RESUMO
BACKGROUND: The molecular heterogeneity of Alzheimer's amyloid-ß (Aß) deposits extends well beyond the classic Aß1-40/Aß1-42 dichotomy, substantially expanded by multiple post-translational modifications that increase the proteome diversity. Numerous truncated fragments consistently populate the brain Aß peptidome, and their homeostatic regulation and potential contribution to disease pathogenesis are largely unknown. Aß4-x peptides have been reported as major components of plaque cores and the limited studies available indicate their relative abundance in Alzheimer's disease (AD). METHODS: Immunohistochemistry was used to assess the topographic distribution of Aß4-x species in well-characterized AD cases using custom-generated monoclonal antibody 18H6-specific for Aß4-x species and blind for full-length Aß1-40/Aß1-42-in conjunction with thioflavin-S and antibodies recognizing Aßx-40 and Aßx-42 proteoforms. Circular dichroism, thioflavin-T binding, and electron microscopy evaluated the biophysical and aggregation/oligomerization properties of full-length and truncated synthetic homologues, whereas stereotaxic intracerebral injections of monomeric and oligomeric radiolabeled homologues in wild-type mice were used to evaluate their brain clearance characteristics. RESULTS: All types of amyloid deposits contained the probed Aß epitopes, albeit expressed in different proportions. Aß4-x species showed preferential localization within thioflavin-S-positive cerebral amyloid angiopathy and cored plaques, strongly suggesting poor clearance characteristics and consistent with the reduced solubility and enhanced oligomerization of their synthetic homologues. In vivo clearance studies demonstrated a fast brain efflux of N-terminally truncated and full-length monomeric forms whereas their oligomeric counterparts-particularly of Aß4-40 and Aß4-42-consistently exhibited enhanced brain retention. CONCLUSIONS: The persistence of aggregation-prone Aß4-x proteoforms likely contributes to the process of amyloid formation, self-perpetuating the amyloidogenic loop and exacerbating amyloid-mediated pathogenic pathways.
Assuntos
Doença de Alzheimer , Doença de Alzheimer/metabolismo , Peptídeos beta-Amiloides/metabolismo , Animais , Anticorpos Monoclonais , Camundongos , Fragmentos de Peptídeos , Placa Amiloide/química , Placa Amiloide/metabolismo , Placa Amiloide/patologiaRESUMO
Impaired clearance in the Alzheimer's Disease (AD) brain is key in the formation of Aß parenchymal plaques and cerebrovascular deposits known as cerebral amyloid angiopathy (CAA), present in >80% of AD patients and ~50% of non-AD elderly subjects. Aß deposits are highly heterogeneous, containing multiple fragments mostly derived from catabolism of Aß40/Aß42, which exhibit dissimilar aggregation properties. Remarkably, the role of these physiologically relevant Aß species in cerebrovascular injury and their impact in vascular pathology is unknown. We sought to understand how heterogeneous Aß species affect cerebral endothelial health and assess whether their diverse effects are associated with the peptides aggregation propensities. We analyzed cerebral microvascular endothelial cell (CMEC) viability, blood-brain barrier (BBB) permeability, and angiogenesis, all relevant aspects of brain microvascular dysfunction. We found that Aß peptides and fragments exerted differential effects on cerebrovascular pathology. Peptides forming mostly oligomeric structures induced CMEC apoptosis, whereas fibrillar aggregates increased BBB permeability without apoptotic effects. Interestingly, all Aß species tested inhibited angiogenesis in vitro. These data link the biological effects of the heterogeneous Aß peptides to their primary structure and aggregation, strongly suggesting that the composition of amyloid deposits influences clinical aspects of the AD vascular pathology. As the presence of predominant oligomeric structures in proximity of the vessel walls may lead to CMEC death and induction of microhemorrhages, fibrillar amyloid is likely responsible for increased BBB permeability and associated neurovascular dysfunction. These results have the potential to unveil more specific therapeutic targets and clarify the multifactorial nature of AD.
