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1.
Aging Cell ; 21(7): e13615, 2022 07.
Artigo em Inglês | MEDLINE | ID: mdl-35662390

RESUMO

Intracellular accumulation of filamentous tau aggregates with progressive neuronal loss is a common characteristic of tauopathies. Although the neurodegenerative mechanism of tau-associated pathology remains unclear, molecular elements capable of degrading and/or sequestering neurotoxic tau species may suppress neurodegenerative progression. Here, we provide evidence that p62/SQSTM1, a ubiquitinated cargo receptor for selective autophagy, acts protectively against neuronal death and neuroinflammation provoked by abnormal tau accumulation. P301S mutant tau transgenic mice (line PS19) exhibited accumulation of neurofibrillary tangles with localization of p62 mostly in the brainstem, but neuronal loss with few neurofibrillary tangles in the hippocampus. In the hippocampus of PS19 mice, the p62 level was lower compared to the brainstem, and punctate accumulation of phosphorylated tau unaccompanied by co-localization of p62 was observed. In PS19 mice deficient in p62 (PS19/p62-KO), increased accumulation of phosphorylated tau, acceleration of neuronal loss, and exacerbation of neuroinflammation were observed in the hippocampus as compared with PS19 mice. In addition, increase of abnormal tau and neuroinflammation were observed in the brainstem of PS19/p62-KO. Immunostaining and dot-blot analysis with an antibody selectively recognizing tau dimers and higher-order oligomers revealed that oligomeric tau species in PS19/p62-KO mice were significantly accumulated as compared to PS19 mice, suggesting the requirement of p62 to eliminate disease-related oligomeric tau species. Our findings indicated that p62 exerts neuroprotection against tau pathologies by eliminating neurotoxic tau species, suggesting that the manipulative p62 and selective autophagy may provide an intrinsic therapy for the treatment of tauopathy.


Assuntos
Proteína Sequestossoma-1 , Tauopatias , Proteínas tau , Animais , Modelos Animais de Doenças , Camundongos , Camundongos Transgênicos , Emaranhados Neurofibrilares/metabolismo , Proteína Sequestossoma-1/genética , Proteína Sequestossoma-1/metabolismo , Tauopatias/metabolismo , Tauopatias/patologia , Proteínas tau/genética , Proteínas tau/metabolismo
2.
EMBO J ; 40(22): e107757, 2021 11 15.
Artigo em Inglês | MEDLINE | ID: mdl-34636430

RESUMO

Positron emission tomography (PET) allows biomolecular tracking but PET monitoring of brain networks has been hampered by a lack of suitable reporters. Here, we take advantage of bacterial dihydrofolate reductase, ecDHFR, and its unique antagonist, TMP, to facilitate in vivo imaging in the brain. Peripheral administration of radiofluorinated and fluorescent TMP analogs enabled PET and intravital microscopy, respectively, of neuronal ecDHFR expression in mice. This technique can be used to the visualize neuronal circuit activity elicited by chemogenetic manipulation in the mouse hippocampus. Notably, ecDHFR-PET allows mapping of neuronal projections in non-human primate brains, demonstrating the applicability of ecDHFR-based tracking technologies for network monitoring. Finally, we demonstrate the utility of TMP analogs for PET studies of turnover and self-assembly of proteins tagged with ecDHFR mutants. These results establish opportunities for a broad spectrum of previously unattainable PET analyses of mammalian brain circuits at the molecular level.


Assuntos
Encéfalo/diagnóstico por imagem , Tomografia por Emissão de Pósitrons/métodos , Compostos Radiofarmacêuticos/química , Tetra-Hidrofolato Desidrogenase/genética , Animais , Encéfalo/citologia , Callithrix , Radioisótopos de Carbono/química , Radioisótopos de Flúor/química , Genes Reporter , Células HEK293 , Humanos , Masculino , Camundongos Endogâmicos C57BL , Imagem Molecular/métodos , Rede Nervosa/diagnóstico por imagem , Proteínas/análise , Proteínas/metabolismo , Compostos Radiofarmacêuticos/síntese química , Tetra-Hidrofolato Desidrogenase/metabolismo , Trimetoprima/análogos & derivados , Trimetoprima/química
3.
Brain Commun ; 3(1): fcab011, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33644757

