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1.
Chem Biol Interact ; 395: 111036, 2024 May 25.
Artigo em Inglês | MEDLINE | ID: mdl-38705443

RESUMO

Gelsemium elegans Benth. (G. elegans) is a traditional medicinal herb that has anti-inflammatory, analgesic, sedative, and detumescence effects. However, it can also cause intestinal side effects such as abdominal pain and diarrhea. The toxicological mechanisms of gelsenicine are still unclear. The objective of this study was to assess enterotoxicity induced by gelsenicine in the nematodes Caenorhabditis elegans (C. elegans). The nematodes were treated with gelsenicine, and subsequently their growth, development, and locomotion behavior were evaluated. The targets of gelsenicine were predicted using PharmMapper. mRNA-seq was performed to verify the predicted targets. Intestinal permeability, ROS generation, and lipofuscin accumulation were measured. Additionally, the fluorescence intensities of GFP-labeled proteins involved in oxidative stress and unfolded protein response in endoplasmic reticulum (UPRER) were quantified. As a result, the treatment of gelsenicine resulted in the inhibition of nematode lifespan, as well as reductions in body length, width, and locomotion behavior. A total of 221 targets were predicted by PharmMapper, and 731 differentially expressed genes were screened out by mRNA-seq. GO and KEGG enrichment analysis revealed involvement in redox process and transmembrane transport. The permeability assay showed leakage of blue dye from the intestinal lumen into the body cavity. Abnormal mRNAs expression of gem-4, hmp-1, fil-2, and pho-1, which regulated intestinal development, absorption and catabolism, transmembrane transport, and apical junctions, was observed. Intestinal lipofuscin and ROS were increased, while sod-2 and isp-1 expressions were decreased. Multiple proteins in SKN-1/DAF-16 pathway were found to bind stably with gelsenicine in a predictive model. There was an up-regulation in the expression of SKN-1:GFP, while the nuclear translocation of DAF-16:GFP exhibited abnormality. The UPRER biomarker HSP-4:GFP was down-regulated. In conclusion, the treatment of gelsenicine resulted in the increase of nematode intestinal permeability. The toxicological mechanisms underlying this effect involved the disruption of intestinal barrier integrity, an imbalance between oxidative and antioxidant processes mediated by the SKN-1/DAF-16 pathway, and abnormal unfolded protein reaction.


Assuntos
Caenorhabditis elegans , Espécies Reativas de Oxigênio , Animais , Caenorhabditis elegans/efeitos dos fármacos , Caenorhabditis elegans/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Quinoxalinas/farmacologia , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Estresse Oxidativo/efeitos dos fármacos , Intestinos/efeitos dos fármacos , Mucosa Intestinal/metabolismo , Mucosa Intestinal/efeitos dos fármacos , Gelsemium/química , Resposta a Proteínas não Dobradas/efeitos dos fármacos , Permeabilidade/efeitos dos fármacos , Lipofuscina/metabolismo , Locomoção/efeitos dos fármacos , Alcaloides Indólicos
2.
Free Radic Biol Med ; 221: 273-282, 2024 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-38740102

RESUMO

Defective mitochondria and autophagy, as well as accumulation of lipid and iron in WDR45 mutant fibroblasts, is related to beta-propeller protein-associated neurodegeneration (BPAN). In this study, we found that enlarged lysosomes in cells derived from patients with BPAN had low enzyme activity, and most of the enlarged lysosomes had an accumulation of iron and oxidized lipid. Cryo-electron tomography revealed elongated lipid accumulation, and spectrometry-based elemental analysis showed that lysosomal iron and oxygen accumulation superimposed with lipid aggregates. Lysosomal lipid aggregates superimposed with autofluorescence as free radical generator, lipofuscin. To eliminate free radical stress by iron accumulation in cells derived from patients with BPAN, we investigated the effects of the iron chelator, 2,2'-bipyridine (bipyridyl, BIP). To study whether the defects in patient-derived cells can be rescued by an iron chelator BIP, we tested whether the level of iron and reactive oxygen species (ROS) in the cells and genes related to oxidative stress were rescued BIP treatment. Although BIP treatment decreased some iron accumulation in the cytoplasm and mitochondria, the accumulation of iron in the lysosomes and levels of cellular ROS were unaffected. In addition, the change of specific RNA levels related to free radical stress in patient fibroblasts was not rescued by BIP. To alleviate free radical stress, we investigated whether l-serine can regulate abnormal structures in cells derived from patients with BPAN through the regulation of free radical stress. l-serine treatment alleviated increase of enlarged lysosomes and iron accumulation and rescued impaired lysosomal activity by reducing oxidized lipid accumulation in the lysosomes of the cells. Lamellated lipids in the lysosomes of the cells were identified as lipofuscin through correlative light and electron microscopy, and l-serine treatment reduced the increase of lipofuscin. These data suggest that l-serine reduces oxidative stress-mediated lysosomal lipid oxidation and iron accumulation by rescuing lysosomal activity.


