RESUMO
GRN mutations cause frontotemporal dementia (GRN-FTD) due to deficiency in progranulin (PGRN), a lysosomal and secreted protein with unclear function. Here, we found that Grn-/- mice exhibit a global deficiency in bis(monoacylglycero)phosphate (BMP), an endolysosomal phospholipid we identified as a pH-dependent PGRN interactor as well as a redox-sensitive enhancer of lysosomal proteolysis and lipolysis. Grn-/- brains also showed an age-dependent, secondary storage of glucocerebrosidase substrate glucosylsphingosine. We investigated a protein replacement strategy by engineering protein transport vehicle (PTV):PGRN-a recombinant protein linking PGRN to a modified Fc domain that binds human transferrin receptor for enhanced CNS biodistribution. PTV:PGRN rescued various Grn-/- phenotypes in primary murine macrophages and human iPSC-derived microglia, including oxidative stress, lysosomal dysfunction, and endomembrane damage. Peripherally delivered PTV:PGRN corrected levels of BMP, glucosylsphingosine, and disease pathology in Grn-/- CNS, including microgliosis, lipofuscinosis, and neuronal damage. PTV:PGRN thus represents a potential biotherapeutic for GRN-FTD.
Assuntos
Produtos Biológicos/uso terapêutico , Encéfalo/metabolismo , Doenças por Armazenamento dos Lisossomos/terapia , Progranulinas/uso terapêutico , Animais , Proteínas Morfogenéticas Ósseas/metabolismo , Endossomos/metabolismo , Feminino , Demência Frontotemporal/sangue , Demência Frontotemporal/líquido cefalorraquidiano , Gliose/complicações , Gliose/patologia , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Inflamação/patologia , Metabolismo dos Lipídeos , Lipofuscina/metabolismo , Lisossomos/metabolismo , Macrófagos/metabolismo , Masculino , Glicoproteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Microglia/metabolismo , Degeneração Neural/patologia , Fenótipo , Progranulinas/deficiência , Progranulinas/metabolismo , Receptores Imunológicos/metabolismo , Receptores da Transferrina/metabolismo , Distribuição TecidualRESUMO
Protein aggregates disrupt cellular homeostasis, causing toxicity linked to neurodegeneration. Selective autophagic elimination of aggregates is critical to protein quality control, but how aggregates are selectively targeted for degradation is unclear. We compared the requirements for autophagy receptor proteins: OPTN, NBR1, p62, NDP52, and TAX1BP1 in clearance of proteotoxic aggregates. Endogenous TAX1BP1 is recruited to and required for the clearance of stress-induced aggregates, whereas ectopic expression of TAX1BP1 increases clearance through autophagy, promoting viability of human induced pluripotent stem cell-derived neurons. In contrast, TAX1BP1 depletion sensitizes cells to several forms of aggregate-induced proteotoxicity. Furthermore, TAX1BP1 is more specifically expressed in the brain compared to other autophagy receptor proteins. In vivo, loss of TAX1BP1 results in accumulation of high molecular weight ubiquitin conjugates and premature lipofuscin accumulation in brains of young TAX1BP1 knockout mice. TAX1BP1 mediates clearance of a broad range of cytotoxic proteins indicating therapeutic potential in neurodegenerative diseases.
Assuntos
Proteínas Reguladoras de Apoptose/deficiência , Autofagia , Encéfalo/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/deficiência , Proteínas de Neoplasias/deficiência , Doenças Neurodegenerativas/metabolismo , Agregação Patológica de Proteínas/metabolismo , Animais , Proteínas Reguladoras de Apoptose/metabolismo , Encéfalo/patologia , Feminino , Células HEK293 , Células HeLa , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Lipofuscina/genética , Lipofuscina/metabolismo , Masculino , Camundongos , Camundongos Knockout , Proteínas de Neoplasias/metabolismo , Doenças Neurodegenerativas/genética , Doenças Neurodegenerativas/patologia , Agregação Patológica de Proteínas/genética , Agregação Patológica de Proteínas/patologia , Ratos , Ratos Sprague-Dawley , Ubiquitina/genética , Ubiquitina/metabolismoRESUMO
Age-related macular degeneration, Stargardt disease, and their Abca4-/- mouse model are characterized by accelerated accumulation of the pigment lipofuscin, derived from photoreceptor disc turnover in the retinal pigment epithelium (RPE); lipofuscin accumulation and retinal degeneration both occur earlier in albino mice. Intravitreal injection of superoxide (O2â¢-) generators reverses lipofuscin accumulation and rescues retinal pathology, but neither the target nor mechanism is known. Here we show that RPE contains thin multi-lamellar membranes (TLMs) resembling photoreceptor discs, which associate with melanolipofuscin granules in pigmented mice but in albinos are 10-fold more abundant and reside in vacuoles. Genetically over-expressing tyrosinase in albinos generates melanosomes and decreases TLM-related lipofuscin. Intravitreal injection of generators of O2â¢- or nitric oxide (â¢NO) decreases TLM-related lipofuscin in melanolipofuscin granules of pigmented mice by ~50% in 2 d, but not in albinos. Prompted by evidence that O2â¢- plus â¢NO creates a dioxetane on melanin that excites its electrons to a high-energy state (termed "chemiexcitation"), we show that exciting electrons directly using a synthetic dioxetane reverses TLM-related lipofuscin even in albinos; quenching the excited-electron energy blocks this reversal. Melanin chemiexcitation assists in safe photoreceptor disc turnover.
