p < 0.0001) of dementia in the 134 NBTR cases and a 3.14 OR (p = 0.0001) in the 202 UPBB cases. We established that the LPC system has good reproducibility and allows classification of all cases into distinct categories. We expect that it will be reliable and useful in routine diagnostic practice and, therefore, suggest that it should be the standard future approach for the basic post-mortem evaluation of LP.
Assuntos
Encéfalo/patologia , Doença por Corpos de Lewy/patologia , Autopsia , Mapeamento Encefálico , Consenso , Humanos , Corpos de Lewy/patologia , Doença por Corpos de Lewy/classificação , Doença por Corpos de Lewy/diagnóstico , Variações Dependentes do Observador , Reprodutibilidade dos TestesRESUMO
Recent large-scale genetic studies have allowed for the first glimpse of the effects of common genetic variability in dementia with Lewy bodies (DLB), identifying risk variants with appreciable effect sizes. However, it is currently well established that a substantial portion of the genetic heritable component of complex traits is not captured by genome-wide significant SNPs. To overcome this issue, we have estimated the proportion of phenotypic variance explained by genetic variability (SNP heritability) in DLB using a method that is unbiased by allele frequency or linkage disequilibrium properties of the underlying variants. This shows that the heritability of DLB is nearly twice as high as previous estimates based on common variants only (31% vs 59.9%). We also determine the amount of phenotypic variance in DLB that can be explained by recent polygenic risk scores from either Parkinson's disease (PD) or Alzheimer's disease (AD), and show that, despite being highly significant, they explain a low amount of variance. Additionally, to identify pleiotropic events that might improve our understanding of the disease, we performed genetic correlation analyses of DLB with over 200 diseases and biomedically relevant traits. Our data shows that DLB has a positive correlation with education phenotypes, which is opposite to what occurs in AD. Overall, our data suggests that novel genetic risk factors for DLB should be identified by larger GWAS and these are likely to be independent from known AD and PD risk variants.
Assuntos
Predisposição Genética para Doença , Variação Genética , Doença por Corpos de Lewy/genética , Bases de Dados Genéticas , HumanosRESUMO
Biallelic DNAJC12 mutations were described in children with hyperphenylalaninemia, neurodevelopmental delay, and dystonia. We identified DNAJC12 homozygous null variants (c.187A>T;p.K63* and c.79-2A>G;p.V27Wfs*14) in two kindreds with early-onset parkinsonism. Both probands had mild intellectual disability, mild nonprogressive, motor symptoms, sustained benefit from small dose of levodopa, and substantial worsening of symptoms after levodopa discontinuation. Neuropathology (Proband-A) revealed no alpha-synuclein pathology, and substantia nigra depigmentation with moderate cell loss. DNAJC12 transcripts were reduced in both patients. Our results suggest that DNAJC12 mutations (absent in 500 early-onset patients with Parkinson's disease) rarely cause dopa-responsive nonprogressive parkinsonism in adulthood, but broaden the clinical spectrum of DNAJC12 deficiency. Ann Neurol 2017;82:640-646.