Assuntos
Fibroblastos , Ferro , Lipofuscina , Lisossomos , Estresse Oxidativo , Espécies Reativas de Oxigênio , Serina , Humanos , Lisossomos/metabolismo , Lisossomos/efeitos dos fármacos , Lipofuscina/metabolismo , Ferro/metabolismo , Fibroblastos/metabolismo , Fibroblastos/efeitos dos fármacos , Fibroblastos/patologia , Estresse Oxidativo/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Serina/metabolismo , Mitocôndrias/metabolismo , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/patologia , Distrofias Neuroaxonais/metabolismo , Distrofias Neuroaxonais/patologia , Distrofias Neuroaxonais/tratamento farmacológico , Distrofias Neuroaxonais/genética , 2,2'-Dipiridil/farmacologia , 2,2'-Dipiridil/análogos & derivados , Quelantes de Ferro/farmacologia
3.
Vet Clin Pathol ; 53(2): 168-178, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38570895

RESUMO

BACKGROUND: Lipofuscin-like cytoplasmic inclusions have been reported in human blood neutrophils and monocytes but have not been described in dogs. In people, these "green granules of death" have been associated with moderate to severe hepatocellular injury and high mortality. OBJECTIVES: To describe clinicopathologic abnormalities, diagnoses, and outcomes of dogs with greenish inclusions in blood neutrophils or monocytes, and to determine if the inclusions have features of lipofuscin. METHODS: Clinical cases were identified prospectively through routine evaluation of CBC samples. Leukocyte inclusions were characterized with routine staining and assessed for iron and autofluorescence. Additional cases were identified by examination of archived blood smears from dogs meeting search criteria for hepatocellular injury, and clinicopathologic findings were recorded. RESULTS: All 7 prospectively identified dogs with inclusions had inflammation and moderate to marked increases in serum alanine aminotransferase (ALT) activity, as did the 4 dogs identified from the 97 meeting retrospective search criteria. The inclusions were Prussian blue-negative (5/5) with broad-spectrum autofluorescence (5/5) and the appearance of lipofuscin with and without Wright staining. Most clinical diagnoses involved hepatic disorders (5/7 prospective and 3/4 retrospective cases) or pancreatitis (3/7 prospective and 2/4 retrospective cases), and some involved both; 8 of 11 dogs died within 7 days of admission. CONCLUSIONS: Blue-green cytoplasmic inclusions uncommonly found in blood neutrophils ± monocytes of routine canine blood smears have stained and unstained properties of lipofuscin and suggest the presence of hepatocellular injury, often severe. Reporting these inclusions is recommended to guide clinical management.


Assuntos
Doenças do Cão , Corpos de Inclusão , Cães , Animais , Doenças do Cão/patologia , Doenças do Cão/sangue , Doenças do Cão/diagnóstico , Masculino , Corpos de Inclusão/patologia , Feminino , Estudos Retrospectivos , Hepatopatias/veterinária , Hepatopatias/patologia , Hepatopatias/sangue , Hepatopatias/diagnóstico , Lipofuscina/metabolismo , Estudos Prospectivos , Neutrófilos/patologia , Leucócitos/patologia , Alanina Transaminase/sangue , Monócitos/patologia , Pancreatite/veterinária , Pancreatite/patologia , Pancreatite/sangue , Pancreatite/diagnóstico
4.
Int J Mol Sci ; 25(7)2024 Mar 23.
Artigo em Inglês | MEDLINE | ID: mdl-38612421