Assuntos
Degeneração Macular , Melaninas , Camundongos , Animais , Melaninas/metabolismo , Lipofuscina/metabolismo , Degeneração Macular/prevenção & controle , Degeneração Macular/patologia , Retina/metabolismo , Epitélio Pigmentado da Retina/metabolismo , Transportadores de Cassetes de Ligação de ATPRESUMO
Dry age-related macular degeneration (AMD) is a prevalent clinical condition that leads to permanent damage to central vision and poses a significant threat to patients' visual health. Although the pathogenesis of dry AMD remains unclear, there is consensus on the role of retinal pigment epithelium (RPE) damage. Oxidative stress and chronic inflammation are major contributors to RPE cell damage, and the NOD-like receptor thermoprotein structural domain-associated protein 3 (NLRP3) inflammasome mediates the inflammatory response leading to apoptosis in RPE cells. Furthermore, lipofuscin accumulation results in oxidative stress, NLRP3 activation, and the development of vitelliform lesions, a hallmark of dry AMD, all of which may contribute to RPE dysfunction. The process of autophagy, involving the encapsulation, recognition, and transport of accumulated proteins and dead cells to the lysosome for degradation, is recognized as a significant pathway for cellular self-protection and homeostasis maintenance. Recently, RPE cell autophagy has been discovered to be closely linked to the development of macular degeneration, positioning autophagy as a cutting-edge research area in the realm of dry AMD. In this review, we present an overview of how lipofuscin, oxidative stress, and the NLRP3 inflammasome damage the RPE through their respective causal mechanisms. We summarized the connection between autophagy, oxidative stress, and NLRP3 inflammatory cytokines. Our findings suggest that targeting autophagy improves RPE function and sustains visual health, offering new perspectives for understanding the pathogenesis and clinical management of dry AMD.
Assuntos
Autofagia , Estresse Oxidativo , Epitélio Pigmentado da Retina , Humanos , Epitélio Pigmentado da Retina/metabolismo , Epitélio Pigmentado da Retina/patologia , Autofagia/fisiologia , Estresse Oxidativo/fisiologia , Inflamassomos/metabolismo , Lipofuscina/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Atrofia Geográfica/metabolismo , Atrofia Geográfica/patologiaRESUMO
The gradual nature of age-related neurodegeneration causes Parkinson's disease (PD) and impairs movement, memory, intellectual ability, and social interaction. One of the most prevalent neurodegenerative conditions affecting the central nervous system (CNS) among the elderly is PD. PD affects both motor and cognitive functions. Degeneration of dopaminergic (DA) neurons and buildup of the protein α-synuclein (α-Syn) in the substantia nigra pars compacta (SNpc) are two major causes of this disorder. Both UPS and ALS systems serve to eliminate α-Syn. Autophagy and UPS deficits, shortened life duration, and lipofuscin buildup accelerate PD. This sickness has no cure. Innovative therapies are halting PD progression. Bioactive phytochemicals may provide older individuals with a natural substitute to help delay the onset of neurodegenerative illnesses. This study examines whether nicotine helps transgenic C. elegans PD models. According to numerous studies, nicotine enhances synaptic plasticity and dopaminergic neuronal survival. Upgrades UPS pathways, increases autophagy, and decreases oxidative stress and mitochondrial dysfunction. At 100, 150, and 200 µM nicotine levels, worms showed reduced α-Syn aggregation, repaired DA neurotoxicity after 6-OHDA intoxication, increased lifetime, and reduced lipofuscin accumulation. Furthermore, nicotine triggered autophagy and UPS. We revealed nicotine's potential as a UPS and autophagy activator to prevent PD and other neurodegenerative diseases.
Assuntos
Doenças Neurodegenerativas , Doença de Parkinson , Animais , Humanos , Idoso , Doença de Parkinson/tratamento farmacológico , Doença de Parkinson/metabolismo , Nicotina/farmacologia , Nicotina/metabolismo , Caenorhabditis elegans/metabolismo , Lipofuscina/metabolismo , Lipofuscina/farmacologia , alfa-Sinucleína/metabolismo , alfa-Sinucleína/farmacologia , Doenças Neurodegenerativas/metabolismo , Neurônios Dopaminérgicos/metabolismo , AutofagiaRESUMO
INTRODUCTION: While several antidepressants have been identified as potential geroprotectors, the effect and mechanism of sertraline on healthspan remain to be elucidated. Here, we explored the role of sertraline in the lifespan and healthspan of Caenorhabditis elegans. METHODS: The optimal effect concentration of sertraline was first screened in wild-type N2 worms under heat stress conditions. Then, we examined the effects of sertraline on lifespan, reproduction, lipofuscin accumulation, mobility, and stress resistance. Finally, the expression of serotonin signaling and aging-related genes was investigated to explore the underlying mechanism, and the lifespan assays were performed in ser-7 RNAi strain, daf-2, daf-16, and aak-2 mutants. RESULTS: Sertraline extended the lifespan in C. elegans with concomitant extension of healthspan as indicated by increasing mobility and reducing fertility and lipofuscin accumulation, as well as enhanced resistance to different abiotic stresses. Mechanistically, ser-7 orchestrated sertraline-induced longevity via the regulation of insulin and AMPK pathways, and sertraline-induced lifespan extension in nematodes was abolished in ser-7 RNAi strain, daf-2, daf-16, and aak-2 mutants. CONCLUSION: Sertraline promotes health and longevity in C. elegans through ser-7-insulin/AMPK pathways.