Assuntos
Antiparkinsonianos/uso terapêutico , Levodopa/uso terapêutico , Mutação/genética , Transtornos Parkinsonianos/tratamento farmacológico , Transtornos Parkinsonianos/genética , Proteínas Repressoras/genética , Adulto , Peptídeos beta-Amiloides/metabolismo , Aminas Biogênicas/metabolismo , Encéfalo/metabolismo , Encéfalo/patologia , Análise Mutacional de DNA , Proteínas de Ligação a DNA/metabolismo , Saúde da Família , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Transtornos Parkinsonianos/patologia , Fenilalanina/metabolismo , Proteína Sequestossoma-1/metabolismo , Adulto Jovem , alfa-Sinucleína/metabolismo , Proteínas tau/metabolismoRESUMO
OBJECTIVE: The relationship between Parkinson disease (PD), PD with dementia (PDD), and dementia with Lewy bodies (DLB) has long been debated. Although PD is primarily considered a motor disorder, cognitive impairment is often present at diagnosis, and only â¼20% of patients remain cognitively intact in the long term. Alpha-synuclein (SNCA) was first implicated in the pathogenesis of the disease when point mutations and locus multiplications were identified in familial parkinsonism with dementia. In worldwide populations, SNCA genetic variability remains the most reproducible risk factor for idiopathic PD. However, few investigators have looked at SNCA variability in terms of cognitive outcomes. METHODS: We have used targeted high-throughput sequencing to characterize the 135kb SNCA locus in a large multinational cohort of patients with PD, PDD, and DLB and healthy controls. RESULTS: An analysis of 43 tagging single nucleotide polymorphisms across the SNCA locus shows 2 distinct association profiles for symptoms of parkinsonism and/or dementia, respectively, toward the 3' or the 5' of the SNCA gene. In addition, we define a specific haplotype in intron 4 that is directly associated with PDD. The PDD risk haplotype has been interrogated at single nucleotide resolution and is uniquely tagged by an expanded TTTCn repeat. INTERPRETATION: Our data show that PD, PDD, and DLB, rather than a disease continuum, have distinct genetic etiologies albeit within one genomic locus. Such results may serve as prognostic biomarkers to these disorders, to inform physicians and patients, and to assist in the design and stratification of clinical trials aimed at disease modification. Ann Neurol 2016;79:991-999.
Assuntos
Disfunção Cognitiva/genética , Demência/genética , Doença por Corpos de Lewy/genética , Doença por Corpos de Lewy/psicologia , Doença de Parkinson/genética , Doença de Parkinson/psicologia , alfa-Sinucleína/genética , Idoso , Idoso de 80 Anos ou mais , Biomarcadores , Estudos de Casos e Controles , Disfunção Cognitiva/complicações , Demência/complicações , Demência/psicologia , Feminino , Predisposição Genética para Doença/genética , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Doença por Corpos de Lewy/complicações , Masculino , Doença de Parkinson/complicações , Polimorfismo de Nucleotídeo Único/genéticaRESUMO
Clinical and neuropathological similarities between dementia with Lewy bodies (DLB), Parkinson's and Alzheimer's diseases (PD and AD, respectively) suggest that these disorders may share etiology. To test this hypothesis, we have performed an association study of 54 genomic regions, previously implicated in PD or AD, in a large cohort of DLB cases and controls. The cohort comprised 788 DLB cases and 2624 controls. To minimize the issue of potential misdiagnosis, we have also performed the analysis including only neuropathologically proven DLB cases (667 cases). The results show that the APOE is a strong genetic risk factor for DLB, confirming previous findings, and that the SNCA and SCARB2 loci are also associated after a study-wise Bonferroni correction, although these have a different association profile than the associations reported for the same loci in PD. We have previously shown that the p.N370S variant in GBA is associated with DLB, which, together with the findings at the SCARB2 locus, suggests a role for lysosomal dysfunction in this disease. These results indicate that DLB has a unique genetic risk profile when compared with the two most common neurodegenerative diseases and that the lysosome may play an important role in the etiology of this disorder. We make all these data available.
