RESUMO
Synucleinopathies are a group of diseases characterized by brain aggregates of α-synuclein (α-syn). The gradual accumulation of α-syn and the role of inflammation in early-stage pathogenesis remain poorly understood. We explored this interaction by inducing chronic inflammation in a common pre-clinical synucleinopathy mouse model. Three weeks post unilateral intra-striatal injections of human α-syn pre-formed fibrils (PFF), mice underwent repeated intraperitoneal injections of 1 mg/ml lipopolysaccharide (LPS) for 3 weeks. Histological examinations of the ipsilateral site showed phospho-α-syn regional spread and LPS-induced neutrophil recruitment to the brain vasculature. Biochemical assessment of the contralateral site confirmed spreading of α-syn aggregation to frontal cortex and a rise in intracerebral TNF-α, IL-1ß, IL-10 and KC/GRO cytokines levels due to LPS. No LPS-induced exacerbation of α-syn pathology load was observed at this stage. Proteomic analysis was performed contralateral to the PFF injection site using LC-MS/MS. Subsequent downstream Reactome Gene-Set Analysis indicated that α-syn pathology alters mitochondrial metabolism and synaptic signaling. Chronic LPS-induced inflammation further lead to an overrepresentation of pathways related to fibrin clotting as well as integrin and B cell receptor signaling. Western blotting confirmed a PFF-induced increase in fibrinogen brain levels and a PFF + LPS increase in Iba1 levels, indicating activated microglia. Splenocyte profiling revealed changes in T and B cells, monocytes, and neutrophils populations due to LPS treatment in PFF injected animals. In summary, early α-syn pathology impacts energy homeostasis pathways, synaptic signaling and brain fibrinogen levels. Concurrent mild systemic inflammation may prime brain immune pathways in interaction with peripheral immunity.
Assuntos
Encéfalo , Inflamação , Lipopolissacarídeos , alfa-Sinucleína , alfa-Sinucleína/metabolismo , Animais , Camundongos , Inflamação/metabolismo , Encéfalo/metabolismo , Encéfalo/efeitos dos fármacos , Encéfalo/patologia , Lipopolissacarídeos/farmacologia , Camundongos Endogâmicos C57BL , Masculino , Humanos , Coagulação Sanguínea/efeitos dos fármacos , Sinucleinopatias/metabolismo , Sinucleinopatias/patologia , Citocinas/metabolismo , Modelos Animais de DoençasRESUMO
This study investigates the presence of antinuclear antibodies (ANA) in three primary synucleinopathies - Parkinson's disease (PD), multiple system atrophy (MSA), and dementia with Lewy bodies (DLB), compared to healthy controls. Autoinflammatory disorders typically involve the immune system mistakenly attacking the body's own cells and start producing ANA. There is an increasing body of evidence that immune-mediated inflammation is a pathological feature linked to synucleinopathies. To investigate whether this could be autoimmune mediated we analyzed for ANA in the plasma of 25 MSA, 25 PD, and 17 DLB patients, along with 25 healthy controls, using the ANA HEp-2 indirect immunofluorescence antibody assay (ANA HEp-2 IFA). Contrary to initial expectations, results showed ANA HEp-2 positivity in 12% of PD, 8% of MSA patients, 18% of DLB patients, and 17% of healthy controls, indicating no increased prevalence of ANA in synucleinopathies compared to age-matched healthy individuals. Various ANA HEp-2 patterns were identified, but no specific pattern was associated with individual synucleinopathies. We conclude hereby that synucleinopathies are not associated with detectable presence of ANA in plasma.
RESUMO
Symptoms in the urogenital organs are common in multiple system atrophy (MSA), also in the years preceding the MSA diagnosis. It is unknown how MSA is triggered and these observations in prodromal MSA led us to hypothesize that synucleinopathy could be triggered by infection of the genitourinary tract causing É-synuclein (ÉSyn) to aggregate in peripheral nerves innervating these organs. As a first proof that peripheral infections could act as a trigger in MSA, this study focused on lower urinary tract infections (UTIs), given the relevance and high frequency of UTIs in prodromal MSA, although other types of infection might also be important triggers of MSA. We performed an epidemiological nested-case control study in the Danish population showing that UTIs are associated with future diagnosis of MSA several years after infection and that it impacts risk in both men and women. Bacterial infection of the urinary bladder triggers synucleinopathy in mice and we propose a novel role of ÉSyn in the innate immune system response to bacteria. Urinary tract infection with uropathogenic E. coli results in the de novo aggregation of ÉSyn during neutrophil infiltration. During the infection, ÉSyn is released extracellularly from neutrophils as part of their extracellular traps. Injection of MSA aggregates into the urinary bladder leads to motor deficits and propagation of ÉSyn pathology to the central nervous system in mice overexpressing oligodendroglial ÉSyn. Repeated UTIs lead to progressive development of synucleinopathy with oligodendroglial involvement in vivo. Our results link bacterial infections with synucleinopathy and show that a host response to environmental triggers can result in ÉSyn pathology that bears semblance to MSA.