RESUMO

Microglia are the resident phagocytes of the central nervous system, and microglial activation is considered to play an important role in the pathogenesis of neurodegenerative diseases. Recent studies with single-cell RNA analysis of CNS cells in Alzheimer's disease and diverse other neurodegenerative conditions revealed that the transition from homeostatic microglia to disease-associated microglia was defined by changes of gene expression levels, including down-regulation of the P2Y12 receptor gene (P2Y12R). However, it is yet to be clarified in Alzheimer's disease brains whether and when this down-regulation occurs in response to amyloid-ß and tau depositions, which are core pathological processes in the disease etiology. To further evaluate the significance of P2Y12 receptor alterations in the neurodegenerative pathway of Alzheimer's disease and allied disorders, we generated an anti-P2Y12 receptor antibody and examined P2Y12 receptor expressions in the brains of humans and model mice bearing amyloid-ß and tau pathologies. We observed that the brains of both Alzheimer's disease and non-Alzheimer's disease tauopathy patients and tauopathy model mice (rTg4510 and PS19 mouse lines) displayed declined microglial P2Y12 receptor levels in regions enriched with tau inclusions, despite an increase in the total microglial population. Notably, diminution of microglial immunoreactivity with P2Y12 receptor was noticeable prior to massive accumulations of phosphorylated tau aggregates and neurodegeneration in rTg4510 mouse brains, despite a progressive increase of total microglial population. On the other hand, Iba1-positive microglia encompassing compact and dense-cored amyloid-ß plaques expressed P2Y12 receptor at varying levels in amyloid precursor protein (APP) mouse models (APP23 and AppNL-F/NL-F mice). By contrast, neuritic plaques in Alzheimer's disease brains were associated with P2Y12 receptor-negative microglia. These data suggest that the down-regulation of microglia P2Y12 receptor, which is characteristic of disease-associated microglia, is intimately associated with tau rather than amyloid-ß pathologies from an early stage and could be a sensitive index for neuroinflammatory responses to Alzheimer's disease-related neurodegenerative processes.

4.
J Neurosci ; 40(17): 3491-3501, 2020 04 22.
Artigo em Inglês | MEDLINE | ID: mdl-32265258

RESUMO

Synaptic dysfunction provoking dysregulated cortical neural circuits is currently hypothesized as a key pathophysiological process underlying clinical manifestations in Alzheimer's disease and related neurodegenerative tauopathies. Here, we conducted PET along with postmortem assays to investigate time course changes of excitatory and inhibitory synaptic constituents in an rTg4510 mouse model of tauopathy, which develops tau pathologies leading to noticeable brain atrophy at 5-6 months of age. Both male and female mice were analyzed in this study. We observed that radiosignals derived from [11C]flumazenil, a tracer for benzodiazepine receptor, in rTg4510 mice were significantly lower than the levels in nontransgenic littermates at 2-3 months of age. In contrast, retentions of (E)-[11C]ABP688, a tracer for mGluR5, were unaltered relative to controls at 2 months of age but then gradually declined with aging in parallel with progressive brain atrophy. Biochemical and immunohistochemical assessment of postmortem brain tissues demonstrated that inhibitory, but not excitatory, synaptic constituents selectively diminished without overt loss of somas of GABAergic interneurons in the neocortex and hippocampus of rTg4510 mice at 2 months of age, which was concurrent with enhanced immunoreactivity of cFos, a well-characterized immediate early gene, suggesting that impaired inhibitory neurotransmission may cause hyperexcitability of cortical circuits. Our findings indicate that tau-induced disruption of the inhibitory synapse may be a critical trigger of progressive neurodegeneration, resulting in massive neuronal loss, and PET assessments of inhibitory versus excitatory synapses potentially offer in vivo indices for hyperexcitability and excitotoxicity early in the etiologic pathway of neurodegenerative tauopathies.SIGNIFICANCE STATEMENT In this study, we examined the in vivo status of excitatory and inhibitory synapses in the brain of the rTg4510 tauopathy mouse model by PET imaging with (E)-[11C]ABP688 and [11C]flumazenil, respectively. We identified inhibitory synapse as being significantly dysregulated before brain atrophy at 2 months of age, while excitatory synapse stayed relatively intact at this stage. In line with this observation, postmortem assessment of brain tissues demonstrated selective attenuation of inhibitory synaptic constituents accompanied by the upregulation of cFos before the formation of tau pathology in the forebrain at young ages. Our findings indicate that selective degeneration of inhibitory synapse with hyperexcitability in the cortical circuit constitutes the critical early pathophysiology of tauopathy.