Assuntos
Proteínas de Caenorhabditis elegans , Caenorhabditis elegans , Animais , Caenorhabditis elegans/genética , Longevidade/fisiologia , Sertralina/farmacologia , Sertralina/metabolismo , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas Quinases Ativadas por AMP/metabolismo , Lipofuscina/metabolismo , Lipofuscina/farmacologia , Insulina , Fatores de Transcrição Forkhead/genéticaRESUMO
Lipofuscin granules enclose mixtures of cross-linked proteins and lipids in proportions that depend on the tissue analyzed. Retinal lipofuscin is unique in that it contains mostly lipids with very little proteins. However, retinal lipofuscin also presents biological and physicochemical characteristics indistinguishable from conventional granules, including indigestibility, tendency to cause lysosome swelling that results in rupture or defective functions, and ability to trigger NLRP3 inflammation, a symptom of low-level disruption of lysosomes. In addition, like conventional lipofuscins, it appears as an autofluorescent pigment, considered toxic waste, and a biomarker of aging. Ocular lipofuscin accumulates in the retinal pigment epithelium (RPE), whereby it interferes with the support of the neuroretina. RPE cell death is the primary cause of blindness in the most prevalent incurable genetic and age-related human disorders, Stargardt disease and age-related macular degeneration (AMD), respectively. Although retinal lipofuscin is directly linked to the cell death of the RPE in Stargardt, the extent to which it contributes to AMD is a matter of debate. Nonetheless, the number of AMD clinical trials that target lipofuscin formation speaks for the potential relevance for AMD as well. Here, we show that retinal lipofuscin triggers an atypical necroptotic cascade, amenable to pharmacological intervention. This pathway is distinct from canonic necroptosis and is instead dependent on the destabilization of lysosomes. We also provide evidence that necroptosis is activated in aged human retinas with AMD. Overall, this cytotoxicity mechanism may offer therapeutic targets and markers for genetic and age-related diseases associated with lipofuscin buildups.
Assuntos
Membranas Intracelulares/metabolismo , Lipofuscina/farmacologia , Lisossomos/metabolismo , Necroptose/efeitos dos fármacos , Transportadores de Cassetes de Ligação de ATP/genética , Transportadores de Cassetes de Ligação de ATP/metabolismo , Envelhecimento , Oxirredutases do Álcool , Animais , Morte Celular , Humanos , Lipofuscina/metabolismo , Degeneração Macular/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Retina/metabolismo , Epitélio Pigmentado da Retina/metabolismoRESUMO
The aging ovary in mammals leads to the reduced production of sex hormones and a deterioration in follicle quality. The interstitial gland originates from the hypertrophy of the theca cells of atretic follicles and represents an accumulative structure of the ovary that may contribute to its aging. Here, reproductive and mature rabbit ovaries are used to determine whether the interstitial gland plays a crucial role in ovarian aging. We demonstrate that, in the mature ovary, interstitial gland cells accumulate lipid droplets and show ultrastructural characteristics of lipophagy. Furthermore, they undergo modifications and present a foamy appearance, do not express the pan-leukocyte CD-45 marker, and express CYP11A1. These cells are the first to present an increase in lipofuscin accumulation. In foamy cells, the expression of p21 remains low, PCNA expression is maintained at mature ages, and their nuclei do not show positivity for H2AX. The interstitial gland shows a significant increase in lipofuscin accumulation compared with the ovaries of younger rabbits, but lipofuscin accumulation remains constant at mature ages. Surprisingly, no accumulation of cells with DNA damage is evident, and an increase in proliferative cells is observed at the age of 36 months. We suggest that the interstitial gland initially uses lipophagy to maintain steroidogenic homeostasis and prevent cellular senescence.
Assuntos
Envelhecimento , Senescência Celular , Lipofuscina , Ovário , Animais , Feminino , Coelhos , Envelhecimento/metabolismo , Ovário/metabolismo , Ovário/citologia , Lipofuscina/metabolismo , Chinchila , Células Tecais/metabolismo , Folículo Ovariano/metabolismo , Folículo Ovariano/citologia , Enzima de Clivagem da Cadeia Lateral do Colesterol/metabolismo , Enzima de Clivagem da Cadeia Lateral do Colesterol/genética , Dano ao DNARESUMO
Accumulation of autofluorescent waste products, amyloids, and products of lipid peroxidation (LPO) are important hallmarks of aging. Until now, these processes have not been documented in Daphnia, a convenient model organism for longevity and senescence studies. We conducted a longitudinal cohort study of autofluorescence and Congo Red (CR) fluorescent staining for amyloids in four clones of D. magna. Additionally, we used a single time point cross-sectional common garden experiment within a single clone in which autofluorescence and BODIPY C11 fluorescence were measured. We observed a robust increase in autofluorescent spots that show diagnostic co-staining by Sudan Black indicating lipofuscin aggregates, particularly in the upper body region. There was also a significant clone-by-age interaction indicating that some genotypes accumulated lipofuscins faster than others. Contrary to predictions, CR fluorescence and lipid peroxidation did not consistently increase with age. CR fluorescence demonstrated a slight non-monotonous relationship with age, achieving the highest values at intermediate ages, possibly due to elimination of physiological heterogeneity in our genetically uniform cohorts. LPO demonstrated a significant ovary status-by-age interaction, decreasing with age when measured in Daphnia with full ovaries (late phase ovarian cycle) and showing no significant trend or slight increase with age when measured during the early phase in the ovarian cycle.
