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Besides motor symptoms, many individuals with Parkinson's disease develop cognitive impairment perhaps due to coexisting α-synuclein and Alzheimer's disease pathologies and impaired brain insulin signalling. Discovering biomarkers for cognitive impairment in Parkinson's disease could help clarify the underlying pathogenic processes and improve Parkinson's disease diagnosis and prognosis. This study used plasma samples from 273 participants: 103 Parkinson's disease individuals with normal cognition, 121 Parkinson's disease individuals with cognitive impairment (81 with mild cognitive impairment, 40 with dementia) and 49 age- and sex-matched controls. Plasma extracellular vesicles enriched for neuronal origin were immunocaptured by targeting the L1 cell adhesion molecule, then biomarkers were quantified using immunoassays. α-Synuclein was lower in Parkinson's disease compared to control individuals (P = 0.004) and in cognitively impaired Parkinson's disease individuals compared to Parkinson's disease with normal cognition (P < 0.001) and control (P < 0.001) individuals. Amyloid-ß42 did not differ between groups. Phosphorylated tau (T181) was higher in Parkinson's disease than control individuals (P = 0.003) and in cognitively impaired compared to cognitively normal Parkinson's disease individuals (P < 0.001) and controls (P < 0.001). Total tau was not different between groups. Tyrosine-phosphorylated insulin receptor substrate-1 was lower in Parkinson's disease compared to control individuals (P = 0.03) and in cognitively impaired compared to cognitively normal Parkinson's disease individuals (P = 0.02) and controls (P = 0.01), and also decreased with increasing motor symptom severity (P = 0.005); serine312-phosphorylated insulin receptor substrate-1 was not different between groups. Mechanistic target of rapamycin was not different between groups, whereas phosphorylated mechanistic target of rapamycin trended lower in cognitively impaired compared to cognitively normal Parkinson's disease individuals (P = 0.05). The ratio of α-synuclein to phosphorylated tau181 was lower in Parkinson's disease compared to controls (P = 0.001), in cognitively impaired compared to cognitively normal Parkinson's disease individuals (P < 0.001) and decreased with increasing motor symptom severity (P < 0.001). The ratio of insulin receptor substrate-1 phosphorylated serine312 to insulin receptor substrate-1 phosphorylated tyrosine was higher in Parkinson's disease compared to control individuals (P = 0.01), in cognitively impaired compared to cognitively normal Parkinson's disease individuals (P = 0.02) and increased with increasing motor symptom severity (P = 0.003). α-Synuclein, phosphorylated tau181 and insulin receptor substrate-1 phosphorylated tyrosine contributed in diagnostic classification between groups. These findings suggest that both α-synuclein and tau pathologies and impaired insulin signalling underlie Parkinson's disease with cognitive impairment. Plasma neuronal extracellular vesicles biomarkers may inform cognitive prognosis in Parkinson's disease.
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Enfermedad de Alzheimer , Disfunción Cognitiva , Insulinas , Enfermedad de Parkinson , Humanos , Enfermedad de Parkinson/complicaciones , alfa-Sinucleína , Receptor de Insulina , Proteínas tau , Péptidos beta-Amiloides , Enfermedad de Alzheimer/complicaciones , Disfunción Cognitiva/complicaciones , BiomarcadoresRESUMEN
INTRODUCTION: Neuronal extracellular vesicle (nEV) tau and insulin signaling biomarkers may detect preclinical Alzheimer's disease and age-associated cognitive decline. METHODS: This case-control study used repeated serum samples from 73 cognitively declining and 73 stable Wisconsin Registry for Alzheimer's Prevention participants (62.4 ± 6.3 years old). We immunocaptured nEVs; measured tau and insulin signaling biomarkers; and examined biomarker differences by group, their performance in group classification in training and test datasets (97, 49 individuals, respectively), and whether they predict cognitive performance change. RESULTS: Declining compared to stable individuals showed higher baseline total, p231-, and p181-tau with older age and higher annualized change for p-IR and p-IGF-1R. Combining biomarkers classified decliners with 94% area under the curve (AUC), 86.0% sensitivity and 86.7% specificity, in training data, and 75% AUC, 71.4% sensitivity, and 77.3% specificity, in test data. Insulin biomarkers predicted cognitive performance change prospectively. DISCUSSION: Combining nEV biomarkers can identify individuals with age-associated cognitive decline.