Assuntos
Doença de Alzheimer/fisiopatologia , Peptídeos beta-Amiloides/metabolismo , Barreira Hematoencefálica/metabolismo , Células Endoteliais/metabolismo , Diferenciação Celular , Feminino , Humanos , MasculinoRESUMO
Extensive parenchymal and vascular Aß deposits are pathological hallmarks of Alzheimer's disease (AD). Besides classic full-length peptides, biochemical analyses of brain deposits have revealed high degree of Aß heterogeneity likely resulting from the action of multiple proteolytic enzymes. In spite of the numerous studies focusing in Aß, the relevance of N- and C-terminal truncated species for AD pathogenesis remains largely understudied. In the present work, using novel antibodies specifically recognizing Aß species N-terminally truncated at position 4 or C-terminally truncated at position 34, we provide a clear assessment of the differential topographic localization of these species in AD brains and transgenic models. Based on their distinct solubility, brain N- and C-terminal truncated species were extracted by differential fractionation and identified via immunoprecipitation coupled to mass spectrometry analysis. Biochemical/biophysical studies with synthetic homologues further confirmed the different solubility properties and contrasting fibrillogenic characteristics of the truncated species composing the brain Aß peptidome. Aß C-terminal degradation leads to the production of more soluble fragments likely to be more easily eliminated from the brain. On the contrary, N-terminal truncation at position 4 favors the formation of poorly soluble, aggregation prone peptides with high amyloidogenic propensity and the potential to exacerbate the fibrillar deposits, self-perpetuating the amyloidogenic loop. Detailed assessment of the molecular diversity of Aß species composing interstitial fluid and amyloid deposits at different disease stages, as well as the evaluation of the truncation profile during various pharmacologic approaches will provide a comprehensive understanding of the still undefined contribution of Aß truncations to the disease pathogenesis and their potential as novel therapeutic targets.
Assuntos
Peptídeos beta-Amiloides/metabolismo , Encéfalo/metabolismo , Placa Amiloide/metabolismo , Placa Amiloide/patologia , Idoso , Idoso de 80 Anos ou mais , Peptídeos beta-Amiloides/química , Precursor de Proteína beta-Amiloide/química , Precursor de Proteína beta-Amiloide/metabolismo , Animais , Encéfalo/patologia , Estudos de Casos e Controles , Feminino , Humanos , Masculino , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/metabolismo , CoelhosRESUMO
We analyze the feasibility of providing Wireless Sensor Network-data-based services in an Internet of Things scenario from an economical point of view. The scenario has two competing service providers with their own private sensor networks, a network operator and final users. The scenario is analyzed as two games using game theory. In the first game, sensors decide to subscribe or not to the network operator to upload the collected sensing-data, based on a utility function related to the mean service time and the price charged by the operator. In the second game, users decide to subscribe or not to the sensor-data-based service of the service providers based on a Logit discrete choice model related to the quality of the data collected and the subscription price. The sinks and users subscription stages are analyzed using population games and discrete choice models, while network operator and service providers pricing stages are analyzed using optimization and Nash equilibrium concepts respectively. The model is shown feasible from an economic point of view for all the actors if there are enough interested final users and opens the possibility of developing more efficient models with different types of services.
RESUMO
Mitochondrial dysfunction has been recognized as an early event in Alzheimer's disease (AD) pathology, preceding and inducing neurodegeneration and memory loss. The presence of cytochrome c (CytC) released from the mitochondria into the cytoplasm is often detected after acute or chronic neurodegenerative insults, including AD. The carbonic anhydrase inhibitor (CAI) methazolamide (MTZ) was identified among a library of drugs as an inhibitor of CytC release and proved to be neuroprotective in Huntington's disease and stroke models. Here, using neuronal and glial cell cultures, in addition to an acute model of amyloid beta (Aß) toxicity, which replicates by intra-hippocampal injection the consequences of interstitial and cellular accumulation of Aß, we analyzed the effects of MTZ on neuronal and glial degeneration induced by the Alzheimer's amyloid. MTZ prevented DNA fragmentation, CytC release and activation of caspase 9 and caspase 3 induced by Aß in neuronal and glial cells in culture through the inhibition of mitochondrial hydrogen peroxide production. Moreover, intraperitoneal administration of MTZ prevented neurodegeneration induced by intra-hippocampal Aß injection in the mouse brain and was effective at reducing caspase 3 activation in neurons and microglia in the area surrounding the injection site. Our results, delineating the molecular mechanism of action of MTZ against Aß-mediated mitochondrial dysfunction and caspase activation, and demonstrating its efficiency in a model of acute amyloid-mediated toxicity, provide the first combined in vitro and in vivo evidence supporting the potential of a new therapy employing FDA-approved CAIs in AD.