Assuntos
Apolipoproteínas E/genética , Doença por Corpos de Lewy/etiologia , Proteínas de Membrana Lisossomal/genética , Lisossomos/patologia , Receptores Depuradores/genética , alfa-Sinucleína/genética , Doença de Alzheimer/etiologia , Doença de Alzheimer/genética , Doença de Alzheimer/patologia , Estudos de Casos e Controles , Estudos de Coortes , Feminino , Estudos de Associação Genética , Loci Gênicos , Humanos , Doença por Corpos de Lewy/genética , Doença por Corpos de Lewy/patologia , Masculino , Doença de Parkinson/etiologia , Doença de Parkinson/genética , Doença de Parkinson/patologia , Fatores de RiscoRESUMO
In recent years, several studies have investigated the potential of immunohistochemical detection of α-synuclein in the gastrointestinal tract to diagnose Parkinson's disease (PD). Although methodological heterogeneity has hindered comparability between studies, it has become increasingly apparent that the high sensitivity and specificity reported in preliminary studies has not been sustained in subsequent large-scale studies. What constitutes pathological α-synuclein in the alimentary canal that could distinguish between PD patients and controls and how this can be reliably detected represent key outstanding questions in the field. In this review, we will comment on and compare the variable technical aspects from previous studies, and by highlighting some advantages and shortcomings we hope to delineate a standardized approach to facilitate the consensus criteria urgently needed in the field. Furthermore, we will describe alternative detection techniques to conventional immunohistochemistry that have recently emerged and may facilitate ease of interpretation and reliability of gastrointestinal α-synuclein detection. Such techniques have the potential to detect the presence of pathological α-synuclein and include the paraffin-embedded tissue blot, the proximity ligation assay, the protein misfolding cyclic amplification technique, and the real-time quaking-induced conversion assay. Finally, we will review 2 nonsynonymous theories that have driven enteric α-synuclein research, namely, (1) that α-synuclein propagates in a prion-like fashion from the peripheral nervous system to the brain via vagal connections and (2) that gastrointestinal α-synuclein deposition may be used as a clinically useful biomarker in PD.
Assuntos
Biomarcadores/metabolismo , Trato Gastrointestinal/metabolismo , Doença de Parkinson/diagnóstico , alfa-Sinucleína/metabolismo , HumanosRESUMO
The pathological end-state of Parkinson disease is well described from postmortem tissue, but there remains a pressing need to define early functional changes to susceptible neurons and circuits. In particular, mechanisms underlying the vulnerability of the dopamine neurons of the substantia nigra pars compacta (SNc) and the importance of protein aggregation in driving the disease process remain to be determined. To better understand the sequence of events occurring in familial and sporadic Parkinson disease, we generated bacterial artificial chromosome transgenic mice (SNCA-OVX) that express wild-type α-synuclein from the complete human SNCA locus at disease-relevant levels and display a transgene expression profile that recapitulates that of endogenous α-synuclein. SNCA-OVX mice display age-dependent loss of nigrostriatal dopamine neurons and motor impairments characteristic of Parkinson disease. This phenotype is preceded by early deficits in dopamine release from terminals in the dorsal, but not ventral, striatum. Such neurotransmission deficits are not seen at either noradrenergic or serotoninergic terminals. Dopamine release deficits are associated with an altered distribution of vesicles in dopaminergic axons in the dorsal striatum. Aged SNCA-OVX mice exhibit reduced firing of SNc dopamine neurons in vivo measured by juxtacellular recording of neurochemically identified neurons. These progressive changes in vulnerable SNc neurons were observed independently of overt protein aggregation, suggesting neurophysiological changes precede, and are not driven by, aggregate formation. This longitudinal phenotyping strategy in SNCA-OVX mice thus provides insights into the region-specific neuronal disturbances preceding and accompanying Parkinson disease.