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Enfermedad de Alzheimer/diagnóstico , Biomarcadores/sangre , Disfunción Cognitiva/diagnóstico , Vesículas Extracelulares , Síntomas Prodrómicos , Enfermedad de Alzheimer/sangre , Estudios de Casos y Controles , Disfunción Cognitiva/sangre , Femenino , Humanos , Masculino , Persona de Mediana Edad , Tomografía de Emisión de Positrones , Wisconsin , Proteínas tau/sangreRESUMEN
OBJECTIVE: Mitochondrial impairments have been implicated in the pathogenesis of Fragile X-associated tremor/ataxia syndrome (FXTAS) based on analysis of mitochondria in peripheral tissues and cultured cells. We sought to assess whether mitochondrial abnormalities present in postmortem brain tissues of patients with FXTAS are also present in plasma neuron-derived extracellular vesicles (NDEVs) from living carriers of fragile X messenger ribonucleoprotein1 (FMR1) gene premutations at an early asymptomatic stage of the disease continuum. METHODS: We utilized postmortem frozen cerebellar and frontal cortex samples from a cohort of eight patients with FXTAS and nine controls and measured the quantity and activity of the mitochondrial proteins complex IV and complex V. In addition, we evaluated the same measures in isolated plasma NDEVs by selective immunoaffinity capture targeting L1CAM from a separate cohort of eight FMR1 premutation carriers and four age-matched controls. RESULTS: Lower complex IV and V quantity and activity were observed in the cerebellum of FXTAS patients compared to controls, without any differences in total mitochondrial content. No patient-control differences were observed in the frontal cortex. In NDEVs, FMR1 premutation carriers compared to controls had lower activity of Complex IV and Complex V, but higher Complex V quantity. INTERPRETATION: Quantitative and functional abnormalities in mitochondrial electron transport chain complexes IV and V seen in the cerebellum of patients with FXTAS are also manifest in plasma NDEVs of FMR1 premutation carriers. Plasma NDEVs may provide further insights into mitochondrial pathologies in this syndrome and could potentially lead to the development of biomarkers for predicting symptomatic FXTAS among premutation carriers and disease monitoring.
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Ataxia , Vesículas Extracelulares , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil , Síndrome del Cromosoma X Frágil , Mitocondrias , Temblor , Humanos , Síndrome del Cromosoma X Frágil/genética , Síndrome del Cromosoma X Frágil/metabolismo , Síndrome del Cromosoma X Frágil/patología , Síndrome del Cromosoma X Frágil/fisiopatología , Temblor/genética , Temblor/metabolismo , Temblor/fisiopatología , Temblor/patología , Vesículas Extracelulares/metabolismo , Ataxia/genética , Ataxia/metabolismo , Ataxia/patología , Ataxia/fisiopatología , Masculino , Anciano , Femenino , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/genética , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/metabolismo , Persona de Mediana Edad , Mitocondrias/metabolismo , Mitocondrias/patología , Cerebelo/metabolismo , Cerebelo/patología , Anciano de 80 o más Años , Encéfalo/metabolismo , Encéfalo/patología , Lóbulo Frontal/metabolismo , Lóbulo Frontal/patologíaRESUMEN
Isolation of neuron-derived extracellular vesicles (NDEVs) with L1 Cell Adhesion Molecule (L1CAM)-specific antibodies has been widely used to identify blood biomarkers of CNS disorders. However, full methodological validation requires demonstration of L1CAM in individual NDEVs and lower levels or absence of L1CAM in individual EVs from other cells. Here, we used multiple single-EV techniques to establish the neuronal origin and determine the abundance of L1CAM-positive EVs in human blood. L1CAM epitopes of the ectodomain are shown to be co-expressed on single-EVs with the neuronal proteins ß-III-tubulin, GAP43, and VAMP2, the levels of which increase in parallel with the enrichment of L1CAM-positive EVs. Levels of L1CAM-positive EVs carrying the neuronal proteins VAMP2 and ß-III-tubulin range from 30% to 63%, in contrast to 0.8%-3.9% of L1CAM-negative EVs. Plasma fluid-phase L1CAM does not bind to single-EVs. Our findings support the use of L1CAM as a target for isolating plasma NDEVs and leveraging their cargo to identify biomarkers reflecting neuronal function.