Assuntos
Daphnia , Lipofuscina , Animais , Feminino , Lipofuscina/metabolismo , Peroxidação de Lipídeos/fisiologia , Daphnia/metabolismo , Estudos Longitudinais , Estudos Transversais , Envelhecimento/fisiologiaRESUMO
Age-related macular degeneration (AMD) is the leading cause of blindness in the elderly. While the histopathology of the different disease stages is well characterized, the cause underlying the progression, from the early drusen stage to the advanced macular degeneration stage that leads to blindness, remains unknown. Here, we show that photoreceptors (PRs) of diseased individuals display increased expression of two key glycolytic genes, suggestive of a glucose shortage during disease. Mimicking aspects of this metabolic profile in PRs of wild-type mice by activation of the mammalian target of rapamycin complex 1 (mTORC1) caused early drusen-like pathologies, as well as advanced AMD-like pathologies. Mice with activated mTORC1 in PRs also displayed other early disease features, such as a delay in photoreceptor outer segment (POS) clearance and accumulation of lipofuscin in the retinal-pigmented epithelium (RPE) and of lipoproteins at the Bruch's membrane (BrM), as well as changes in complement accumulation. Interestingly, formation of drusen-like deposits was dependent on activation of mTORC1 in cones. Both major types of advanced AMD pathologies, including geographic atrophy (GA) and neovascular pathologies, were also seen. Finally, activated mTORC1 in PRs resulted in a threefold reduction in di-docosahexaenoic acid (DHA)-containing phospholipid species. Feeding mice a DHA-enriched diet alleviated most pathologies. The data recapitulate many aspects of the human disease, suggesting that metabolic adaptations in photoreceptors could contribute to disease progression in AMD. Identifying the changes downstream of mTORC1 that lead to advanced pathologies in mouse might present new opportunities to study the role of PRs in AMD pathogenesis.
Assuntos
Envelhecimento/patologia , Macula Lutea/patologia , Degeneração Macular/patologia , Células Fotorreceptoras Retinianas Cones/patologia , Idoso , Idoso de 80 Anos ou mais , Animais , Lâmina Basilar da Corioide/metabolismo , Modelos Animais de Doenças , Feminino , Humanos , Lipofuscina/metabolismo , Lipoproteínas/metabolismo , Macula Lutea/citologia , Masculino , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Camundongos , Células Fotorreceptoras Retinianas Cones/metabolismo , Epitélio Pigmentado da Retina/metabolismoRESUMO
OBJECTIVE: Skin brightness and spot have a significant impact on youthful and beautiful appearance. One important factor influencing skin brightness is the amount of internal reflected light from the skin. Observers recognize the total surface-reflected light and internal reflected light as skin brightness. The more internal reflected light from the skin, the more attractive and brighter the skin appears. This study aims to identify a new natural cosmetic ingredient that increases the skin's internal reflected light, decreases spot and provides a youthful and beautiful skin appearance. METHODS: Lipofuscin in epidermal keratinocytes, the aggregating complex of denatured proteins and peroxidized lipids, is one factor that decreases skin brightness and causes of spot. Aggregates block light transmission, and peroxidized lipids lead to skin yellowness, dullness and age spot. Lipofuscin is known to accumulate intracellularly with ageing. Rapid removal of intracellular denatured proteins prevents lipofuscin formation and accumulation in cells. We focused a proteasome system that efficiently removes intracellular denatured proteins. To identify natural ingredients that increase proteasome activity, we screened 380 extracts derived from natural products. The extract with the desired activity was fractionated and purified to identify active compounds that lead to proteasome activation. Finally, the efficacy of the proteasome-activating extract was evaluated in a human clinical study. RESULTS: We discovered that Juniperus communis fruits (Juniper berry) extract (JBE) increases proteasome activity and suppresses lipofuscin accumulation in human epidermal keratinocytes. We found Anthricin and Yatein, which belong to the lignan family, to be major active compounds responsible for the proteasome-activating effect of JBE. In a human clinical study, an emulsion containing 1% JBE was applied to half of the face twice daily for 4 weeks, resulting in increased internal reflected light, brightness improvement (L-value) and reduction in yellowness (b-value) and spot in the cheek area. CONCLUSION: This is the first report demonstrating that JBE containing Anthricin and Yatein decreases lipofuscin accumulation in human epidermal keratinocytes through proteasome activation, increases brightness and decreases surface spots in human skin. JBE would be an ideal natural cosmetic ingredient for creating a more youthful and beautiful skin appearance with greater brightness and less spot.