Assuntos
Envelhecimento/metabolismo , Corpo Estriado/metabolismo , Neurônios Dopaminérgicos/metabolismo , Transtornos Parkinsonianos/metabolismo , Substância Negra/metabolismo , Transmissão Sináptica , Envelhecimento/patologia , Animais , Cromossomos Artificiais Bacterianos/genética , Cromossomos Artificiais Bacterianos/metabolismo , Corpo Estriado/patologia , Corpo Estriado/fisiopatologia , Neurônios Dopaminérgicos/patologia , Humanos , Camundongos , Transtornos Parkinsonianos/genética , Transtornos Parkinsonianos/patologia , Transtornos Parkinsonianos/fisiopatologia , Substância Negra/patologia , Substância Negra/fisiopatologia , alfa-Sinucleína/biossíntese , alfa-Sinucleína/genéticaRESUMO
α-Synuclein (α-syn), a small protein that has the intrinsic propensity to aggregate, is implicated in several neurodegenerative diseases including Parkinson's disease (PD), dementia with Lewy bodies (DLB), and multiple system atrophy (MSA), which are collectively known as synucleinopathies. Genetic, pathological, biochemical, and animal modeling studies provided compelling evidence that α-syn aggregation plays a key role in the pathogenesis of PD and related synucleinopathies. It is therefore of utmost importance to develop reliable tools that can detect the aggregated forms of α-syn. We describe here the generation and characterization of six novel conformation-specific monoclonal antibodies that recognize specifically α-syn aggregates but not the soluble, monomeric form of the protein. The antibodies described herein did not recognize monomers or fibrils generated from other amyloidogenic proteins including ß-syn, γ-syn, ß-amyloid, tau protein, islet amyloid polypeptide and ABri. Interestingly, the antibodies did not react to overlapping linear peptides spanning the entire sequence of α-syn, confirming further that they only detect α-syn aggregates. In immunohistochemical studies, the new conformation-specific monoclonal antibodies showed underappreciated small micro-aggregates and very thin neurites in PD and DLB cases that were not observed with generic pan antibodies that recognize linear epitope. Furthermore, employing one of our conformation-specific antibodies in a sandwich based ELISA, we observed an increase in levels of α-syn oligomers in brain lysates from DLB compared to Alzheimer's disease and control samples. Therefore, the conformation-specific antibodies portrayed herein represent useful tools for research, biomarkers development, diagnosis and even immunotherapy for PD and related pathologies.
Assuntos
Anticorpos Monoclonais/imunologia , Encéfalo/metabolismo , alfa-Sinucleína/química , alfa-Sinucleína/imunologia , Proteínas Adaptadoras de Transdução de Sinal , Doença de Alzheimer/metabolismo , Peptídeos beta-Amiloides/metabolismo , Animais , Anticorpos Monoclonais/isolamento & purificação , Anticorpos Monoclonais/metabolismo , Encéfalo/patologia , Escherichia coli , Polipeptídeo Amiloide das Ilhotas Pancreáticas/metabolismo , Doença por Corpos de Lewy/metabolismo , Doença por Corpos de Lewy/patologia , Glicoproteínas de Membrana/metabolismo , Camundongos , Proteínas de Neoplasias/imunologia , Proteínas de Neoplasias/metabolismo , Doença de Parkinson/metabolismo , Doença de Parkinson/patologia , Fragmentos de Peptídeos/metabolismo , Conformação Proteica , Multimerização Proteica , Proteínas Recombinantes/química , Proteínas Recombinantes/imunologia , Proteínas Recombinantes/metabolismo , alfa-Sinucleína/metabolismo , beta-Sinucleína/imunologia , beta-Sinucleína/metabolismo , gama-Sinucleína/imunologia , gama-Sinucleína/metabolismo , Proteínas tau/metabolismoRESUMO
Parkinson's disease (PD) is a neurodegenerative disorder classically characterized by the death of dopamine (DA) neurons in the substantia nigra pars compacta and by intracellular Lewy bodies composed largely of α-synuclein. Approximately 5-10% of PD patients have a familial form of Parkinsonism, including mutations in α-synuclein. To better understand the cell-type specific role of α-synuclein on DA neurotransmission, and the effects of the disease-associated A30P mutation, we generated and studied a novel transgenic model of PD. We expressed the A30P mutant form of human α-synuclein in a spatially-relevant manner from the 111kb SNCA genomic DNA locus on a bacterial artificial chromosome (BAC) insert on a mouse null (Snca-/-) background. The BAC transgenic mice expressed α-synuclein in tyrosine hydroxylase-positive neurons and expression of either A30P α-synuclein or wildtype α-synuclein restored the sensitivity of DA neurons to MPTP in resistant Snca-/- animals. A30P α-synuclein mice showed no Lewy body-like aggregation, and did not lose catecholamine neurons in substantia nigra or locus coeruleus. However, using cyclic voltammetry at carbon-fiber microelectrodes we identified a deficit in evoked DA release in the caudate putamen, but not in the nucleus accumbens, of SNCA-A30P Snca-/- mice but no changes to release of another catecholamine, norepinephrine (NE), in the NE-rich ventral bed nucleus of stria terminalis. SNCA-A30P Snca-/- mice had no overt behavioral impairments but exhibited a mild increase in wheel-running. In summary, this refined PD mouse model shows that A30P α-synuclein preferentially perturbs the dopaminergic system in the dorsal striatum, reflecting the region-specific change seen in PD.
