RESUMO
Vascular endothelial cells play an important role in maintaining brain health, but their contribution to Alzheimer's disease (AD) is obscured by limited understanding of the cellular heterogeneity in normal aged brain and in disease. To address this, we performed single nucleus RNAseq on tissue from 32 human AD and non-AD donors (19 female, 13 male) each with five cortical regions: entorhinal cortex, inferior temporal gyrus, prefrontal cortex, visual association cortex, and primary visual cortex. Analysis of 51,586 endothelial cells revealed unique gene expression patterns across the five regions in non-AD donors. Alzheimer's brain endothelial cells were characterized by upregulated protein folding genes and distinct transcriptomic differences in response to amyloid ß plaques and cerebral amyloid angiopathy. This dataset demonstrates previously unrecognized regional heterogeneity in the endothelial cell transcriptome in both aged non-AD and AD brain.SIGNIFICANCE STATEMENT In this work, we show that vascular endothelial cells collected from five different brain regions display surprising variability in gene expression. In the presence of Alzheimer's disease pathology, endothelial cell gene expression is dramatically altered with clear differences in regional and temporal changes. These findings help explain why certain brain regions appear to differ in susceptibility to disease-related vascular remodeling events that may impact blood flow.
Assuntos
Doença de Alzheimer , Angiopatia Amiloide Cerebral , Masculino , Feminino , Humanos , Idoso , Doença de Alzheimer/metabolismo , Peptídeos beta-Amiloides/metabolismo , Células Endoteliais/metabolismo , Encéfalo/metabolismo , Angiopatia Amiloide Cerebral/genética , Placa Amiloide/patologia , Núcleo Solitário/metabolismo , Córtex Entorrinal/metabolismoRESUMO
Insoluble pathogenic proteins accumulate along blood vessels in conditions of cerebral amyloid angiopathy (CAA), exerting a toxic effect on vascular cells and impacting cerebral homeostasis. In this work, we provide new evidence from three-dimensional human brain histology that tau protein, the main component of neurofibrillary tangles, can similarly accumulate along brain vascular segments. We quantitatively assessed n = 6 Alzheimer's disease (AD), and n = 6 normal aging control brains and saw that tau-positive blood vessel segments were present in all AD cases. Tau-positive vessels are enriched for tau at levels higher than the surrounding tissue and appear to affect arterioles across cortical layers (I-V). Further, vessels isolated from these AD tissues were enriched for N-terminal tau and tau phosphorylated at T181 and T217. Importantly, tau-positive vessels are associated with local areas of increased tau neurofibrillary tangles. This suggests that accumulation of tau around blood vessels may reflect a local clearance failure. In sum, these data indicate that tau, like amyloid beta, accumulates along blood vessels and may exert a significant influence on vasculature in the setting of AD.
Assuntos
Doença de Alzheimer , Encéfalo , Emaranhados Neurofibrilares , Proteínas tau , Humanos , Doença de Alzheimer/patologia , Doença de Alzheimer/metabolismo , Proteínas tau/metabolismo , Emaranhados Neurofibrilares/patologia , Emaranhados Neurofibrilares/metabolismo , Encéfalo/patologia , Encéfalo/metabolismo , Feminino , Masculino , Idoso , Idoso de 80 Anos ou mais , Pessoa de Meia-Idade , FosforilaçãoRESUMO
Apolipoprotein E (ApoE) is a multifaceted secreted molecule synthesized in the CNS by astrocytes and microglia, and in the periphery largely by the liver. ApoE has been shown to impact the integrity of the blood-brain barrier, and, in humans, the APOE4 allele of the gene is reported to lead to a leaky blood-brain barrier. We used allele specific knock-in mice expressing each of the common (human) ApoE alleles, and longitudinal multiphoton intravital microscopy, to directly monitor the impact of various ApoE isoforms on blood-brain barrier integrity. We found that humanized APOE4, but not APOE2 or APOE3, mice show a leaky blood-brain barrier, increased MMP9, impaired tight junctions, and reduced astrocyte end-foot coverage of blood vessels. Removal of astrocyte-produced ApoE4 led to the amelioration of all phenotypes while the removal of astrocyte-produced ApoE3 had no effect on blood-brain barrier integrity. This work shows a cell specific gain of function effect of ApoE4 in the dysfunction of the BBB and implicates astrocyte production of ApoE4, possibly as a function of astrocytic end foot interactions with vessels, as a key regulator of the integrity of the blood-brain barrier.