OBJECTIF: La luminosité et les taches de peau ont un impact significatif sur la jeunesse et la beauté de l'apparence. L'un des facteurs importants influençant la luminosité de la peau est la quantité de lumière interne réfléchie par la peau. Pour les observateurs, la luminosité de la peau correspond à la somme de la lumière réfléchie par la surface et de la lumière réfléchie par l'intérieur de la peau. Plus la quantité de lumière interne réfléchie par la peau est importante, plus la peau semble attrayante et lumineuse. Cette étude vise à identifier un nouvel ingrédient cosmétique naturel qui augmente la lumière interne réfléchie par la peau, diminue les taches et donne à la peau une apparence jeune et belle. MÉTHODES: La lipofuscine dans les kératinocytes de l'épiderme, le complexe agrégé de protéines dénaturées et de lipides peroxydés, est un facteur qui diminue l'éclat de la peau et qui est à l'origine des taches. Les agrégats bloquent la transmission de la lumière et les lipides peroxydés entraînent une coloration jaune de la peau, un aspect terne et des taches de vieillesse. On sait que la lipofuscine s'accumule au niveau intracellulaire avec le vieillissement. L'élimination rapide des protéines dénaturées intracellulaires empêche la formation et l'accumulation de lipofuscine dans les cellules. Nous avons mis l'accent sur un système de protéasome qui élimine efficacement les protéines dénaturées intracellulaires. Pour identifier les ingrédients naturels qui augmentent l'activité du protéasome, nous avons passé au crible 380 extraits dérivés de produits naturels. L'extrait présentant l'activité souhaitée a été fractionné et purifié afin d'identifier les composés actifs qui conduisent à l'activation du protéasome. Enfin, l'efficacité de l'extrait activant le protéasome a été évaluée dans une étude clinique humaine. RÉSULTATS: Nous avons découvert que l'extrait de Juniperus communis fruits (baie de genièvre) augmente l'activité du protéasome et supprime l'accumulation de lipofuscine dans les kératinocytes épidermiques humains. Nous avons découvert que l'anthricine et la yateine, qui appartiennent à la famille des lignanes, sont les principaux composés actifs responsables de l'effet activateur du protéasome de l'extrait de baies de genévrier. Dans une étude clinique humaine, une émulsion contenant 1 % de JBE a été appliquée sur la moitié du visage deux fois par jour pendant 4 semaines, ce qui a entraîné une augmentation de la lumière interne réfléchie, une amélioration de la luminosité (valeur L) et une réduction de la jaunisse (valeur b) et des taches dans la zone des joues. CONCLUSION: Il s'agit du premier rapport démontrant que l'EBJ contenant de l'anthricine et de la yateine diminue l'accumulation de lipofuscine dans les kératinocytes épidermiques humains par l'activation du protéasome, augmente la luminosité et diminue les taches superficielles de la peau humaine. Le JBE serait un ingrédient cosmétique naturel idéal pour créer une peau plus jeune et plus belle, plus lumineuse et moins tachetée.
Assuntos
Juniperus , Complexo de Endopeptidases do Proteassoma , Humanos , Lipofuscina/metabolismo , Juniperus/metabolismo , Frutas/metabolismo , Queratinócitos/metabolismo , ProteínasRESUMO
Aging is a risk factor for multiple retinal degeneration diseases. Entraining brain gamma oscillations with gamma-flicker light (γFL) has been confirmed to coordinate pathological changes in several Alzheimer's disease mouse models and aged mice. However, the direct effect of γFL on retinal aging remains unknown. We assessed retinal senescence-associated beta-galactosidase (ß-gal) and autofluorescence in 20-month-old mice and found reduced ß-gal-positive cells in the inner retina and diminished lipofuscin accumulation around retinal vessels after 6 days of γFL. In immunofluorescence, γFL was further demonstrated to ameliorate aging-related retinal changes, including a decline in microtubule-associated protein 1 light chain 3 beta expression, an increase in complement C3 activity, and an imbalance between the anti-oxidant factor catalase and pro-oxidant factor carboxymethyl lysine. Moreover, we found that γFL can increase the expression of activating transcription factor 4 (ATF4) in the inner retina, while revealing a decrease of ATF4 expression in the inner retina and positive expression in the outer segment of photoreceptor and RPE layer for aged mice. Western blotting was then used to confirm the immunofluorescence results. After mRNA sequencing (NCBI Sequence Read Archive database: PRJNA748184), we found several main mechanistic clues, including mitochondrial function and chaperone-mediated protein folding. Furthermore, we extended γFL to aged Apoe-/- mice and showed that 1-m γFL treatment even improved the structures of retinal-pigment-epithelium basal infolding and Bruch's membrane. Overall, γFL can orchestrate various pathological characteristics of retinal aging in mice and might be a noninvasive, convenient, and tissue-specific therapeutic strategy for retinal aging.