Assuntos
Atrofia de Múltiplos Sistemas , Sinucleinopatias , Infecções Urinárias , Camundongos , Feminino , Animais , Sinucleinopatias/patologia , Estudos de Casos e Controles , Escherichia coli , Camundongos Transgênicos , alfa-Sinucleína , Atrofia de Múltiplos Sistemas/complicações , Atrofia de Múltiplos Sistemas/patologia , Infecções Urinárias/complicações , Imunidade InataRESUMO
BACKGROUND: Multiple system atrophy (MSA) is a rare, progressive, neurodegenerative disorder presenting glia pathology. Still, disease etiology and pathophysiology are unknown, but neuro-inflammation and vascular disruption may be contributing factors to the disease progression. Here, we performed an ex vivo deep proteome profiling of the prefrontal cortex of MSA patients to reveal disease-relevant molecular neuropathological processes. Observations were validated in plasma and cerebrospinal fluid (CSF) of novel cross-sectional patient cohorts. METHODS: Brains from 45 MSA patients and 30 normal controls (CTRLs) were included. Brain samples were homogenized and trypsinized for peptide formation and analyzed by high-performance liquid chromatography tandem mass spectrometry (LC-MS/MS). Results were supplemented by western blotting, immuno-capture, tissue clearing and 3D imaging, immunohistochemistry and immunofluorescence. Subsequent measurements of glial fibrillary acid protein (GFAP) and neuro-filament light chain (NFL) levels were performed by immunoblotting in plasma of 20 MSA patients and 20 CTRLs. Finally, we performed a proteome profiling of 144 CSF samples from MSA and CTRLs, as well as other parkinsonian disorders. Data were analyzed using relevant parametric and non-parametric two-sample tests or linear regression tests followed by post hoc tests corrected for multiple testing. Additionally, high-throughput bioinformatic analyses were applied. RESULTS: We quantified more than 4,000 proteins across samples and identified 49 differentially expressed proteins with significantly different abundances in MSA patients compared with CTRLs. Pathway analyses showed enrichment of processes related to fibrinolysis and complement cascade activation. Increased fibrinogen subunit ß (FGB) protein levels were further verified, and we identified an enriched recognition of FGB by IgGs as well as intra-parenchymal accumulation around blood vessels. We corroborated blood-brain barrier leakage by a significant increase in GFAP and NFL plasma levels in MSA patients that correlated to disease severity and/or duration. Proteome profiling of CSF samples acquired during the disease course, confirmed increased total fibrinogen levels and immune-related components in the soluble fraction of MSA patients. This was also true for the other atypical parkinsonian disorders, dementia with Lewy bodies and progressive supra-nuclear palsy, but not for Parkinson's disease patients. CONCLUSION: Our results implicate activation of the fibrinolytic cascade and immune system in the brain as contributing factors in MSA associated with a more severe disease course.
Assuntos
Atrofia de Múltiplos Sistemas , Doença de Parkinson , Transtornos Parkinsonianos , Encéfalo/metabolismo , Cromatografia Líquida , Estudos Transversais , Progressão da Doença , Fibrinogênio/metabolismo , Proteína Glial Fibrilar Ácida/metabolismo , Humanos , Atrofia de Múltiplos Sistemas/metabolismo , Doença de Parkinson/metabolismo , Transtornos Parkinsonianos/metabolismo , Transtornos Parkinsonianos/patologia , Proteoma/metabolismo , Espectrometria de Massas em TandemRESUMO
Familial Parkinson disease (PD) is associated with rare genetic mutations, but the etiology in most patients with sporadic (s)PD is largely unknown, and the basis for its progression to dementia (sPDD) is poorly characterized. We have identified that loss of IFNß or IFNAR1, the receptor for IFNα/ß, causes pathological and behavioral changes resembling PDD, prompting us to hypothesize that dysregulated genes in IFNß-IFNAR signaling pathway predispose one to sPD. By transcriptomic analysis, we found defective neuronal IFNß-IFNAR signaling, including particularly elevated PIAS2 associated with sPDD. With meta-analysis of GWASs, we identified sequence variants in IFNß-IFNAR-related genes in sPD patients. Furthermore, sPDD patients expressed higher levels of PIAS2 mRNA and protein in neurons. To determine its function in brain, we overexpressed PIAS2 under a neuronal promoter, alone or with human α-synuclein, in the brains of mice, which caused motor and cognitive impairments and correlated with intraneuronal phosphorylated (p)α-synuclein accumulation and dopaminergic neuron loss. Ectopic expression of neuronal PIAS2 blocked mitophagy, increased the accumulation of senescent mitochondrial and oxidative stress, as evidenced by excessive oxDJ1 and 8OHdG, by inactivating ERK1/2-P53 signaling. Conversely, PIAS2 knockdown rescued the clinicopathological manifestations of PDD in Ifnb-/- mice on restoring mitochondrial homeostasis, oxidative stress, and pERK1/2-pP53 signaling. The regulation of JAK-STAT2-PIAS2 signaling was crucial for neurite outgrowth and neuronal survival and excitability and thus might prevent cognitive impairments. Our findings provide insights into the progression of sPD and dementia and have implications for new therapeutic approaches.