Assuntos
Apolipoproteína E4 , Astrócitos , Humanos , Animais , Camundongos , Apolipoproteína E4/genética , Apolipoproteína E4/metabolismo , Apolipoproteína E3/genética , Astrócitos/metabolismo , Barreira Hematoencefálica/metabolismo , Metaloproteinase 9 da Matriz , Isoformas de Proteínas/metabolismoRESUMO
Tau aggregation within neurons is a critical feature of Alzheimer's disease (AD) and related tauopathies. It is believed that soluble pathologic tau species seed the formation of tau aggregates in a prion-like manner and propagate through connected neurons during the progression of disease. Both soluble and aggregated forms of tau are thought to have neurotoxic properties. In addition, different strains of misfolded tau may cause differential neurotoxicity. In this work, we present an accelerated human neuronal model of tau-induced neurotoxicity that incorporates both soluble tau species and tau aggregation. Using patient-derived induced pluripotent stem cell (iPSC) neurons expressing a tau aggregation biosensor, we develop a cell culture system that allows continuous assessment of both induced tau aggregation and neuronal viability at single-cell resolution for periods of >1 week. We show that exogenous tau "seed" uptake, as measured by tau repeat domain (TauRD) reporter aggregation, increases the risk for subsequent neuronal death in vitro These results are the first to directly visualize neuronal TauRD aggregation and subsequent cell death in single human iPSC neurons. Specific morphologic strains or patterns of TauRD aggregation are then identified and associated with differing neurotoxicity. Furthermore, we demonstrate that familial AD iPSC neurons expressing the PSEN1 L435F mutation exhibit accelerated TauRD aggregation kinetics and a tau strain propagation bias when compared with control iPSC neurons.SIGNIFICANCE STATEMENT Neuronal intracellular aggregation of the microtubule binding protein tau occurs in Alzheimer's disease and related neurodegenerative tauopathies. Tau aggregates are believed to spread from neuron to neuron via prion-like misfolded tau seeds. Our work develops a human neuronal live-imaging system to visualize seeded tau aggregation and tau-induced neurotoxicity within single neurons. Using an aggregation-sensing tau reporter, we find that neuronal uptake and propagation of tau seeds reduces subsequent survival. In addition, human induced pluripotent stem cell (iPSC) neurons carrying an Alzheimer's disease-causing mutation in presenilin-1 undergo tau seeding more rapidly than control iPSC neurons. However, they do not show subsequent differences in neuronal survival. Finally, specific morphologies of tau aggregates are associated with increased neurotoxicity.
Assuntos
Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Síndromes Neurotóxicas/patologia , Tauopatias/patologia , Proteínas tau/toxicidade , Idoso de 80 Anos ou mais , Doença de Alzheimer/genética , Doença de Alzheimer/metabolismo , Animais , Sobrevivência Celular , Células Cultivadas , Feminino , Humanos , Masculino , Camundongos , Pessoa de Meia-Idade , Presenilina-1/biossíntese , Presenilina-1/genética , Proteínas tau/genética , Proteínas tau/metabolismoRESUMO
The transition between soluble intrinsically disordered tau protein and aggregated tau in neurofibrillary tangles in Alzheimer's disease is unknown. Here, we propose that soluble tau species can undergo liquid-liquid phase separation (LLPS) under cellular conditions and that phase-separated tau droplets can serve as an intermediate toward tau aggregate formation. We demonstrate that phosphorylated or mutant aggregation prone recombinant tau undergoes LLPS, as does high molecular weight soluble phospho-tau isolated from human Alzheimer brain. Droplet-like tau can also be observed in neurons and other cells. We found that tau droplets become gel-like in minutes, and over days start to spontaneously form thioflavin-S-positive tau aggregates that are competent of seeding cellular tau aggregation. Since analogous LLPS observations have been made for FUS, hnRNPA1, and TDP43, which aggregate in the context of amyotrophic lateral sclerosis, we suggest that LLPS represents a biophysical process with a role in multiple different neurodegenerative diseases.