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Biomarcadores , Vesículas Extracelulares , Molécula L1 de Adhesión de Célula Nerviosa , Neuronas , Proteína 2 de Membrana Asociada a Vesículas , Humanos , Molécula L1 de Adhesión de Célula Nerviosa/metabolismo , Vesículas Extracelulares/metabolismo , Biomarcadores/metabolismo , Biomarcadores/sangre , Neuronas/metabolismo , Proteína 2 de Membrana Asociada a Vesículas/metabolismo , Tubulina (Proteína)/metabolismoRESUMEN
Background: Cognitive decline in Alzheimer's disease (AD) is associated with prion-like tau propagation between neurons along synaptically connected networks, in part via extracellular vesicles (EV). EV biogenesis is triggered by ceramide enrichment at the plasma membrane from neutral sphingomyelinase2(nSMase2)-mediated cleavage of sphingomyelin. We report, for the first time, that tau expression triggers an elevation in brain ceramides and nSMase2 activity. Methods: To determine the therapeutic benefit of inhibiting this elevation, we evaluated the efficacy of PDDC, the first potent, selective, orally bioavailable, and brain-penetrable nSMase2 inhibitor, in the PS19 tau transgenic AD murine model. Changes in brain ceramide and sphingomyelin levels, Tau content, histopathology, and nSMase2 target engagement were monitored, as well as changes in the number of brain-derived EVs in plasma and their Tau content. Additionally, we evaluated the ability of PDDC to impede tau propagation in a murine model where an adeno-associated virus(AAV) encoding for P301L/S320F double mutant human tau was stereotaxically-injected unilaterally into the hippocampus and the contralateral transfer to the dentate gyrus was monitored. Results: Similar to human AD, PS19 mice exhibited increased brain ceramides and nSMase2 activity; both were completely normalized by PDDC treatment. PS19 mice exhibited elevated tau immunostaining, thinning of hippocampal neuronal cell layers, increased mossy fiber synaptophysin immunostaining, and glial activation, all pathologic features of human AD. PDDC treatment significantly attenuated these aberrant changes. Mouse plasma isolated from PDDC-treated PS19 mice exhibited reduced levels of neuron- and microglia-derived EVs, the former carrying lower phosphorylated Tau(pTau) levels, compared to untreated mice. In the AAV tau propagation model, PDDC normalized the tau-induced increase in brain ceramides and significantly decreased tau spreading to the contralateral side. Conclusions: PDDC is a first-in-class therapeutic candidate that normalizes elevated brain ceramides and nSMase2 activity leading to the slowing of tau spread in AD mice.