Assuntos
Demência , Interferon beta/metabolismo , Doença de Parkinson , Proteínas Inibidoras de STAT Ativados , Transdução de Sinais , Animais , Demência/genética , Neurônios Dopaminérgicos/metabolismo , Humanos , Camundongos , Camundongos Knockout , Degeneração Neural , Doença de Parkinson/genética , Proteínas Inibidoras de STAT Ativados/genética , alfa-Sinucleína/metabolismoRESUMO
Multiple-system trophy (MSA) and Parkinson's Disease (PD) are both progressive, neurodegenerative diseases characterized by neuropathological deposition of aggregated alpha-synuclein (αSyn). The causes behind this aggregation are still unknown. We have reported aberrancies in MSA and PD patients in naturally occurring autoantibodies (nAbs) against αSyn (anti-αSyn-nAbs), which are important partakers in anti-aggregatory processes, immune-mediated clearance, and anti-inflammatory functions. To elaborate further on the timeline of autoimmune aberrancies towards αSyn, we investigated here the Immunoglobulin (Ig) affinity profile and subclass composition (IgG-total, IgG1-4 and IgM) of anti-αSyn-nAbs in serum samples from prodromal (p) phases of MSA and PD. Using an electrochemiluminescence competition immunoassay, we confirmed that the repertoire of high-affinity anti-αSyn-nAbs is significantly reduced in pMSA and pPD. Further, we demonstrated that pPD had increased anti-αSyn IgG-total levels compared to pMSA and controls, concordant with increased anti-αSyn IgG1 levels in pPD. Anti-αSyn IgG2 and IgG4 levels were reduced in pMSA and pPD compared with controls, whereas anti-αSyn IgG3 levels were reduced in pMSA compared to pPD and controls. The results indicate that the impaired reactivity towards αSyn occurs prior to disease onset. The apparent lack of high-affinity anti-αSyn nAbs may result in reduced clearance of αSyn, leading to aggregation of the protein. Thus, this study provides novel insights into possible causes behind the pathogenesis in synucleinopathies such as MSA and PD.
Assuntos
Atrofia de Múltiplos Sistemas , Doença de Parkinson , Autoanticorpos , Humanos , Imunoglobulina G , Doença de Parkinson/metabolismo , alfa-Sinucleína/metabolismoRESUMO
Aggregation of α-synuclein is a hallmark of Parkinson's disease and dementia with Lewy bodies. We here investigate the relationship between cytosolic Ca2+ and α-synuclein aggregation. Analyses of cell lines and primary culture models of α-synuclein cytopathology reveal an early phase with reduced cytosolic Ca2+ levels followed by a later Ca2+ increase. Aggregated but not monomeric α-synuclein binds to and activates SERCA in vitro, and proximity ligation assays confirm this interaction in cells. The SERCA inhibitor cyclopiazonic acid (CPA) normalises both the initial reduction and the later increase in cytosolic Ca2+ CPA protects the cells against α-synuclein-aggregate stress and improves viability in cell models and in Caenorhabditis elegans in vivo Proximity ligation assays also reveal an increased interaction between α-synuclein aggregates and SERCA in human brains affected by dementia with Lewy bodies. We conclude that α-synuclein aggregates bind SERCA and stimulate its activity. Reducing SERCA activity is neuroprotective, indicating that SERCA and down-stream processes may be therapeutic targets for treating α-synucleinopathies.