Assuntos
Doença de Alzheimer/metabolismo , Esclerose Lateral Amiotrófica/metabolismo , Encéfalo/metabolismo , Neurônios/metabolismo , Agregação Patológica de Proteínas/metabolismo , Proteínas tau/química , Proteínas tau/isolamento & purificação , Proteínas tau/metabolismo , Idoso de 80 Anos ou mais , Sequência de Aminoácidos , Animais , Benzotiazóis/metabolismo , Fenômenos Biofísicos , Clonagem Molecular , Proteínas de Ligação a DNA/metabolismo , Escherichia coli/genética , Feminino , Células HEK293 , Ribonucleoproteína Nuclear Heterogênea A1/metabolismo , Humanos , Extração Líquido-Líquido , Camundongos , Camundongos Transgênicos , Peso Molecular , Neuroblastoma/metabolismo , Doenças Neurodegenerativas/metabolismo , Emaranhados Neurofibrilares/metabolismo , Fosforilação , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Análise de Sequência de Proteína , Células Sf9RESUMO
Mixed pathology, with both Alzheimer's disease and vascular abnormalities, is the most common cause of clinical dementia in the elderly. While usually thought to be concurrent diseases, the fact that changes in cerebral blood flow are a prominent early and persistent alteration in Alzheimer's disease raises the possibility that vascular alterations and Alzheimer pathology are more directly linked. Here, we report that aged tau-overexpressing mice develop changes to blood vessels including abnormal, spiraling morphologies; reduced blood vessel diameters; and increased overall blood vessel density in cortex. Blood flow in these vessels was altered, with periods of obstructed flow rarely observed in normal capillaries. These changes were accompanied by cortical atrophy as well as increased expression of angiogenesis-related genes such as Vegfa, Serpine1, and Plau in CD31-positive endothelial cells. Interestingly, mice overexpressing nonmutant forms of tau in the absence of frank neurodegeneration also demonstrated similar changes. Furthermore, many of the genes we observe in mice are also altered in human RNA datasets from Alzheimer patients, particularly in brain regions classically associated with tau pathology such as the temporal lobe and limbic system regions. Together these data indicate that tau pathological changes in neurons can impact brain endothelial cell biology, altering the integrity of the brain's microvasculature.
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Doença de Alzheimer/patologia , Peptídeos beta-Amiloides/metabolismo , Indutores da Angiogênese/metabolismo , Encéfalo/irrigação sanguínea , Circulação Cerebrovascular/fisiologia , Neurônios/patologia , Proteínas tau/metabolismo , Envelhecimento , Doença de Alzheimer/genética , Doença de Alzheimer/metabolismo , Animais , Encéfalo/metabolismo , Encéfalo/patologia , Células Cultivadas , Humanos , Camundongos , Camundongos Transgênicos , Neurônios/metabolismo , Proteínas tau/genéticaRESUMO
Phosphatase and tensin homolog (PTEN) regulates synaptic density in development; however, whether PTEN also regulates synapse loss in a neurodegenerative disorder such as frontotemporal lobar degeneration with Tau deposition (FTLD-Tau) has not been explored. Here, we found that pathological Tau promotes early activation of PTEN, which precedes apoptotic caspase-3 cleavage in the rTg4510 mouse model of FTLD-Tau. We further demonstrate increased synaptic and neuronal exposure of the apoptotic signal phosphatidylserine that tags neuronal structures for microglial uptake, thereby linking PTEN activation to synaptic and neuronal structure elimination. By applying pharmacological inhibition of PTEN's protein phosphatase activity, we observed that microglial uptake can be decreased in Tau transgenic mice. Finally, we reveal a dichotomous relationship between PTEN activation and age in FTLD-Tau patients and healthy controls. Together, our findings suggest that in tauopathy, PTEN has a role in the synaptotoxicity of pathological Tau and promotes microglial removal of affected neuronal structures.