Assuntos
Gânglios da Base/metabolismo , Dopamina/metabolismo , Neurônios Dopaminérgicos/metabolismo , Norepinefrina/metabolismo , alfa-Sinucleína/genética , Fatores Etários , Animais , Cromossomos Artificiais Bacterianos , Modelos Animais de Doenças , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Núcleos Septais/metabolismo , alfa-Sinucleína/metabolismoRESUMO
BACKGROUND: Dementia is one of the milestones of advanced Parkinson's disease (PD), with its neuropathological substrate still being a matter of debate, particularly regarding its potential mechanistic implications. OBJECTIVE: The aim of this study was to review the relative importance of Lewy-related α-synuclein and Alzheimer's tau and amyloid-ß (Aß) pathologies in disease progression and dementia in PD. METHODS: We reviewed studies conducted at the Queen Square Brain Bank, Institute of Neurology, University College London, using large PD cohorts. RESULTS: Cortical Lewy- and Alzheimer-type pathologies are associated with milestones of poorer prognosis and with non-tremor predominance, which have been, in turn, linked to dementia. The combination of these pathologies is the most robust neuropathological substrate of PD-related dementia, with cortical Aß burden determining a faster progression to dementia. CONCLUSION: The shared relevance of these pathologies in PD progression and dementia is in line with experimental data suggesting synergism between α-synuclein, tau and Aß and with studies testing these proteins as disease biomarkers, hence favouring the eventual testing of therapeutic strategies targeting these proteins in PD.
Assuntos
Peptídeos beta-Amiloides/metabolismo , Demência/patologia , Doença de Parkinson/patologia , alfa-Sinucleína/metabolismo , Proteínas tau/metabolismo , Encéfalo/metabolismo , Encéfalo/patologia , Demência/etiologia , Demência/metabolismo , Progressão da Doença , Humanos , Doença de Parkinson/complicações , Doença de Parkinson/metabolismo , Estudos RetrospectivosRESUMO
Motivation: Pooled designs for single-cell RNA sequencing, where many cells from distinct samples are processed jointly, offer increased throughput and reduced batch variation. This study describes expression-aware demultiplexing (EAD), a computational method that employs differential co-expression patterns between individuals to demultiplex pooled samples without any extra experimental steps. Results: We use synthetic sample pools and show that the top interindividual differentially co-expressed genes provide a distinct cluster of cells per individual, significantly enriching the regulation of metabolism. Our application of EAD to samples of six isogenic inbred mice demonstrated that controlling genetic and environmental effects can solve interindividual variations related to metabolic pathways. We utilized 30 samples from both sepsis and healthy individuals in six batches to assess the performance of classification approaches. The results indicate that combining genetic and EAD results can enhance the accuracy of assignments (Min. 0.94, Mean 0.98, Max. 1). The results were enhanced by an average of 1.4% when EAD and barcoding techniques were combined (Min. 1.25%, Median 1.33%, Max. 1.74%). Furthermore, we demonstrate that interindividual differential co-expression analysis within the same cell type can be used to identify cells from the same donor in different activation states. By analysing single-nuclei transcriptome profiles from the brain, we demonstrate that our method can be applied to nonimmune cells. Availability and implementation: EAD workflow is available at https://isarnassiri.github.io/scDIV/ as an R package called scDIV (acronym for single-cell RNA-sequencing data demultiplexing using interindividual variations).