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BACKGROUND: Cognitive decline in Alzheimer's disease (AD) is associated with hyperphosphorylated tau (pTau) propagation between neurons along synaptically connected networks, in part via extracellular vesicles (EVs). EV biogenesis is triggered by ceramide enrichment at the plasma membrane from neutral sphingomyelinase2 (nSMase2)-mediated cleavage of sphingomyelin. We report, for the first time, that human tau expression elevates brain ceramides and nSMase2 activity. METHODS: To determine the therapeutic benefit of inhibiting this elevation, we evaluated PDDC, the first potent, selective, orally bioavailable, and brain-penetrable nSMase2 inhibitor in the transgenic PS19 AD mouse model. Additionally, we directly evaluated the effect of PDDC on tau propagation in a mouse model where an adeno-associated virus (AAV) encoding P301L/S320F double mutant human tau was stereotaxically-injected unilaterally into the hippocampus. The contralateral transfer of the double mutant human tau to the dentate gyrus was monitored. We examined ceramide levels, histopathological changes, and pTau content within EVs isolated from the mouse plasma. RESULTS: Similar to human AD, the PS19 mice exhibited increased brain ceramide levels and nSMase2 activity; both were completely normalized by PDDC treatment. The PS19 mice also exhibited elevated tau immunostaining, thinning of hippocampal neuronal cell layers, increased mossy fiber synaptophysin immunostaining, and glial activation, all of which were pathologic features of human AD. PDDC treatment reduced these changes. The plasma of PDDC-treated PS19 mice had reduced levels of neuronal- and microglial-derived EVs, the former carrying lower pTau levels, compared to untreated mice. In the tau propagation model, PDDC normalized the tau-induced increase in brain ceramides and significantly reduced the amount of tau propagation to the contralateral side. CONCLUSIONS: PDDC is a first-in-class therapeutic candidate that normalizes elevated brain ceramides and nSMase2 activity, leading to the slowing of tau spread in AD mice.
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Enfermedad de Alzheimer , Animales , Humanos , Ratones , Enfermedad de Alzheimer/tratamiento farmacológico , Enfermedad de Alzheimer/genética , Ceramidas/metabolismo , Ratones Transgénicos , Neuronas/metabolismoRESUMEN
The hallmarks of Alzheimer's disease (AD) pathology are senile plaques containing amyloid-beta (Aß) and neurofibrillary tangles containing hyperphosphorylated tau. Additional pathologies often co-exist, whereas multiple pathogenic mechanisms are involved in AD, especially synaptic degeneration, which necessitate the need for synaptic integrity-related biomarkers alongside Aß- and tau-related biomarkers. Plasma neuron-derived Extracellular Vesicles EVs (NDEVs) provide biomarkers related to Aß and tau and synaptic degeneration. Here, to further establish the latter as a "liquid biopsy" for AD, we examined their relationship with ante-mortem cognition in pathologically-confirmed AD cases. We immunoprecipitated NDEVs by targeting neuronal marker L1CAM from ante-mortem plasma samples from 61 autopsy-confirmed cases of pure AD or AD with additional pathologies and measured Aß42, p181-Tau, total Tau, synaptophysin, synaptopodin and three canonical EV markers, CD63, CD81 and CD9. Higher NDEV Aß42 levels were consistently associated with better cognitive status, memory, fluency, working memory and executive function. Higher levels of NDEV synaptic integrity-related biomarkers were associated with better performance on executive function tasks. Our findings motivate the hypothesis that releasing Aß42-laden NDEVs may be an adaptive mechanism in AD.
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Enfermedad de Alzheimer , Disfunción Cognitiva , Enfermedad de Alzheimer/patología , Péptidos beta-Amiloides , Biomarcadores , Humanos , Neuronas/patología , Proteínas tauRESUMEN
Mitochondria provide energy to neurons through oxidative phosphorylation and eliminate Reactive Oxygen Species (ROS) through Superoxide Dismutase 1 (SOD1). Dysfunctional mitochondria, manifesting decreased activity of electron transport chain (ETC) complexes and high ROS levels, are involved in Alzheimer's disease (AD) pathogenesis. We hypothesized that neuronal mitochondrial dysfunction in AD is reflected in ETC and SOD1 levels and activity in plasma neuron-derived extracellular vesicles (NDEVs). We immunoprecipitated NDEVs targeting neuronal marker L1CAM from two cohorts: one including 22 individuals with early AD and 29 control subjects; and another including 14 individuals with early AD and 14 control subjects. In the first cohort, we measured levels of complexes I, III, IV, ATP synthase, and SOD1; in the second cohort, we measured levels and catalytic activity of complexes IV and ATP synthase. AD individuals had lower levels of complexes I (p < 0.0001), III (p < 0.0001), IV (p = 0.0061), and V (p < 0.0001), and SOD1 (p < 0.0001) compared to controls. AD individuals also had lower levels of catalytic activity of complex IV (p = 0.0214) and ATP synthase (p < 0.0001). NDEVs confirm quantitative and functional abnormalities in ECT complexes and SOD1 previously observed in AD models and during autopsy, opening the way for using them as biomarkers for mitochondrial dysfunction in AD.