Assuntos
Microglia/metabolismo , Neurônios/patologia , PTEN Fosfo-Hidrolase/metabolismo , Sinapses/patologia , Tauopatias/patologia , Idoso , Idoso de 80 Anos ou mais , Animais , Feminino , Humanos , Masculino , Camundongos , Pessoa de Meia-Idade , Tauopatias/metabolismoRESUMO
Several studies have now supported the use of a tau lowering agent as a possible therapy in the treatment of tauopathy disorders, including Alzheimer's disease. In human Alzheimer's disease, however, concurrent amyloid-ß deposition appears to synergize and accelerate tau pathological changes. Thus far, tau reduction strategies that have been tested in vivo have been examined in the setting of tau pathology without confounding amyloid-ß deposition. To determine whether reducing total human tau expression in a transgenic model where there is concurrent amyloid-ß plaque formation can still reduce tau pathology and protect against neuronal loss, we have taken advantage of the regulatable tau transgene in APP/PS1 × rTg4510 mice. These mice develop both neurofibrillary tangles as well as amyloid-ß plaques throughout the cortex and hippocampus. By suppressing human tau expression for 6 months in the APP/PS1 × rTg4510 mice using doxycycline, AT8 tau pathology, bioactivity, and astrogliosis were reduced, though importantly to a lesser extent than lowering tau in the rTg4510 alone mice. Based on non-denaturing gels and proteinase K digestions, the remaining tau aggregates in the presence of amyloid-ß exhibit a longer-lived aggregate conformation. Nonetheless, lowering the expression of the human tau transgene was sufficient to equally ameliorate thioflavin-S positive tangles and prevent neuronal loss equally well in both the APP/PS1 × rTg4510 mice and the rTg4510 cohort. Together, these results suggest that, although amyloid-ß stabilizes tau aggregates, lowering total tau levels is still an effective strategy for the treatment of tau pathology and neuronal loss even in the presence of amyloid-ß deposition.
Assuntos
Placa Amiloide/patologia , Tauopatias/metabolismo , Proteínas tau/metabolismo , Doença de Alzheimer/patologia , Peptídeos beta-Amiloides/metabolismo , Precursor de Proteína beta-Amiloide/metabolismo , Animais , Modelos Animais de Doenças , Hipocampo/metabolismo , Humanos , Camundongos , Camundongos Transgênicos , Emaranhados Neurofibrilares/patologia , Neurônios/metabolismo , Fosforilação , Placa Amiloide/metabolismo , Presenilina-1/metabolismoRESUMO
BACKGROUND: Activation of inflammation pathways in the brain occurs in Alzheimer's disease and may contribute to the accumulation and spread of pathological proteins including tau. The goal of this study was to identify how changes in microglia, a key inflammatory cell type, may contribute to tau protein accumulation and pathology-associated changes in immune and non-immune cell processes such as neuronal degeneration, astrocyte physiology, cytokine expression, and blood vessel morphology. METHODS: We used PLX3397 (290 mg/kg), a colony-stimulating factor receptor 1 (CSF1R) inhibitor, to reduce the number of microglia in the brains of a tau-overexpressing mouse model. Mice were fed PLX3397 in chow or a control diet for 3 months beginning at 12 months of age and then were subsequently analyzed for changes in blood vessel morphology by in vivo two-photon microscopy and tissues were collected for biochemistry and histology. RESULTS: PLX3397 reduced microglial numbers by 30% regardless of genotype compared to control diet-treated mice. No change in tau burden, cortical atrophy, blood vessels, or astrocyte activation was detected. All Tg4510 mice were observed to have an increased in "disease-associated" microglial gene expression, but PLX3397 treatment did not reduce expression of these genes. Surprisingly, PLX3397 treatment resulted in upregulation of CD68 and Tgf1ß. CONCLUSIONS: Manipulating microglial activity may not be an effective strategy to combat tau pathological lesions. Higher doses of PLX3397 may be required or earlier intervention in the disease course. Overall, this indicates a need for a better understanding of specific microglial changes and their relation to the disease process.