Assuntos
Cálcio/química , Cálcio/metabolismo , Citosol/química , Agregados Proteicos , ATPases Transportadoras de Cálcio do Retículo Sarcoplasmático/metabolismo , alfa-Sinucleína/metabolismo , Animais , Encéfalo/patologia , Caenorhabditis elegans , Linhagem Celular , Células Cultivadas , Retículo Endoplasmático/metabolismo , Humanos , Indóis/farmacologia , Corpos de Lewy , Masculino , Camundongos , Doença de Parkinson/patologia , Ratos , ATPases Transportadoras de Cálcio do Retículo Sarcoplasmático/antagonistas & inibidoresRESUMO
OBJECTIVE: Intestinal inflammation has been suggested to play a role in development of Parkinson's disease (PD) and multiple system atrophy (MSA). To test the hypothesis that IBD is associated with risk of PD and MSA, we performed a nationwide population-based cohort study. DESIGN: The cohort consisted of all individuals diagnosed with IBD in Denmark during 1977-2014 (n=76 477) and non-IBD individuals from the general population, who were comparable in terms of gender, age and vital status (n=7 548 259). All cohort members were followed from IBD diagnosis/index date to occurrence of PD and MSA (according to the Danish National Patient Register). RESULTS: Patients with IBD had a 22% increased risk of PD as compared with non-IBD individuals (HR=1.22; 95% CI 1.09 to 1.35). The increased risk was present independently of age at IBD diagnosis, gender or length of follow-up. The overall incidence of MSA was low in our study, and the regression analysis suggested a tendency towards higher risk of developing MSA in patients with IBD as compared with non-IBD individuals (HR=1.41; 95% CI 0.82 to 2.44). Estimates were similar for women and men. The increased risk of parkinsonism was significantly higher among patients with UC (HR=1.35; 95% CI 1.20 to 1.52) and not significantly different among patients with Crohn's disease (HR=1.12; 95% CI 0.89 to 1.40). CONCLUSIONS: This nationwide, unselected, cohort study shows a significant association between IBD and later occurrence of PD, which is consistent with recent basic scientific findings of a potential role of GI inflammation in development of parkinsonian disorders.
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Doenças Inflamatórias Intestinais/complicações , Doenças Inflamatórias Intestinais/epidemiologia , Doença de Parkinson/epidemiologia , Doença de Parkinson/etiologia , Adulto , Idoso , Dinamarca/epidemiologia , Feminino , Humanos , Incidência , Masculino , Pessoa de Meia-Idade , RiscoRESUMO
To determine the extent of neocortical involvement in multiple system atrophy (MSA), we used design-based stereological methods to estimate the total numbers of neurons, oligodendrocytes, astrocytes, and microglia in the frontal, parietal, temporal, and occipital cortex of brains from 11 patients with MSA and 11 age- and gender-matched control subjects. The stereological data were supported by cell marker expression analyses in tissue samples from the prefrontal cortex. We found significantly fewer neurons in the frontal and parietal cortex of MSA brains compared with control brains. Significantly more astrocytes and microglia were observed in the frontal, parietal, and temporal cortex of MSA brains, whereas no change in the total number of oligodendrocytes was seen in any of the neocortical regions. There were significantly fewer neurons in the frontal cortex of MSA patients with impaired executive function than in patients with normal executive function. Our results indicate that the involvement of the neocortex in MSA is far more widespread and substantial than previously thought. In addition, our results suggest that the increasingly recognized cognitive impairment in MSA may be related to neuronal loss in the frontal cortex.
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Atrofia de Múltiplos Sistemas/patologia , Neocórtex/patologia , Idoso , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Atrofia de Múltiplos Sistemas/diagnóstico por imagem , Neocórtex/diagnóstico por imagemRESUMO
Parkinson's Disease (PD) and Multiple System Atrophy (MSA) are neurodegenerative diseases characterized neuropathologically by alpha-synuclein accumulation in brain cells. This accumulation is hypothesized to contribute to constitutive neuroinflammation, and to participate in the neurodegeneration. Cytokines, which are the main inflammatory signalling molecules, have been identified in blood and cerebrospinal fluid of PD patients, but studies investigating the human brain levels are scarce. It is documented that neurotrophins, necessary for survival of brain cells and known to interact with cytokines, are altered in the basal ganglia of PD patients. In regards to MSA, no major study has investigated brain cytokine or neurotrophin protein expression. Here, we measured protein levels of 18 cytokines (IL-2, 4-8, 10, 12, 13, 17, G-CSF, GM-CSF, IFN-γ, MCP-1, MIP-1α and 1ß, TNF-α) and 5 neurotrophins (BDNF, GDNF, bFGF, PDGF-BB, VEGF) in the dorsomedial prefrontal cortex in brains of MSA and PD patients and control subjects. We found altered expression of IL-2, IL-13, and G-CSF, but no differences in neurotrophin levels. Further, in MSA patients we identified increased mRNA levels of GSK3ß that is involved in neuroinflammatory pathways. Lastly, we identified increased expression of the neurodegenerative marker S100B, but not CRP, in PD and MSA patients, indicating local rather than systemic inflammation. Supporting this, in both diseases we observed increased MHC class II+ and CD45+ positive cells, and low numbers of infiltrating CD3+ cells. In conclusion, we identified neuroinflammatory responses in PD and MSA which seems more widespread in the brain than neurotrophic changes.