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Mitochondrial function relies on multiple quality control mechanisms, including the release of mitochondrial vesicles. To investigate the ultrastructure and prevalence of mitochondrial membranous protrusions (and, by extension, vesicles) in neurons, we surveyed mitochondria in rat and planarian brains using transmission electron microscopy (EM). We observed that mitochondrial protrusions mostly extend from the outer membrane. Leveraging available 3D EM datasets of the brain, we further analyzed mitochondrial protrusions in neurons of mouse and Drosophila brains, identifying high-resolution spatial views of these protrusions. To assess whether the abundance of mitochondrial protrusions and mitochondria-derived vesicles respond to cellular stress, we examined neurons expressing fluorescently tagged mitochondrial markers using confocal microscopy with Airyscan and found increased numbers of mitochondrial protrusions and vesicles with mild stress. Future studies using improved spatial resolution with added temporal information may further define the functional implications of mitochondrial protrusions and vesicles in neurons.
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We have previously shown that blood astrocytic-origin extracellular vesicles (AEVs) from Alzheimer's disease (AD) patients contain high complement levels. To test the hypothesis that circulating EVs from AD patients can induce complement-mediated neurotoxicity involving Membrane Attack Complex (MAC) formation, we assessed the effects of immunocaptured AEVs (using anti-GLAST antibody), in comparison with neuronal-origin (N)EVs (using anti-L1CAM antibody), and nonspecific CD81+ EVs (using anti-CD81 antibody), from the plasma of AD, frontotemporal lobar degeneration (FTLD), and control participants. AEVs (and, less effectively, NEVs) of AD participants induced Membrane Attack Complex (MAC) expression on recipient neurons (by immunohistochemistry), membrane disruption (by EthD-1 assay), reduced neurite density (by Tuj-1 immunohistochemistry), and decreased cell viability (by MTT assay) in rat cortical neurons and human iPSC-derived neurons. Demonstration of decreased cell viability was replicated in a separate cohort of autopsy-confirmed AD patients. These effects were not produced by CD81+ EVs from AD participants or AEVs/NEVs from FTLD or control participants, and were suppressed by the MAC inhibitor CD59 and other complement inhibitors. Our results support the stated hypothesis and should motivate future studies on the roles of neuronal MAC deposition and AEV/NEV uptake, as effectors of neurodegeneration in AD.
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Astrocitos/metabolismo , Vesículas Extracelulares/inmunología , Vesículas Extracelulares/metabolismo , Neuronas/metabolismo , Animales , Antígenos CD59/metabolismo , Estudios de Casos y Controles , Células Cultivadas , Activación de Complemento/inmunología , Complejo de Ataque a Membrana del Sistema Complemento/inmunología , Complejo de Ataque a Membrana del Sistema Complemento/metabolismo , Ensayo de Inmunoadsorción Enzimática , Humanos , Células Madre Pluripotentes Inducidas/metabolismo , Masculino , RatasRESUMEN
Alzheimer's disease (AD) is the most common type of dementia. Amyloid ß (Aß) plaques, tau-containing neurofibrillary tangles, and neuronal loss leading to brain atrophy are pathologic hallmarks of AD. Given the importance of early diagnosis, extensive efforts have been undertaken to identify diagnostic and prognostic biomarkers for AD. Circulating extracellular vesicles (EVs) provide a platform for "liquid biopsy" biomarkers for AD. Here, we characterized the RNA contents of plasma EVs of age-matched individuals who were cognitively normal (healthy controls (HC)) or had mild cognitive impairment (MCI) due to AD or had mild AD dementia (AD). Using RNA sequencing analysis, we found that mitochondrial (mt)-RNAs, including MT-ND1-6 mRNAs and other protein-coding and non-coding mt-RNAs, were strikingly elevated in plasma EVs of MCI and AD individuals compared with HC. EVs secreted from cultured astrocytes, microglia, and neurons after exposure to toxic conditions relevant to AD pathogenesis (Aß aggregates and H2O2), contained mitochondrial structures (detected by electron microscopy) and mitochondrial RNA and protein. We propose that in the AD brain, toxicity-causing mitochondrial damage results in the packaging of mitochondrial components for export in EVs and further propose that mt-RNAs in plasma EVs can be diagnostic and prognostic biomarkers for MCI and AD.