Assuntos
Envelhecimento , Microglia/patologia , Tauopatias/patologia , Proteínas tau/genética , Proteínas tau/metabolismo , Aminopiridinas/farmacologia , Animais , Vasos Sanguíneos/patologia , Proteínas de Ligação ao Cálcio/metabolismo , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/genética , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/metabolismo , Citocinas/genética , Citocinas/metabolismo , Modelos Animais de Doenças , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Proteínas dos Microfilamentos/metabolismo , Microglia/metabolismo , Mutação/genética , Pirróis/farmacologia , RNA Mensageiro/metabolismo , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/antagonistas & inibidores , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/metabolismo , Tauopatias/genéticaRESUMO
Amyloid plaques and neurofibrillary tangles co-occur in Alzheimer disease, but with different topological and temporal patterns. Whether these two lesions are independent or pathobiologically related is uncertain. For example, amyloid deposition in the neocortex precedes the spread of tau neurofibrillary tangles from the limbic areas to the cortex. We examined the aggregation properties of tau isolated from human cases with early tau pathology (Braak II) with and without plaques. Using a well-established HEK cell biosensor assay, we show that tau from cases with plaques has an enhanced ability to induce tau aggregates compared to tau from cases without plaques. To further explore this effect, we combined mice carrying the APP/PS1 transgene array that develop plaques with rTg4510 mice carrying the P301L mutant human tau transgene that develop extensive tau pathology with age. The resulting APP/PS1-rTg4510 mice had a threefold increase in tau seeding activity over the rTg4510 strain, without change in tau production or extracellular release. Surprisingly, this effect was observed before overt amyloid deposition. The enhancement of tau aggregation was also apparent by an increase in histological measures of tau pathology in young APP/PS1-rTg4510 mice and an increase in high-molecular-weight tau. Overall, these data provide evidence that amyloid ß acts to enhance tau pathology by increasing the formation of tau species capable of seeding new aggregates.
Assuntos
Doença de Alzheimer/patologia , Peptídeos beta-Amiloides/metabolismo , Emaranhados Neurofibrilares/patologia , Placa Amiloide/patologia , Proteínas tau/metabolismo , Idoso , Idoso de 80 Anos ou mais , Doença de Alzheimer/metabolismo , Peptídeos beta-Amiloides/genética , Animais , Modelos Animais de Doenças , Feminino , Humanos , Masculino , Camundongos , Camundongos Transgênicos , Pessoa de Meia-Idade , Neocórtex/metabolismo , Neocórtex/patologia , Emaranhados Neurofibrilares/metabolismo , Fosforilação , Placa Amiloide/metabolismo , Agregação Patológica de Proteínas , Proteínas tau/genéticaRESUMO
OBJECTIVE: The 2013 Canadian Forces Mental Health Survey (CFMHS) collected detailed information on mental health problems, their impacts, occupational and nonoccupational determinants of mental health, and the use of mental health services from a random sample of 8200 serving personnel. The objective of this article is to provide a firm scientific foundation for understanding and interpreting the CFMHS findings. METHODS: This narrative review first provides a snapshot of the Canadian Armed Forces (CAF), focusing on 2 key determinants of mental health: the deployment of more than 40,000 personnel in support of the mission in Afghanistan and the extensive renewal of the CAF mental health system. The findings of recent population-based CAF mental health research are reviewed, with a focus on findings from the very similar mental health survey done in 2002. Finally, key aspects of the methods of the 2013 CFMHS are presented. RESULTS: The findings of 20 peer-reviewed publications using the 2002 mental health survey data are reviewed, along with those of 25 publications from other major CAF mental health research projects executed over the past decade. CONCLUSIONS: More than a decade of population-based mental health research in the CAF has provided a detailed picture of its mental health and use of mental health services. This knowledge base and the homology of the 2013 survey with the 2002 CAF survey and general population surveys in 2002 and 2012 will provide an unusual opportunity to use the CFMHS to situate mental health in the CAF in a historical and societal perspective.