Assuntos
Citocinas/metabolismo , Atrofia de Múltiplos Sistemas/imunologia , Doença de Parkinson/imunologia , Córtex Pré-Frontal/imunologia , Idoso , Estudos de Coortes , Feminino , Humanos , Imuno-Histoquímica , Inflamação/metabolismo , Inflamação/patologia , Masculino , Atrofia de Múltiplos Sistemas/patologia , Neurônios/imunologia , Neurônios/patologia , Doença de Parkinson/patologia , Córtex Pré-Frontal/patologia , RNA Mensageiro/metabolismoRESUMO
BACKGROUND: Multiple system atrophy (MSA) is a sporadic progressive neurodegenerative disorder with adult onset and unknown etiology. Clinically it is characterized by autonomic failure, cerebellar ataxia, parkinsonism, and corticospinal dysfunction in any combination and with varying severity. OBJECTIVES AND METHODS: To establish the extent of involvement of the white matter in the disease, we have used stereology to quantify the total number of neurons and glial cells (oligodendrocytes, astrocytes, and microglia) in the brains from 10 MSA patients and 11 controls. RESULTS: The mean total number of white matter interstitial neurons in the patient brains was 0.5 × 109 (coefficient of variation = standard deviation/mean = 0.37), which was significantly lower than the 1.1 × 109 (0.41) in the control brains (P = .001) and equal to a reduction by â¼50%. The patient brains had a significantly higher number of white matter microglia, 1.5 × 109 (0.47) versus 0.7 × 109 (0.39) microglia in the control subjects (P = .003) and equal to an increase by â¼ 100%. There was no significant difference in mean total numbers of white matter oligodendrocytes and astrocytes between the groups. CONCLUSIONS: We found widespread microgliosis without concomitant astrogliosis in brain white matter in MSA patients and demonstrated an absence of significant oligodendrocyte degeneration. The exact role of oligodendrocytes in MSA pathogenesis, including neurodegeneration, remains to be elucidated. © 2017 International Parkinson and Movement Disorder Society.
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Atrofia de Múltiplos Sistemas/patologia , Neuroglia , Neurônios , Substância Branca/patologia , Idoso , Contagem de Células , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Substância Branca/citologiaRESUMO
Following the failure of a Phase II clinical study evaluating human retinal pigment epithelial (hRPE) cell implants as a potential treatment option for Parkinson's disease, speculation has centered on implant function and survival as possible contributors to the therapeutic outcomes. We recently reported that neonatal hRPE cells, similar to hRPE cells used in the Phase II clinical study, produced short-lived in vitro and limited in vivo trophic factors, which supports that assumption. We hypothesize that the switch from fetal to neonatal hRPE cells, between the Phase I and the Phase II clinical trial may be partly responsible for the later negative outcomes. To investigate this hypothesis, we used two neonatal hRPE cell lots, prepared in a similar manner to neonatal hRPE cells used in the Phase II clinical study, and compared them to previously evaluated fetal hRPE cells for behavioral changes following unilateral striatal implantation in 6-hydroxydopamine-lesioned rats. The results showed that only fetal, not neonatal, hRPE cell implants, were able to improve behavioral outcomes following striatal implantation in the lesioned rats. These data suggest that fetal hRPE cells may be preferential to neonatal hRPE cells in restoring behavioral deficits.