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BACKGROUND: TREM2 is a transmembrane receptor that is predominantly expressed by microglia in the central nervous system. Rare variants in the TREM2 gene increase the risk for late-onset Alzheimer's disease (AD). Soluble TREM2 (sTREM2) resulting from shedding of the TREM2 ectodomain can be detected in the cerebrospinal fluid (CSF) and is a surrogate measure of TREM2-mediated microglia function. CSF sTREM2 has been previously reported to increase at different clinical stages of AD, however, alterations in relation to Amyloid ß-peptide (Aß) deposition or additional pathological processes in the amyloid cascade (such as tau pathology or neurodegeneration) remain unclear. In the current cross-sectional study, we employed the biomarker-based classification framework recently proposed by the NIA-AA consensus guidelines, in combination with clinical staging, in order to examine the CSF sTREM2 alterations at early asymptomatic and symptomatic stages of AD. METHODS: A cross-sectional study of 1027 participants of the Alzheimer's Disease Imaging Initiative (ADNI) cohort, including 43 subjects carrying TREM2 rare genetic variants, was conducted to measure CSF sTREM2 using a previously validated enzyme-linked immunosorbent assay (ELISA). ADNI participants were classified following the A/T/N framework, which we implemented based on the CSF levels of Aß1-42 (A), phosphorylated tau (T) and total tau as a marker of neurodegeneration (N), at different clinical stages defined by the clinical dementia rating (CDR) score. RESULTS: CSF sTREM2 differed between TREM2 variants, whereas the p.R47H variant had higher CSF sTREM2, p.L211P had lower CSF sTREM2 than non-carriers. We found that CSF sTREM2 increased in early symptomatic stages of late-onset AD but, unexpectedly, we observed decreased CSF sTREM2 levels at the earliest asymptomatic phase when only abnormal Aß pathology (A+) but no tau pathology or neurodegeneration (TN-), is present. CONCLUSIONS: Aß pathology (A) and tau pathology/neurodegeneration (TN) have differing associations with CSF sTREM2. While tau-related neurodegeneration is associated with an increase in CSF sTREM2, Aß pathology in the absence of downstream tau-related neurodegeneration is associated with a decrease in CSF sTREM2.
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Enfermedad de Alzheimer/líquido cefalorraquídeo , Enfermedad de Alzheimer/patología , Glicoproteínas de Membrana/líquido cefalorraquídeo , Degeneración Nerviosa/patología , Anciano , Anciano de 80 o más Años , Enfermedad de Alzheimer/genética , Péptidos beta-Amiloides/líquido cefalorraquídeo , Biomarcadores/líquido cefalorraquídeo , Estudios Transversales , Femenino , Humanos , Masculino , Glicoproteínas de Membrana/genética , Persona de Mediana Edad , Receptores Inmunológicos/genética , Proteínas tau/líquido cefalorraquídeoRESUMEN
Sulforaphane (SFN) is a natural product with cytoprotective, anti-inflammatory, and antioxidant effects. In this study, we evaluated the mechanisms of its effects on lipopolysaccharide (LPS)-induced cell death, inflammation, oxidative stress, and polarization in murine microglia. We found that SFN protects N9 microglial cells upon LPS-induced cell death and suppresses LPS-induced levels of secreted pro-inflammatory cytokines, tumor necrosis factor-alpha, interleukin-1 beta, and interleukin-6. SFN is also a potent inducer of redox sensitive transcription factor, nuclear factor erythroid 2-related factor 2 (Nrf2), which is responsible for the transcription of antioxidant, cytoprotective, and anti-inflammatory genes. SFN induced translocation of Nrf2 to the nucleus via extracellular signal-regulated kinase 1/2 (ERK1/2) pathway activation. siRNA-mediated knockdown study showed that the effects of SFN on LPS-induced reactive oxygen species, reactive nitrogen species, and pro-inflammatory cytokine production and cell death are partly Nrf2 dependent. Mox phenotype is a novel microglial phenotype that has roles in oxidative stress responses. Our results suggested that SFN induced the Mox phenotype in murine microglia through Nrf2 pathway. SFN also alleviated LPS-induced expression of inflammatory microRNA, miR-155. Finally, SFN inhibits microglia-mediated neurotoxicity as demonstrated by conditioned medium and co-culture experiments. In conclusion, SFN exerts protective effects on microglia and modulates the microglial activation state.