Assuntos
Inquéritos Epidemiológicos/estatística & dados numéricos , Serviços de Saúde Mental/estatística & dados numéricos , Saúde Mental/estatística & dados numéricos , Militares/estatística & dados numéricos , Doenças Profissionais/epidemiologia , Canadá/epidemiologia , HumanosRESUMO
OBJECTIVE: More than 40,000 Canadian Armed Forces (CAF) personnel have deployed in support of the mission in Afghanistan since 2002. Over the same period, the CAF strengthened its mental health system. This article explores the effect of these events on the prevalence of past-year mental disorders over the period 2002-2013. METHOD: The data sources were 2 highly comparable population-based mental health surveys of CAF Regular Force personnel done in 2002 and 2013 (n = 5155 and 6996, respectively). Data were collected via in-person interviews with Statistics Canada personnel using the World Health Organization's Composite International Diagnostic Interview to assess past-year disorders. RESULTS: In 2013, 16.5% had 1 or more of the 6 past-year disorders assessed in the survey, with the most common conditions being major depressive episode (MDE), posttraumatic stress disorder (PTSD), and generalized anxiety disorder (GAD), which affected 8.0%, 5.3%, and 4.7%, respectively. The prevalence of PTSD, GAD, and panic disorder has increased significantly since 2002 (adjusted odds ratios from logistic regression models = 2.1, 3.0, and 1.9, respectively), while no change was seen for MDE. The comorbidity of mood and anxiety disorders increased significantly over time, being seen in 27.4% and 41.0% of those with mental disorders in 2002 and 2013, respectively. CONCLUSION: There has been an increase in the prevalence of PTSD and other anxiety disorders and of the extent of comorbidity of mood and anxiety disorders in CAF personnel over the period 2002-2013.
Assuntos
Transtornos de Ansiedade/epidemiologia , Transtorno Depressivo Maior/epidemiologia , Saúde Mental/estatística & dados numéricos , Militares/estatística & dados numéricos , Transtornos de Estresse Pós-Traumáticos/epidemiologia , Adolescente , Adulto , Idoso , Canadá/epidemiologia , Comorbidade , Feminino , Inquéritos Epidemiológicos/estatística & dados numéricos , Humanos , Masculino , Pessoa de Meia-Idade , Prevalência , Adulto JovemRESUMO
In recent years, there has been an increasing interest in the pathophysiology of repetitive concussive traumatic brain injury (rcTBI) in large part due to the association with dramatic cases of progressive neurological deterioration in professional athletes, military personnel, and others. However, our understanding of the pathophysiology of rcTBI is less advanced than for more severe brain injuries. Most prominently, the mechanisms underlying traumatic axonal injury, microglial activation, amyloid-beta accumulation, and progressive tau pathology are not yet known. In addition, the role of injury to dendritic spine cytoskeletal structures, vascular reactivity impairments, and microthrombi are intriguing and subjects of ongoing inquiry. Methods for quantitative analysis of axonal injury, dendritic injury, and synaptic loss need to be refined for the field to move forward in a rigorous fashion. We and others are attempting to develop translational approaches to assess these specific pathophysiological events in both animals and humans to facilitate clinically relevant pharmacodynamic assessments of candidate therapeutics. In this article, we review and discuss several of the recent experimental results from our lab and others. We include new initial data describing the difficulty in modeling progressive tau pathology in experimental rcTBI, and results demonstrating that sertraline can alleviate social interaction deficits and depressive-like behaviors following experimental rcTBI plus foot shock stress. Furthermore, we propose a discrete set of open, experimentally tractable questions that may serve as a framework for future investigations. In addition, we also raise several important questions that are less experimentally tractable at this time, in hopes that they may stimulate future methodological developments to address them. This article is part of a Special Issue entitled "Traumatic Brain Injury".