Assuntos
Transplante de Células , Transtornos Parkinsonianos/cirurgia , Epitélio Pigmentado da Retina/citologia , Epitélio Pigmentado da Retina/embriologia , Anfetamina/farmacologia , Animais , Sobrevivência Celular , Senescência Celular , Estimulantes do Sistema Nervoso Central/farmacologia , Corpo Estriado/cirurgia , Células Epiteliais/transplante , Feminino , Humanos , Recém-Nascido , Masculino , Atividade Motora/efeitos dos fármacos , Atividade Motora/fisiologia , Oxidopamina , Transtornos Parkinsonianos/fisiopatologia , Distribuição Aleatória , Ratos Sprague-Dawley , Epitélio Pigmentado da Retina/crescimento & desenvolvimento , Caminhada/fisiologiaRESUMO
Together with Parkinson's disease (PD) and dementia with Lewy bodies, multiple system atrophy (MSA) is a member of a diverse group of neurodegenerative disorders termed α-synucleinopathies. Previously, it has been shown that α-synuclein, parkin, and synphilin-1 display disease-specific transcription patterns in frontal cortex in PD, dementia with Lewy bodies, and MSA, and thus may mediate the development of α-synucleinopathies. In this study, the differential expression of α-synuclein isoforms on transcriptional and translational levels was ascertained in MSA patients in comparison with PD cases and normal controls using isoform-specific primers and exon-specific antibodies in substantia nigra, striatum, cerebellar cortex, and nucleus dentatus. These regions are severely affected by α-synuclein pathology and neurodegeneration. Furthermore, we have also investigated transcript levels for parkin and synphilin-1 isoforms. In MSA brains, α-synuclein140 and α-synuclein 112 isoform levels were significantly increased, whereas levels of the α-synuclein 126 isoform were decreased in the substantia nigra, striatum, cerebellar cortex, and nucleus dentatus versus controls. Moreover, in MSA cases, we showed increased levels of parkin isoforms lacking the N-terminal ubiquitin-like domain and an aggregation-prone synphilin-1A isoform that causes neuronal toxicity in MSA. In PD brains, parkin transcript variant 3, 7, and 11 were significantly and specifically over-expressed in the striatum and cerebellar cortex, together with synphilin-1A and 1C. The changes of isoform expression profiles in neurodegenerative diseases suggest alterations in the regulation of transcription and/or splicing events, leading to regional/cellular events that may be important for the highly increased aggregation of α-synuclein in the brain. We report differential expression of α-synuclein, parkin, and synphilin-1 isoforms in multiple system atrophy (MSA) versus Parkinson's disease and normal control brains. We have focused on brain regions that are severely affected by α-synuclein pathology and neurodegeneration in MSA. The reported changes of isoform expression profiles suggest alterations in the regulation of transcription that may be important for aggregation of α-synuclein in the brain.
Assuntos
Encéfalo/metabolismo , Proteínas de Transporte/biossíntese , Atrofia de Múltiplos Sistemas/metabolismo , Proteínas do Tecido Nervoso/biossíntese , Ubiquitina-Proteína Ligases/biossíntese , alfa-Sinucleína/biossíntese , Idoso , Idoso de 80 Anos ou mais , Encéfalo/patologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Atrofia de Múltiplos Sistemas/patologia , Doença de Parkinson/metabolismo , Doença de Parkinson/patologia , Isoformas de Proteínas/biossínteseRESUMO
Human retinal pigment epithelial (hRPE) cell implants into the striatum have been investigated as a potential cell-based treatment for Parkinson's disease in a Phase II clinical trial that recently failed. We hypothesize that the trophic factor potential of the hRPE cells could potentially influence the function and/or survival of the implants and may be involved in an alternative mechanism of action. However, it is unclear if hRPE cells secreted trophic factors when handled in the manner used in the clinical Phase II trial. To address these questions, we investigated two neonatal hRPE cell lots, cultured in a similar manner to hRPE cells used in a Phase II clinical study, and longitudinally determined brain-derived neurotrophic factor (BDNF), fibroblast growth factor 2 (FGF2), and pigment epithelium-derived factor concentrations in vitro and following striatal implantation into 6-hydroxydopamine-lesioned rats. The results demonstrate short-lived BDNF and FGF2 concentrations in vitro from hRPE cells grown alone or attached to gelatin microcarriers (GM)s as well as limited trophic factor concentration differences in vivo following striatal implantation of hRPE-GM in 6-hydroxydopamine lesioned rats compared to sham (GM-only). The data suggest that trophic factors from neonatal hRPE cell implants likely did not participate in an alternative mechanism of action, which adds supports to a hypothesis that additional factors may have been necessary for the survival and/or function of hRPE implants and potentially the success of the Phase II clinical trial.
Assuntos
Ensaios Clínicos Fase II como Assunto , Corpo Estriado/cirurgia , Fatores de Crescimento Neural/metabolismo , Transtornos Parkinsonianos/cirurgia , Epitélio Pigmentado da Retina/transplante , Animais , Células Cultivadas , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Recém-Nascido , Masculino , Ratos , Ratos Sprague-Dawley , Epitélio Pigmentado da Retina/citologia , Epitélio Pigmentado da Retina/metabolismoRESUMO
Total numbers of neurons, oligodendrocytes, astrocytes, and microglia in the basal ganglia and red nucleus were estimated in brains from 11 patients with multiple system atrophy (MSA) and 11 age- and gender-matched control subjects with unbiased stereological methods. Compared to the control subjects, the MSA patients had a substantially lower number of neurons in the substantia nigra (p=0.001), putamen (p=0.001), and globus pallidus (p<0.001), and, to a lesser extent in the caudate nucleus (p=0.03). A significantly lower number of oligodendrocytes were only observed in the putamen (p=0.04) and globus pallidus (p=0.01). In the MSA brains the total number of astrocytes was significantly higher in the putamen (p=0.04) and caudate nucleus (p=0.01). In all examined regions a higher number of microglia were found in the MSA brains with the greatest difference observed in the otherwise unaffected red nucleus (p=0.001). The results from the stereological study were supported by cell marker expression analyses showing increased markers for activated microglia. Our results suggest that microgliosis is a consistent and severe neuropathological feature of MSA, whereas no widespread and substantial loss of oligodendrocytes was observed. We have demonstrated significant neuronal loss in the substantia nigra, striatum, and globus pallidus of patients with MSA, while neurons in other basal ganglia nuclei were spared, supporting the region-specific patterns of neuropathological changes in MSA.