Assuntos
Doença de Alzheimer/fisiopatologia , Neurônios GABAérgicos/fisiologia , Hipocampo/fisiopatologia , Neocórtex/fisiopatologia , Sinapses/fisiologia , Tauopatias/fisiopatologia , Proteínas tau/metabolismo , Doença de Alzheimer/diagnóstico por imagem , Doença de Alzheimer/metabolismo , Animais , Modelos Animais de Doenças , Feminino , Hipocampo/diagnóstico por imagem , Hipocampo/metabolismo , Imageamento por Ressonância Magnética , Masculino , Camundongos , Camundongos Transgênicos , Neocórtex/diagnóstico por imagem , Neocórtex/metabolismo , Inibição Neural/fisiologia , Tomografia por Emissão de Pósitrons , Tauopatias/diagnóstico por imagem , Tauopatias/metabolismo
5.
J Alzheimers Dis ; 61(3): 1037-1052, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29332041

RESUMO

BACKGROUND: Tau imaging using PET is a promising tool for the diagnosis and evaluation of tau-related neurodegenerative disorders, but the relationship among PET-detectable tau, neuroinflammation, and neurodegeneration is not yet fully understood. OBJECTIVE: We aimed to elucidate sequential changes in tau accumulation, neuroinflammation, and brain atrophy by PET and MRI in a tauopathy mouse model. METHODS: rTg4510 transgenic (tg) mice expressing P301L mutated tau and non-tg mice were examined with brain MRI and PET imaging (analyzed numbers: tg = 17, non-tg = 13; age 2.5∼14 months). As PET probes, [11C]PBB3 (Pyridinyl-Butadienyl-Benzothiazole 3) and [11C]AC-5216 were used to visualize tau pathology and 18-kDa translocator protein (TSPO) neuroinflammation. Tau pathology and microglia activation were subsequently analyzed by histochemistry. RESULTS: PET studies revealed age-dependent increases in [11C]PBB3 and [11C]AC-5216 signals, which were correlated with age-dependent volume reduction in the forebrain on MRI. However, the increase in [11C]PBB3 signals reached a plateau at age 7 months, and therefore its significant correlation with [11C]AC-5216 disappeared after age 7 months. In contrast, [11C]AC-5216 showed a strong correlation with both age and volume reduction until age 14 months. Histochemical analyses confirmed the relevance of pathological tau accumulation and elevated TSPO immunoreactivity in putative microglia. CONCLUSION: Our results showed that tau accumulation is associated with neuroinflammation and brain atrophy in a tauopathy mouse model. The time-course of the [11C]PBB3- and TSPO-PET finding suggests that tau deposition triggers progressive neuroinflammation, and the sequential changes can be evaluated in vivo in mouse brains.


Assuntos
Encéfalo/diagnóstico por imagem , Encéfalo/patologia , Modelos Animais de Doenças , Microglia/metabolismo , Tauopatias/diagnóstico por imagem , Proteínas tau/metabolismo , Animais , Atrofia , Benzotiazóis , Feminino , Imageamento por Ressonância Magnética , Masculino , Camundongos , Camundongos Transgênicos , Tomografia por Emissão de Pósitrons , Receptores de GABA/metabolismo
6.
J Neurosci ; 36(45): 11544-11558, 2016 11 09.
Artigo em Inglês | MEDLINE | ID: mdl-27911758