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Antiinflamatorios/farmacología , Antioxidantes/farmacología , Isotiocianatos/farmacología , MicroARNs/biosíntesis , Microglía/metabolismo , Factor 2 Relacionado con NF-E2/metabolismo , Animales , Elementos de Respuesta Antioxidante/fisiología , Apoptosis/efectos de los fármacos , Línea Celular , Técnicas de Cocultivo , Medios de Cultivo Condicionados/farmacología , Citocinas/metabolismo , Activación Enzimática/efectos de los fármacos , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Humanos , Inflamación/tratamiento farmacológico , Lipopolisacáridos , Factor de Transcripción MafK/metabolismo , Ratones , Factor 2 Relacionado con NF-E2/genética , Estrés Oxidativo/efectos de los fármacos , Interferencia de ARN , ARN Interferente Pequeño/genética , SulfóxidosRESUMEN
Progranulin (PGRN) is predominantly expressed by microglia in the brain, and genetic and experimental evidence suggests a critical role in Alzheimer's disease (AD). We asked whether PGRN expression is changed in a disease severity-specific manner in AD We measured PGRN in cerebrospinal fluid (CSF) in two of the best-characterized AD patient cohorts, namely the Dominant Inherited Alzheimer's Disease Network (DIAN) and the Alzheimer's Disease Neuroimaging Initiative (ADNI). In carriers of AD causing dominant mutations, cross-sectionally assessed CSF PGRN increased over the course of the disease and significantly differed from non-carriers 10 years before the expected symptom onset. In late-onset AD, higher CSF PGRN was associated with more advanced disease stages and cognitive impairment. Higher CSF PGRN was associated with higher CSF soluble TREM2 (triggering receptor expressed on myeloid cells 2) only when there was underlying pathology, but not in controls. In conclusion, we demonstrate that, although CSF PGRN is not a diagnostic biomarker for AD, it may together with sTREM2 reflect microglial activation during the disease.
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Enfermedad de Alzheimer/fisiopatología , Disfunción Cognitiva/fisiopatología , Glicoproteínas de Membrana/metabolismo , Progranulinas/líquido cefalorraquídeo , Progranulinas/metabolismo , Receptores Inmunológicos/metabolismo , Adulto , Estudios Transversales , Femenino , Humanos , Masculino , Microglía/fisiología , Persona de Mediana EdadRESUMEN
Neurotransmitters and peptide hormones are secreted into outside the cell by a vesicle fusion process. Although non-coding RNA (ncRNA) that include microRNA (miRNA) regulates gene expression inside the cell where they are transcribed, extracellular miRNA has been recently discovered outside the cells, proposing that miRNA might be released to participate in cell-to-cell communication. Despite its importance of extracellular miRNA, the molecular mechanisms by which miRNA can be stored in vesicles and released by vesicle fusion remain enigmatic. Using next-generation sequencing, vesicle purification techniques, and synthetic neurotransmission, we observe that large dense-core vesicles (LDCVs) contain a variety of miRNAs including miR-375. Furthermore, miRNA exocytosis is mediated by the SNARE complex and accelerated by Ca2+. Our results suggest that miRNA can be a novel neuromodulator that can be stored in vesicles and released by vesicle fusion together with classical neurotransmitters.