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Gânglios da Base/patologia , Atrofia de Múltiplos Sistemas/patologia , Núcleo Rubro/patologia , Substância Negra/patologia , Núcleo Subtalâmico/patologia , Idoso , Idoso de 80 Anos ou mais , Astrócitos/metabolismo , Astrócitos/patologia , Gânglios da Base/metabolismo , Contagem de Células , Feminino , Humanos , Masculino , Microglia/metabolismo , Microglia/patologia , Pessoa de Meia-Idade , Atrofia de Múltiplos Sistemas/metabolismo , Neurônios/metabolismo , Neurônios/patologia , Oligodendroglia/metabolismo , Oligodendroglia/patologia , Tamanho do Órgão , Reação em Cadeia da Polimerase em Tempo Real , Núcleo Rubro/metabolismo , Substância Negra/metabolismo , Núcleo Subtalâmico/metabolismoRESUMO
Asphyctic brain injury is a major cause of neuronal inflammation in the perinatal period. Fetal asphyctic preconditioning has been shown to modulate the cerebral inflammatory cytokine response, hereby protecting the brain against asphyctic injury at birth. This study was designated to examine the effects of perinatal asphyxia and fetal asphyctic preconditioning on the inflammatory cytokine response in the cerebellum. Fetal asphyxia was induced at embryonic day 17 by clamping the uterine vasculature for 30 min. At term birth, global perinatal asphyxia was induced by placing the uterine horns in saline for 19 min. Pro- and anti-inflammatory cytokine expression were assessed by real-time PCR and immunohistochemistry in cerebella of newborn rats. We found that tumor necrosis factor alpha and interleukin-10 mRNA were increased 12 h after fetal asphyxia, while the inflammatory cytokine response was decreased 96 h postfetal asphyxia. When applied as preconditioning stimulus, fetal asphyxia attenuates the cerebellar cytokine response. These results indicate that sublethal fetal asphyxia may protect the cerebellum from perinatal asphyxia-induced damage via inhibition of inflammation.
Assuntos
Asfixia Neonatal/patologia , Cerebelo/metabolismo , Citocinas/metabolismo , Hipóxia Fetal/patologia , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Precondicionamento Isquêmico/métodos , Fatores Etários , Análise de Variância , Animais , Animais Recém-Nascidos , Citocinas/genética , Modelos Animais de Doenças , Embrião de Mamíferos , Feminino , Masculino , Gravidez , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Fatores de TempoRESUMO
Multiple sclerosis (MS) is considered to be an autoimmune disease with an unknown cause and with immune system dysregulation. Among environmental factors, viruses are most often connected with the etiology of MS. Human endogenous retroviruses (HERVs) constitute 5 to 8% of human genomic DNA and have been detected as transcripts and proteins in the central nervous system (CNS) and peripheral blood, frequently in the context of neuroinflammation. HERV-Fc1, which belongs to the HERV-H/F family, has received our attention largely because of the genetic association with MS. We studied the expression of a capsid (Gag) protein of HERV-H/F origin by flow cytometry in peripheral blood mononuclear cells (PBMCs) from healthy controls and from MS patients with nonactive or active disease. There was a significant increase in HERV-H/F Gag expression in CD4(+) (P < 0.001) and CD8(+) (P < 0.001) T lymphocytes and in monocytes (P = 0.0356) in PBMCs from MS patients with active disease. Furthermore, we have undertaken the first rigorous SYBR green-based absolute quantitative PCR (Q-PCR) evaluation approach to quantify extracellular HERV-Fc1 RNA viral loads in plasma from MS patients and healthy controls. We found a 4-fold increase in extracellular HERV-Fc1 RNA titers in patients with active MS compared with healthy controls (P < 0.001). These findings strengthen the link between HERV-Fc1 and the pathology of MS. The cause and biological consequences of these differential expression levels will be the subject of further investigation. HERV-Fc1 biology could be a compelling area for understanding the pathology of MS and possibly other autoimmune disorders.