RESUMO
Idiopathic Parkinson's disease is determined by a combination of genetic and environmental factors. Recently, the first genome-wide association study on short-tandem repeats in Parkinson's disease reported on eight suggestive short-tandem repeat-based risk loci (α = 5.3 × 10-6), of which four were novel, i.e. they had not been implicated in Parkinson's disease risk by genome-wide association analyses of single-nucleotide polymorphisms before. Here, we tested these eight candidate short-tandem repeats in a large, independent Parkinson's disease case-control dataset (n = 4757). Furthermore, we combined the results from both studies by meta-analysis resulting in the largest Parkinson's disease genome-wide association study of short-tandem repeats to date (n = 43 844). Lastly, we investigated whether leading short-tandem repeat risk variants exert functional effects on gene expression regulation based on methylation quantitative trait locus data in human 'post-mortem' brain (n = 142). None of the eight previously reported short-tandem repeats were significantly associated with Parkinson's disease in our independent dataset after multiple testing correction (α = 6.25 × 10-3). However, we observed modest support for short-tandem repeats near CCAR2 and NCOR1 in the updated meta-analyses of all available data. While the genome-wide meta-analysis did not reveal additional study-wide significant (α = 6.3 × 10-7) short-tandem repeat signals, we identified seven novel suggestive Parkinson's disease short-tandem repeat risk loci (α = 5.3 × 10-6). Of these, especially a short-tandem repeat near MEIOSIN showed consistent evidence for association across datasets. CCAR2, NCOR1 and one novel suggestive locus identified here (LINC01012) emerged from colocalization analyses showing evidence for a shared causal short-tandem repeat variant affecting both Parkinson's disease risk and cis DNA methylation in brain. Larger studies, ideally using short-tandem repeats called from whole-sequencing data, are needed to more fully investigate their role in Parkinson's disease.
RESUMO
Multiple system atrophy (MSA) is an adult-onset, sporadic synucleinopathy characterized by parkinsonism, cerebellar ataxia, and dysautonomia. The genetic architecture of MSA is poorly understood, and treatments are limited to supportive measures. Here, we performed a comprehensive analysis of whole genome sequence data from 888 European-ancestry MSA cases and 7,128 controls to systematically investigate the genetic underpinnings of this understudied neurodegenerative disease. We identified four significantly associated risk loci using a genome-wide association study approach. Transcriptome-wide association analyses prioritized USP38-DT, KCTD7, and lnc-KCTD7-2 as novel susceptibility genes for MSA within these loci, and single-nucleus RNA sequence analysis found that the associated variants acted as cis-expression quantitative trait loci for multiple genes across neuronal and glial cell types. In conclusion, this study highlights the role of genetic determinants in the pathogenesis of MSA, and the publicly available data from this study represent a valuable resource for investigating synucleinopathies.
Assuntos
Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Atrofia de Múltiplos Sistemas , Atrofia de Múltiplos Sistemas/genética , Humanos , Predisposição Genética para Doença/genética , Feminino , Masculino , Idoso , Locos de Características Quantitativas/genética , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo ÚnicoRESUMO
The abnormal aggregation and accumulation of alpha-synuclein (aSyn) in the brain is a defining hallmark of synucleinopathies. Various aSyn conformations and post-translationally modified forms accumulate in pathological inclusions and vary in abundance among these disorders. Relying on antibodies that have not been assessed for their ability to detect the diverse forms of aSyn may lead to inaccurate estimations of aSyn pathology in human brains or disease models. To address this challenge, we developed and characterized an expanded antibody panel that targets different sequences and post-translational modifications along the length of aSyn, and that recognizes all monomeric, oligomeric, and fibrillar aSyn conformations. Next, we profiled aSyn pathology across sporadic and familial Lewy body diseases (LBDs) and reveal heterogeneous forms of aSyn pathology, rich in Serine 129 phosphorylation, Tyrosine 39 nitration and N- and C-terminal tyrosine phosphorylations, scattered both to neurons and glia. In addition, we show that aSyn can become hyperphosphorylated during processes of aggregation and inclusion maturation in neuronal and animal models of aSyn seeding and spreading. The validation pipeline we describe for these antibodies paves the way for systematic investigations into aSyn pathological diversity in the human brain, peripheral tissues, as well as in cellular and animal models of synucleinopathies.