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Exocitosis , Fusión de Membrana , MicroARNs/metabolismo , Vesículas Secretoras/metabolismo , Animales , Bovinos , Ratones , Modelos Biológicos , Células PC12 , Ratas , Análisis de Secuencia de ARN , Proteína 2 de Membrana Asociada a Vesículas/metabolismoRESUMEN
Cerebrospinal fluid (CSF) biomarkers may support the diagnosis of Alzheimer's disease (AD). We studied if the diagnostic power of AD CSF biomarker concentrations, i.e., Aß42, total tau (t-tau), and phosphorylated tau (p-tau), is affected by differences in lateral ventricular volume (VV), using CSF biomarker data and magnetic resonance imaging (MRI) scans of 730 subjects, from 13 European Memory Clinics. We developed a Matlab-algorithm for standardized automated segmentation analysis of T1 weighted MRI scans in SPM8 for determining VV, and computed its ratio with total intracranial volume (TIV) as proxy for total CSF volume. The diagnostic power of CSF biomarkers (and their combination), either corrected for VV/TIV ratio or not, was determined by ROC analysis. CSF Aß42 levels inversely correlated to VV/TIV in the whole study population (Aß42: râ=â-0.28; pâ<â0.0001). For CSF t-tau and p-tau, this association only reached statistical significance in the combined MCI and AD group (t-tau: râ=â-0.15; p-tau: râ=â-0.13; both pâ<â0.01). Correction for differences in VV/TIV improved the differentiation of AD versus controls based on CSF Aß42 alone (AUC: 0.75 versus 0.81) or in combination with t-tau (AUC: 0.81 versus 0.91). In conclusion, differences in VV may be an important confounder in interpreting CSF Aß42 levels.
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Enfermedad de Alzheimer/líquido cefalorraquídeo , Enfermedad de Alzheimer/diagnóstico por imagen , Ventrículos Cerebrales/diagnóstico por imagen , Disfunción Cognitiva/líquido cefalorraquídeo , Disfunción Cognitiva/diagnóstico por imagen , Anciano , Algoritmos , Péptidos beta-Amiloides/líquido cefalorraquídeo , Área Bajo la Curva , Atrofia , Biomarcadores/líquido cefalorraquídeo , Femenino , Hipocampo/diagnóstico por imagen , Humanos , Interpretación de Imagen Asistida por Computador , Imagen por Resonancia Magnética , Masculino , Persona de Mediana Edad , Tamaño de los Órganos , Reconocimiento de Normas Patrones Automatizadas , Fragmentos de Péptidos/líquido cefalorraquídeo , Curva ROC , Proteínas tau/líquido cefalorraquídeoRESUMEN
Decreased levels of alpha-synuclein (aSyn) in cerebrospinal fluid (CSF) in Parkinson's disease and related synucleinopathies have been reported, however, not consistently in all cross-sectional studies. To test the performance of one recently released human-specific enzyme-linked immunosorbent assay (ELISA) for the quantification of aSyn in CSF, we carried out a round robin trial with 18 participating laboratories trained in CSF ELISA analyses within the BIOMARKAPD project in the EU Joint Program - Neurodegenerative Disease Research. CSF samples (homogeneous aliquots from pools) and ELISA kits (one lot) were provided centrally and data reported back to one laboratory for data analysis. Our study showed that although factors such as preanalytical sample handling and lot-to-lot variability were minimized by our study design, we identified high variation in absolute values of CSF aSyn even when the same samples and same lots of assays were applied. We further demonstrate that although absolute concentrations differ between laboratories the quantitative results are comparable. With further standardization this assay may become an attractive tool for comparing aSyn measurements in diverse settings. Recommendations for further validation experiments and improvement of the interlaboratory results obtained are given.