Assuntos
Complemento C3 , Lipofuscina , Fator 4 Ativador da Transcrição/metabolismo , Animais , Antioxidantes/metabolismo , Apolipoproteínas E/metabolismo , Catalase/metabolismo , Complemento C3/metabolismo , Lipofuscina/metabolismo , Lisina/metabolismo , Camundongos , Proteínas Associadas aos Microtúbulos/metabolismo , RNA Mensageiro/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Retina/metabolismo , beta-Galactosidase/metabolismoRESUMO
PURPOSE: Agaro-oligosaccharides (AGO), hydrolysis products of agarose, is known to have antioxidant and anti-inflammatory properties. Speculating that AGO is effective for preventing aging, we investigated the longevity-supporting effects of AGO and their mechanisms using Caenorhabditis elegans. METHODS: Caenorhabditis elegans were fed AGO from young adulthood. The lifespan, locomotory activity, lipofuscin accumulation, and heat stress resistance of the worms were examined. To elucidate mechanisms of AGO-mediated longevity, we conducted comprehensive expression analysis using microarrays. Moreover, we used quantitative real-time PCR (qRT-PCR) to verify the genes showing differential expression levels. Furthermore, we measured the lifespan of loss-of-function mutants to determine the genes related to AGO-mediated longevity. RESULTS: AGO extended the lifespan of C. elegans, reduced lipofuscin accumulation, and maintained vigorous locomotion. The microarray analysis revealed that the endoplasmic reticulum-unfolded protein response (ER-UPR) and insulin/insulin-like growth factor-1-mediated signaling (IIS) pathway were activated in AGO-fed worms. The qRT-PCR analysis showed that AGO treatment suppressed sir-2.1 expression, which is a negative regulator of ER-UPR. In loss-of-function mutant of sir-2.1, AGO-induced longevity and heat stress resistance were decreased or cancelled completely. Furthermore, the pro-longevity effect of AGO was decreased in loss-of-function mutants of abnormal Dauer formation (daf) -2 and daf-16, which are IIS pathway-related genes. CONCLUSION: AGO delays the C. elegans aging process and extends their lifespan through the activations of ER-UPR and the IIS pathway.
Assuntos
Proteínas de Caenorhabditis elegans , Insulinas , Sirtuínas , Animais , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Longevidade/fisiologia , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Fator de Crescimento Insulin-Like I/metabolismo , Ágar/metabolismo , Ágar/farmacologia , Antioxidantes/farmacologia , Sefarose/metabolismo , Sefarose/farmacologia , Lipofuscina/metabolismo , Lipofuscina/farmacologia , Resposta a Proteínas não Dobradas , Oligossacarídeos/farmacologia , Oligossacarídeos/metabolismo , Insulinas/genética , Insulinas/metabolismo , Insulinas/farmacologia , Fatores de Transcrição Forkhead/genética , Sirtuínas/genética , Sirtuínas/metabolismoRESUMO
ß-Amyloid toxicity (Aß) is an important pathological factor of Alzheimer's disease (AD). Studies have shown that genistein can reduce the toxicity of Aß to a certain extent; however, the specific mechanism is still uncertain. In the study, we applied Caenorhabditis elegans strains expressing Aß peptides to evaluate the role of genistein inhibiting Aß toxicity and the undying mechanism. Genistein influencing the sterol metabolism pathway, the HSP-16.2 pathway, and lipofuscin in different strains of C. elegans were studied using reverse transcription-polymerase chain reaction, fluorescence labeling, RNA interference (RNAi), and so forth. Our results showed that genistein alleviated the paralysis of transgenic C. elegans strains. Furthermore, in AD C. elegans, genistein reduced the fluorescence of lipofuscin, downregulated the messenger RNA (mRNA) level of vit-3 and vit-6 which were related to the sterol metabolism pathway, significantly increased the mRNA level and protein level of HSP-16.2, increased the nuclear translocation of the DAF-16 transcription factor and increased the survival rate after heat stress, which was closely associated with HSP-16.2 levels. However, the paralysis-alleviating effect of genistein was greatly reduced because of RNAi-mediated inhibition of hsp-16.2, indicating that the anti-Aß toxicity effect of genistein was greatly dependent on HSP-16.2. The above results suggest that genistein inhibiting the toxicity of Aß in C. elegans, is involved in the modulation of the sterol metabolism pathway by promoting transcription factor DAF-16 translocation into the nucleus, increasing the expression level of HSP-16.2, and reducing the levels of lipofuscin through its antioxidant activity.
Assuntos
Doença de Alzheimer , Proteínas de Caenorhabditis elegans , Animais , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/metabolismo , Fatores de Transcrição Forkhead/farmacologia , Genisteína/farmacologia , Lipofuscina/metabolismo , Lipofuscina/farmacologia , Paralisia , RNA Mensageiro/metabolismo , Transdução de Sinais , Esteróis/farmacologia , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
INTRODUCTION: The accumulation of lipofuscin is a hallmark in the pathogenesis of Stargardt disease type 1 (STGD1) and geographic atrophy (GA) secondary to age-related macular degeneration. Limiting lipofuscin accumulation by inhibiting the retinol-binding protein 4 (RBP4) is being explored as a potential treatment target for those diseases. In this study, we aimed to establish the concentration of RBP4 in the systemic circulation in different age cohorts of healthy individuals and to check if patients with STGD1 or GA may show abnormal RBP4 levels. METHODS: Forty healthy subjects of various age-groups, 15 Stargardt patients, and 15 GA patients were included in the study. We measured RBP4 levels, serum retinol (SR) levels, complete blood count, and blood chemistry including liver function tests. RESULTS: Mean RBP4 for all cohorts was 26,911.40 ± 6,198.61 ng/mL, and mean SR 1.75 ± 0.36 µmol/L. Age was not found to significantly impact levels neither of RBP4 and SR nor of the RBP4-to-SR ratio. Also, the 2 patient groups showed similar blood levels to their age-matched controls. CONCLUSION: Serum RBP4 and SR do not appear to be affected by age in healthy individuals and remain within normal limits in both STGD1 and GA.