RESUMO
BACKGROUND: Studies on DNA methylation (DNAm) in Alzheimer's disease (AD) have recently highlighted several genomic loci showing association with disease onset and progression. METHODS: Here, we conducted an epigenome-wide association study (EWAS) using DNAm profiles in entorhinal cortex (EC) from 149 AD patients and control brains and combined these with two previously published EC datasets by meta-analysis (total n = 337). RESULTS: We identified 12 cytosine-phosphate-guanine (CpG) sites showing epigenome-wide significant association with either case-control status or Braak's tau-staging. Four of these CpGs, located in proximity to CNFN/LIPE, TENT5A, PALD1/PRF1, and DIRAS1, represent novel findings. Integrating DNAm levels with RNA sequencing-based mRNA expression data generated in the same individuals showed significant DNAm-mRNA correlations for 6 of the 12 significant CpGs. Lastly, by calculating rates of epigenetic age acceleration using two recently proposed "epigenetic clock" estimators we found a significant association with accelerated epigenetic aging in the brains of AD patients vs. controls. CONCLUSION: In summary, our study represents the hitherto most comprehensive EWAS in AD using EC and highlights several novel differentially methylated loci with potential effects on gene expression.
Assuntos
Doença de Alzheimer , Epigenoma , Humanos , Epigênese Genética , Doença de Alzheimer/genética , Córtex Entorrinal , Ilhas de CpG , Metilação de DNA , Estudo de Associação Genômica Ampla , GTP Fosfo-Hidrolases/genética , Proteínas Supressoras de Tumor/genéticaRESUMO
The relative importance of Lewy- and Alzheimer-type pathologies to dementia in Parkinson's disease remains unclear. We have examined the combined associations of α-synuclein, tau and amyloid-ß accumulation in 56 pathologically confirmed Parkinson's disease cases, 29 of whom had developed dementia. Cortical and subcortical amyloid-ß scores were obtained, while tau and α-synuclein pathologies were rated according to the respective Braak stages. Additionally, cortical Lewy body and Lewy neurite scores were determined and Lewy body densities were generated using morphometry. Non-parametric statistics, together with regression models, receiver-operating characteristic curves and survival analyses were applied. Cortical and striatal amyloid-ß scores, Braak tau stages, cortical Lewy body, Lewy neurite scores and Lewy body densities, but not Braak α-synuclein stages, were all significantly greater in the Parkinson's disease-dementia group (P<0.05), with all the pathologies showing a significant positive correlation to each other (P<0.05). A combination of pathologies [area under the receiver-operating characteristic curve=0.95 (0.88-1.00); P<0.0001] was a better predictor of dementia than the severity of any single pathology. Additionally, cortical amyloid-ß scores (r=-0.62; P=0.043) and Braak tau stages (r=-0.52; P=0.028), but not Lewy body scores (r=-0.25; P=0.41) or Braak α-synuclein stages (r=-0.44; P=0.13), significantly correlated with mini-mental state examination scores in the subset of cases with this information available within the last year of life (n=15). High cortical amyloid-ß score (P=0.017) along with an older age at onset (P=0.001) were associated with a shorter time-to-dementia period. A combination of Lewy- and Alzheimer-type pathologies is a robust pathological correlate of dementia in Parkinson's disease, with quantitative and semi-quantitative assessment of Lewy pathology being more informative than Braak α-synuclein stages. Cortical amyloid-ß and age at disease onset seem to determine the rate to dementia.