RESUMO

Chemogenetic manipulation of neuronal activities has been enabled by a designer receptor (designer receptor exclusively activated by designer drugs, DREADD) that is activated exclusively by clozapine-N-oxide (CNO). Here, we applied CNO as a functional reporter probe to positron emission tomography (PET) of DREADD in living brains. Mutant human M4 DREADD (hM4Di) expressed in transgenic (Tg) mouse neurons was visualized by PET with microdose [11C]CNO. Deactivation of DREADD-expressing neurons in these mice by nonradioactive CNO at a pharmacological dose could also be captured by arterial spin labeling MRI (ASL-MRI). Neural progenitors derived from hM4Di Tg-induced pluripotent stem cells were then implanted into WT mouse brains and neuronal differentiation of the grafts could be imaged by [11C]CNO-PET. Finally, ASL-MRI captured chemogenetic functional manipulation of the graft neurons. Our data provide the first demonstration of multimodal molecular/functional imaging of cells expressing a functional gene reporter in the brain, which would be translatable to humans for therapeutic gene transfers and cell replacements. SIGNIFICANCE STATEMENT: The present work provides the first successful demonstration of in vivo positron emission tomographic (PET) visualization of a chemogenetic designer receptor (designer receptor exclusively activated by designer drugs, DREADD) expressed in living brains. This technology has been applied to longitudinal PET reporter imaging of neuronal grafts differentiated from induced pluripotent stem cells. Differentiated from currently used reporter genes for neuroimaging, DREADD has also been available for functional manipulation of target cells, which could be visualized by functional magnetic resonance imaging (fMRI) in a real-time manner. Multimodal imaging with PET/fMRI enables the visualization of the differentiation of iPSC-derived neural progenitors into mature neurons and DREADD-mediated functional manipulation along the time course of the graft and is accordingly capable of fortifying the utility of stem cells in cell replacement therapies.


Assuntos
Encéfalo/citologia , Genes Reporter , Células-Tronco Pluripotentes Induzidas/citologia , Imagem Multimodal/métodos , Células-Tronco Neurais/transplante , Neurônios/citologia , Neurônios/metabolismo , Animais , Encéfalo/diagnóstico por imagem , Encéfalo/metabolismo , Células Cultivadas , Humanos , Células-Tronco Pluripotentes Induzidas/transplante , Camundongos , Camundongos Transgênicos , Células-Tronco Neurais/citologia , Tomografia por Emissão de Pósitrons/métodos , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Transplante de Células-Tronco/métodos
7.
Brain Res ; 1533: 152-62, 2013 Oct 02.
Artigo em Inglês | MEDLINE | ID: mdl-23973859

RESUMO

Cyclooxygenase-2 (COX-2) plays crucial roles in progressive neuronal death in ischemic brain injury. In the present study, we evaluated two radiolabeled COX-2 selective inhibitors, [11C]celecoxib and [11C]rofecoxib, as positron emission tomography (PET) tracers for COX-2 imaging in normal and ischemic mouse brains. We also took advantage of our newly-generated antibody highly selective for mouse COX-2 to prove accumulation of the radioligands in regions enriched with COX-2. In vitro autoradiography demonstrated specific binding of high-concentration [11C]rofecoxib but not [11C]celecoxib to the cerebellum and brain stem of normal brains wherein COX-2 immunoreactivity in neurons was most abundantly observed. Meanwhile, both of these radioligands failed to detect COX-2 expression in PET assays despite their excellent brain permeability. Hypoperfusion-induced ischemia caused marked necrotic neuron death accompanied by gliosis and enhancement of neuronal COX-2 immunoreactivity in the hippocampus. Correspondingly, in vitro autoradiographic binding of [11C]rofecoxib was increased in the injured hippocampus compared to the uninjured contralateral region, but failed in living brains of ischemia model likewise. Our work provides the rationale for monitoring COX-2 as a biomarker reflecting ischemic brain injuries and demonstrates that [11C]rofecoxib, not [11C]celecoxib, is useful for in vitro assays of COX-2, but its affinity would be insufficient for in vivo PET visualization.


Assuntos
Isquemia Encefálica/diagnóstico por imagem , Encéfalo/diagnóstico por imagem , Ciclo-Oxigenase 2/metabolismo , Lactonas/administração & dosagem , Pirazóis/administração & dosagem , Sulfonamidas/administração & dosagem , Sulfonas/administração & dosagem , Animais , Anticorpos , Encéfalo/enzimologia , Isquemia Encefálica/enzimologia , Radioisótopos de Carbono , Celecoxib , Ciclo-Oxigenase 2/imunologia , Modelos Animais de Doenças , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Tomografia por Emissão de Pósitrons/métodos
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