Assuntos
Atrofia Geográfica , Proteínas Plasmáticas de Ligação ao Retinol , Doença de Stargardt , Vitamina A , Atrofia Geográfica/sangue , Voluntários Saudáveis , Humanos , Lipofuscina/metabolismo , Proteínas Plasmáticas de Ligação ao Retinol/análise , Doença de Stargardt/sangue , Vitamina A/sangueRESUMO
Autosomal recessive retinal degenerative diseases cause visual impairment and blindness in both humans and dogs. Currently, no standard treatment is available, but pioneering gene therapy-based canine models have been instrumental for clinical trials in humans. To study a novel form of retinal degeneration in Labrador retriever dogs with clinical signs indicating cone and rod degeneration, we used whole-genome sequencing of an affected sib-pair and their unaffected parents. A frameshift insertion in the ATP binding cassette subfamily A member 4 (ABCA4) gene (c.4176insC), leading to a premature stop codon in exon 28 (p.F1393Lfs*1395), was identified. In contrast to unaffected dogs, no full-length ABCA4 protein was detected in the retina of an affected dog. The ABCA4 gene encodes a membrane transporter protein localized in the outer segments of rod and cone photoreceptors. In humans, the ABCA4 gene is associated with Stargardt disease (STGD), an autosomal recessive retinal degeneration leading to central visual impairment. A hallmark of STGD is the accumulation of lipofuscin deposits in the retinal pigment epithelium (RPE). The discovery of a canine homozygous ABCA4 loss-of-function mutation may advance the development of dog as a large animal model for human STGD.
Assuntos
Membro 4 da Subfamília A de Transportadores de Cassetes de Ligação de ATP/genética , Doenças do Cão/genética , Degeneração Macular/congênito , Mutação , Membro 4 da Subfamília A de Transportadores de Cassetes de Ligação de ATP/química , Membro 4 da Subfamília A de Transportadores de Cassetes de Ligação de ATP/metabolismo , Transportadores de Cassetes de Ligação de ATP/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Códon sem Sentido , Modelos Animais de Doenças , Doenças do Cão/metabolismo , Doenças do Cão/patologia , Cães , Feminino , Genes Recessivos , Homozigoto , Humanos , Lipofuscina/metabolismo , Degeneração Macular/genética , Degeneração Macular/metabolismo , Degeneração Macular/veterinária , Masculino , Microscopia de Fluorescência , Modelos Moleculares , Mutagênese Insercional , Linhagem , Conformação Proteica , Retina/metabolismo , Retina/patologia , Doença de Stargardt , Sequenciamento Completo do GenomaRESUMO
Lipofuscin granules from retinal pigment epithelium (RPE) cells contain bisretinoid fluorophores, which are photosensitizers and are phototoxic to cells. In the presence of oxygen, bisretinoids are oxidized to form various products, containing aldehydes and ketones, which are also potentially cytotoxic. In a prior study, we identified that bisretinoid oxidation and degradation products have both hydrophilic and amphiphilic properties, allowing their diffusion through the lipofuscin granule membrane into the RPE cell cytoplasm, and are thiobarbituric acid (TBA)-active. The purpose of the present study was to determine if these products exhibit a toxic effect to the RPE cell also in the absence of light. The experiments were performed using the lipofuscin-fed ARPE-19 cell culture. The RPE cell viability analysis was performed with the use of flow cytofluorimetry and laser scanning confocal microscopy. The results obtained indicated that the cell viability of the lipofuscin-fed ARPE-19 sample was clearly reduced not immediately after visible light irradiation for 18 h, but after 4 days maintaining in the dark. Consequently, we could conclude that bisretinoid oxidation products have a damaging effect on the RPE cell in the dark and can be considered as an aggravating factor in age-related macular degeneration progression.
Assuntos
Lipofuscina , Fármacos Fotossensibilizantes , Lipofuscina/metabolismo , Fármacos Fotossensibilizantes/farmacologia , Fármacos Fotossensibilizantes/metabolismo , Retinoides/metabolismo , Epitélio Pigmentado da Retina/metabolismo , Aldeídos/metabolismo , Oxigênio/metabolismo , Cetonas/metabolismoRESUMO
Neurodegenerative diseases are debilitating impairments that affect millions of people worldwide and are characterized by progressive degeneration of structure and function of the central or peripheral nervous system. Effective biomarkers for neurodegenerative diseases can be used to improve the diagnostic workup in the clinic as well as facilitate the development of effective disease-modifying therapies. Progranulin (PGRN) has been reported to be involved in various neurodegenerative disorders. Hence, in the current study we systematically compared the inflammation and accumulation of typical neurodegenerative disease markers in the brain tissue between PGRN knockout (PGRN KO) and wildtype (WT) mice. We found that PGRN deficiency led to significant neuron loss as well as activation of microglia and astrocytes in aged mice. Several characteristic neurodegenerative markers, including α-synuclein, TAR DNA-binding protein 43 (TDP-43), Tau, and ß-amyloid, were all accumulated in the brain of PGRN-deficient mice as compared to WT mice. Moreover, higher aggregation of lipofuscin was observed in the brain tissue of PGRN-deficient mice compared with WT mice. In addition, the autophagy was also defective in the brain of PGRN-deficient mice, indicated by the abnormal expression level of autophagy marker LC3-II. Collectively, comprehensive assays support the idea that PGRN plays an important role during the development of neurodegenerative disease, indicating that PGRN might be a useful biomarker for neurodegenerative diseases in clinical settings.