Assuntos
Retrovirus Endógenos/genética , Esclerose Múltipla/virologia , Regulação para Cima , Adulto , Retrovirus Endógenos/classificação , Retrovirus Endógenos/isolamento & purificação , Feminino , Regulação Viral da Expressão Gênica , Produtos do Gene gag/genética , Humanos , Leucócitos Mononucleares/patologia , Leucócitos Mononucleares/virologia , Masculino , Pessoa de Meia-Idade , Esclerose Múltipla/patologia , Linfócitos T/virologia , Adulto JovemRESUMO
The family of Toll-like receptors (TLRs) plays a key role in controlling innate immune responses to a wide variety of pathogen-associated molecules. It was recently suggested that TLRs have an important role in the crosstalk between neurons and glial cells in the central nervous system, thus their deregulation may play a role in neurodegeneration. Multiple system atrophy (MSA) together with Parkinson's disease belongs to a diverse group of neurodegenerative conditions termed α-synucleinopathies. MSA is a fatal late onset disease characterized by the presence of α-synuclein positive glial cytoplasmic inclusions in oligodendrocytes. α-Synuclein can act as a danger-associated molecular pattern and alter TLR expression thereby activating inflammatory responses in the brain. In this study, using real-time PCR, we assessed the expression of TLRs (TLR1-10) in selected areas of MSA brains (substantia nigra, striatum, cerebral cortex, and nucleus dentatus) in comparison with normal controls. We show evidence for increased levels of mRNA-encoding hTLR-3, hTLR-4, and hTLR-5 in substantia nigra, striatum, cerebral cortex, and nucleus dentatus from MSA brains versus normal controls. The levels of expression of hTLR-1 mRNA were elevated in substantia nigra and striatum whereas levels of hTLR-8 and hTLR-9 mRNAs were significantly higher in cerebella from MSA patients. The concerted alteration of expression of multiple TLRs in MSA brains can be of relevance for understanding the pathogenesis of the disease.
Assuntos
Encéfalo/fisiopatologia , Atrofia de Múltiplos Sistemas/fisiopatologia , Receptores Toll-Like/biossíntese , Idoso , Idoso de 80 Anos ou mais , Encéfalo/patologia , Córtex Cerebelar/metabolismo , Núcleos Cerebelares/metabolismo , Corpo Estriado/metabolismo , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Atrofia de Múltiplos Sistemas/patologia , Neuroglia/metabolismo , Oligodendroglia/metabolismo , Oligodendroglia/patologia , Substância Negra/metabolismoRESUMO
Background: Alcohol use disorder (AUD) is a major problem of our society and is often characterized and worsened by relapse. Prolonged alcohol exposure leads to numerous biochemical alterations that, upon cessation of alcohol intake, cause an array of immediate and lasting withdrawal symptoms. Acute withdrawal and neuroinflammation can be harmful in themselves, and lasting withdrawal symptoms contribute to relapse. Here, we conducted an initial feasibility study assessing several behavioral and neurochemical factors in female C3H/HeNRj (C3H) and C57BL/6JRj (B6) mice to determine which strain showed the clearest alcohol withdrawal symptoms during long-term abstinence and neurochemical alterations following re-exposure. Methods: Female C3H and B6 mice (n = 12 per group/strain) were intermittently exposed to alcohol-containing or control liquid diets for 3 weeks. Acute and prolonged withdrawal symptoms were assessed over a period of 3 weeks using a battery of behavioral test, comprised of alcohol self-administration, anhedonia, hyperalgesia, anxiety-like and depressive-like disturbances. Brain inflammation was measured by multiplex cytokine assay. Monoamine levels in the hippocampus and striatum, as well as exploratory analyses of cations levels in the cerebellum, were assessed by High-Performance Liquid Chromatography (HPLC). Results: Both C3H and B6 alcohol-exposed mice displayed decreased saccharin intake or preference and higher stress levels assessed by ultrasonic vocalizations (USVs) recordings. B6 but not C3H alcohol-exposed mice also exhibited a slower decline of alcohol oral self-administration (OSA), hyperalgesia, elevated brain TNF-α and elevated serotonin turnover. Conclusion: Our findings highlight the suitability of the B6 strain to study the behavioral and neurochemical alterations caused by alcohol withdrawal and the potential efficacy of experimental treatments, not only in early detoxification, but also in prolonged abstinence. The feasibility of these assays is important because long-lasting withdrawal symptoms are often the main cause of relapse in alcohol-dependent patients.