Assuntos
Axônios/patologia , Concussão Encefálica/fisiopatologia , Lesões Encefálicas/fisiopatologia , Encéfalo/fisiopatologia , Animais , Encéfalo/patologia , Modelos Animais de Doenças , Humanos , Microglia/patologiaRESUMO
Insoluble pathogenic proteins accumulate along blood vessels in conditions of cerebral amyloid angiopathy (CAA), exerting a toxic effect on vascular cells and impacting cerebral homeostasis. In this work we provide new evidence from three-dimensional human brain histology that tau protein, the main component of neurofibrillary tangles, can similarly accumulate along brain vascular segments. We quantitatively assessed n=6 Alzheimer's disease (AD), and n=6 normal aging control brains and saw that tau-positive blood vessel segments were present in all AD cases. Tau-positive vessels are enriched for tau at levels higher than the surrounding tissue and appear to affect arterioles across cortical layers (I-V). Further, vessels isolated from these AD tissues were enriched for N-terminal tau and tau phosphorylated at T181 and T217. Importantly, tau-positive vessels are associated with local areas of increased tau neurofibrillary tangles. This suggests that accumulation of tau around blood vessels may reflect a local clearance failure. In sum, these data indicate tau, like amyloid beta, accumulates along blood vessels and may exert a significant influence on vasculature in the setting of AD.
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White matter hyperintensities (WMHs) are commonly detected on T2-weighted magnetic resonance imaging (MRI) scans, occurring in both typical aging and Alzheimer's disease. Despite their frequent appearance and their association with cognitive decline, the molecular factors contributing to WMHs remain unclear. In this study, we investigated the transcriptomic profiles of two commonly affected brain regions with coincident AD pathology-frontal subcortical white matter (frontal-WM) and occipital subcortical white matter (occipital-WM)-and compared with age-matched healthy controls. Through RNA-sequencing in frontal- and occipital-WM bulk tissues, we identified an upregulation of genes associated with brain vasculature function in AD white matter. To further elucidate vasculature-specific transcriptomic features, we performed RNA-seq analysis on blood vessels isolated from these white matter regions, which revealed an upregulation of genes related to protein folding pathways. Finally, comparing gene expression profiles between AD individuals with high- versus low-WMH burden showed an increased expression of pathways associated with immune function. Taken together, our study characterizes the diverse molecular profiles of white matter changes in AD compared to normal aging and provides new mechanistic insights processes underlying AD-related WMHs.
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A prevailing hypothesis is that neurofibrillary tangles play a causal role in driving cognitive decline in Alzheimer's disease (AD) because tangles correlate anatomically with areas that undergo neuronal loss. We used two-photon longitudinal imaging to directly test this hypothesis and observed the fate of individual neurons in two mouse models. At any time point, neurons without tangles died at >3 times the rate as neurons with tangles. Additionally, prior to dying, they became >20% more distant from neighboring neurons across imaging sessions. Similar microstructural changes were evident in a population of non-tangle-bearing neurons in Alzheimer's donor tissues. Together, these data suggest that nonfibrillar tau puts neurons at high risk of death, and surprisingly, the presence of a tangle reduces this risk. Moreover, cortical microstructure changes appear to be a better predictor of imminent cell death than tangle status is and a promising tool for identifying dying neurons in Alzheimer's.
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Difficulty achieving complete, specific, and homogenous staining is a major bottleneck preventing the widespread use of tissue clearing techniques to image large volumes of human tissue. In this manuscript, we describe a procedure to rapidly design immunostaining protocols for antibody labeling of cleared brain tissue. We prepared libraries of 0.5-1.0 mm thick tissue sections that are fixed, pre-treated, and cleared via similar, but different procedures to optimize staining conditions for a panel of antibodies. Results from a library of mouse tissue correlate well with results from a similarly prepared library of human brain tissue, suggesting mouse tissue is an adequate substitute for protocol optimization. These data show that procedural differences do not influence every antibody-antigen pair in the same way, and minor changes can have deleterious effects, therefore, optimization should be conducted for each target. The approach outlined here will help guide researchers to successfully label a variety of targets, thus removing a major hurdle to accessing the rich 3D information available in large, cleared human tissue volumes.