Assuntos
Envelhecimento/patologia , Biomarcadores/metabolismo , Doenças Neurodegenerativas/metabolismo , Progranulinas/deficiência , Peptídeos beta-Amiloides/metabolismo , Animais , Astrócitos/metabolismo , Astrócitos/patologia , Autofagia , Encéfalo/patologia , Encéfalo/ultraestrutura , Proteínas de Ligação a DNA/metabolismo , Lipofuscina/metabolismo , Camundongos Knockout , Microglia/metabolismo , Microglia/patologia , Neurônios/metabolismo , Neurônios/patologia , Fosforilação , Progranulinas/metabolismo , Agregados Proteicos , alfa-Sinucleína/metabolismo , Proteínas tau/metabolismoRESUMO
Near-infrared autofluorescence (NIA) is a non-invasive retinal imaging technique for examination of the retinal pigment epithelium (RPE) based on the autofluorescence of melanin. Melanin has several functions within the RPE cells, in one of them it serves as a protective antioxidative factor within the RPE cells and is involved in the phagocytosis of photoreceptor outer segments. Disorders that affect the photoreceptor-RPE complex result in alterations of RPE cells which are detectable by alterations of NIA. Therefore, NIA allows to detect early alterations in inherited and acquired chorioretinal disorders, frequently prior to ophthalmoscopical visualisation and often prior to alterations in lipofuscin associated fundus autofluorescence (FAF) or optical coherence tomography (OCT). Although NIA and FAF relate to disorders affecting the RPE, findings between both imaging methods differ and the area involved has been demonstrated to be larger in NIA compared to FAF in several disorders (e.g., age-related macular degeneration, retinitis pigmentosa, ABCA4-gene associated Stargardt disease and cone-rod dystrophy, light damage), indicating that NIA detects earlier alterations compared to FAF. In addition, due to the absence of blue-light filtering which limits foveal visualisation in FAF, foveal alterations can be much better detected using NIA. A reduced subfoveal NIA intensity is the earliest sign of autosomal dominant BEST1-associated disease, when FAF and OCT are still normal. In other disorders, a normal subfoveal NIA intensity is associated with good visual acuity. This review summarizes the present knowledge on NIA and demonstrates biomarkers for various chorioretinal disorders.
Assuntos
Melaninas , Epitélio Pigmentado da Retina , Transportadores de Cassetes de Ligação de ATP , Bestrofinas , Angiofluoresceinografia/métodos , Fundo de Olho , Humanos , Lipofuscina/metabolismo , Melaninas/metabolismo , Epitélio Pigmentado da Retina/diagnóstico por imagem , Tomografia de Coerência Óptica/métodosRESUMO
Background and Objectives: Age-related macular degeneration is a slow-progressing disease in which lipofuscin accumulates in the retina, causing inflammation and apoptosis of retinal pigment epithelial (RPE) cells. This study aimed to identify N-methyl-D-aspartate (NMDA) signaling as a novel mechanism for scavenging N-retinylidene-N-retinylethanolamine (A2E), a component of ocular lipofuscin, in human RPE cells. Materials and Methods: A2E degradation assays were performed in ARPE-19 cells using fluorescently labeled A2E. The autophagic activity in ARPE-19 cells was measured upon blue light (BL) exposure, after A2E treatment. Autophagy flux was determined by measuring LC3-II formation using immunoblotting and confocal microscopy. To determine whether autophagy via the NMDA receptor is involved in A2E clearance, ATG5-deficient cells were used. Results: Ro 25-6981, an NR2B-selective NMDA receptor antagonist, effectively cleared A2E. Ro 25-6981 reduced A2E accumulation in the lysosomes of ARPE-19 cells at sub-cytotoxic concentrations, while increasing the formation of LC3-II and decreasing p62 protein levels in a concentration-dependent manner. The autophagic flux monitored by RFP-GFP-LC3 and bafilomycin A1 assays was significantly increased by Ro 25-6981. A2E clearance by Ro 25-6981 was abolished in ATG5-depleted ARPE-19 cells, suggesting that A2E degradation by Ro 25-6981 was mediated by autophagy. Furthermore, treatment with other NMDA receptor antagonists, CP-101,606 and AZD6765, showed similar effects on autophagy activation and A2E degradation in ARPE-19 cells. In contrast, glutamate, an NMDA receptor agonist, exhibited a contrasting effect, suggesting that both the activation of autophagy and the degradation of A2E by Ro 25-6981 in ARPE-19 cells occur through inhibition of the NMDA receptor pathway. Conclusions: This study demonstrates that NMDA receptor antagonists degrade lipofuscin via autophagy in human RPE cells and suggests that NMDA receptor antagonists could be promising new therapeutics for retinal degenerative diseases.