Assuntos
Anticorpos , Imageamento Tridimensional , Humanos , Animais , Camundongos , Coloração e Rotulagem , Imageamento Tridimensional/métodos , EncéfaloRESUMO
Studies of post-mortem human tissue provide insight into pathological processes, but are inherently limited by practical considerations that limit the scale at which tissue can be examined, and the obvious issue that the tissue reflects only one time point in a continuous disease process. We approached this problem by adapting new tissue clearance techniques to an entire cortical area of human brain, which allows surveillance of hundreds of thousands of neurons throughout the depth of the entire cortical thickness. This approach allows detection of 'rare' events that may be difficult to detect in standard 5 micrometre-thick paraffin sections. For example, it is well established that neurofibrillary tangles begin within a neuron, and ultimately, in at least some instances, persist in the brain even after the neuron has died. These are referred to as 'ghost tangles', a term that appropriately implies their 'difficult to see' ephemeral qualities. We set out to find ghost tangles as one example of the power of the tissue clearance/image analysis techniques to detect rare events, and to learn what happens at the end-point of a tangle's life history. We were able to identify 8103 tau tangles, 132 465 neurons and 299 640 nuclei in tissue samples from three subjects with severe Alzheimer's disease (Braak V-VI) and 4 tau tangles, 200 447 neurons and 462 715 nuclei in tissue samples from three subjects with no significant tau pathology (Braak 0-I). Among these data, we located 57 ghost tangles, which makes them only 0.7% of the total tau tangles observed. We found that ghost tangles are more likely to be found in cortical layers 3 and 5 (49/57), with a select few scattered across other layers 1, 2, 4 and 6. This ability to find rare events, such as ghost tangles, in large enough quantities to statistically test their distribution exemplifies how tissue clearing can be used as a powerful tool for studying selective vulnerability or resilience to pathology across brain regions.
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BACKGROUND: The prion-like propagation of tau in neurodegenerative disorders implies that misfolded pathological tau can recruit the normal protein and template its aggregation. Here, we report the methods for the development of sensitive biosensor cell lines for the detection of tau seeding activity. RESULTS: We performed the rational design of novel tau probes based on the current structural knowledge of pathological tau aggregates in Alzheimer's disease. We generated Förster resonance energy transfer (FRET)-based biosensor stable cell lines and characterized their sensitivity, specificity, and overall ability to detect bioactive tau in human samples. As compared to the reference biosensor line, the optimized probe design resulted in an increased efficiency in the detection of tau seeding. The increased sensitivity allowed for the detection of lower amount of tau seeding competency in human brain samples, while preserving specificity for tau seeds found in Alzheimer's disease. CONCLUSIONS: This next generation of FRET-based biosensor cells is a novel tool to study tau seeding activity in Alzheimer's disease human samples, especially in samples with low levels of seeding activity, which may help studying early tau-related pathological events.
Assuntos
Doença de Alzheimer , Técnicas Biossensoriais , Tauopatias , Humanos , Doença de Alzheimer/metabolismo , Tauopatias/metabolismo , Proteínas tau/metabolismo , Encéfalo/metabolismoRESUMO
Cerebrovascular dysfunction is a significant contributor to Alzheimer's disease (AD) progression. AD mouse models show altered capillary morphology, density, and diminished blood flow in areas of tau and beta-amyloid accumulation. The purpose of this study was to examine alterations in vascular structure and their contributions to perfusion deficits in the hippocampus in AD and mild cognitive impairment (MCI). Seven individuals with AD and MCI (1 AD/6 MCI), nine cognitively intact older healthy adults, and seven younger healthy adults underwent pseudo-continuous arterial spin labeling (PCASL) and gradient-echo/spin-echo (GESE) dynamic susceptibility contrast (DSC) MRI. Cerebral blood flow (CBF), cerebral blood volume, relative vessel size index (rVSI), and mean vessel density were calculated from model fitting. Lower CBF from PCASL and SE DSC MRI was observed in the hippocampus of AD/MCI group. rVSI in the hippocampus of the AD/MCI group was larger than that of the two healthy groups (FDR-P = 0.02). No difference in vessel density was detected between the groups. We also explored relationship of tau burden from 18F-flortaucipir positron emission tomography and vascular measures from MRI. Tau burden was associated with larger vessel size and lower CBF in the hippocampus. We postulate that larger vessel size may be associated with vascular alterations in AD/MCI.