RESUMO

The retinal pigment epithelium (RPE), which ensures the normal functioning of the neural retina, is a pigmented single-cell layer that separates the retina from the Bruch's membrane and the choroid. There are three main types of pigment granules in the RPE cells of the human eye: lipofuscin granules (LG) containing the fluorescent "age pigment" lipofuscin, melanoprotein granules (melanosomes, melanolysosomes) containing the screening pigment melanin and complex melanolipofuscin granules (MLG) containing both types of pigments simultaneously-melanin and lipofuscin. This review examines the functional role of pigment granules in the aging process and in the development of oxidative stress and associated pathologies in RPE cells. The focus is on the process of light-induced oxidative degradation of pigment granules caused by reactive oxygen species. The reasons leading to increased oxidative stress in RPE cells as a result of the oxidative degradation of pigment granules are considered. A mechanism is proposed to explain the phenomenon of age-related decline in melanin content in RPE cells. The essence of the mechanism is that when the lipofuscin part of the melanolipofuscin granule is exposed to light, reactive oxygen species are formed, which destroy the melanin part. As more melanolipofuscin granules are formed with age and the development of degenerative diseases, the melanin in pigmented epithelial cells ultimately disappears.


Assuntos
Melaninas , Epitélio Pigmentado da Retina , Humanos , Lipofuscina , Espécies Reativas de Oxigênio , Retina
5.
Invest Ophthalmol Vis Sci ; 65(4): 32, 2024 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-38648041

RESUMO

Purpose: To undertake the first ultrastructural characterization of human retinal pigment epithelial (RPE) differentiation from fetal development to adolescence. Methods: Ten fetal eyes and three eyes aged six, nine, and 17 years were examined in the temporal retina adjacent to the optic nerve head by transmission electron microscopy. The area, number, and distribution of RPE organelles were quantified and interpreted within the context of adjacent photoreceptors, Bruch's membrane, and choriocapillaris maturation. Results: Between eight to 12 weeks' gestation (WG), pseudostratified columnar epithelia with apical tight junctions differentiate to a simple cuboidal epithelium with random distribution of melanosomes and mitochondria. Between 12 to 26 WG, cells enlarge and show long apical microvilli and apicolateral junctional complexes. Coinciding with eye opening at 26 WG, melanosomes migrate apically whereas mitochondria distribute to perinuclear regions, with the first appearance of phagosomes, complex granules, and basolateral extracellular space (BES) formation. Significantly, autophagy and heterophagy, as evidenced by organelle recycling, and the gold standard of ultrastructural evidence for autophagy of double-membrane autophagosomes and mitophagosomes were evident from 32 WG, followed by basal infoldings of RPE cell membrane at 36 WG. Lipofuscin formation and deposition into the BES evident at six years increased at 17 years. Conclusions: We provide compelling ultrastructural evidence that heterophagy and autophagy begins in the third trimester of human fetal development and that deposition of cellular byproducts into the extracellular space of RPE takes place via exocytosis. Transplanted RPE cells must also demonstrate the capacity to subserve autophagic and heterophagic functions for effective disease mitigation.


Assuntos
Autofagia , Exocitose , Lipofuscina , Microscopia Eletrônica de Transmissão , Epitélio Pigmentado da Retina , Humanos , Epitélio Pigmentado da Retina/metabolismo , Epitélio Pigmentado da Retina/ultraestrutura , Epitélio Pigmentado da Retina/embriologia , Adolescente , Autofagia/fisiologia , Criança , Lipofuscina/metabolismo , Exocitose/fisiologia , Espaço Extracelular/metabolismo , Idade Gestacional , Feminino , Masculino , Desenvolvimento Fetal/fisiologia , Mitocôndrias/metabolismo , Mitocôndrias/ultraestrutura , Diferenciação Celular/fisiologia
6.
Exp Eye Res ; 242: 109889, 2024 May.
Artigo em Inglês | MEDLINE | ID: mdl-38593971

RESUMO

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/patologia
7.
Exp Brain Res ; 242(4): 971-986, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38430248

RESUMO

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 , Autofagia
8.
Food Funct ; 15(7): 3353-3364, 2024 Apr 02.
Artigo em Inglês | MEDLINE | ID: mdl-38481358

RESUMO

Cyclocodon lancifolius fruit is a promising commercial fruit with antioxidant activity and is rich in polyphenolic compounds. In this study, the anti-aging activity of C. lancifolius fruit extract (CF) on Caenorhabditis elegans (C. elegans) was evaluated by observing the longevity, stress response, reproduction, oscillation, lipofuscin, and antioxidant enzymes of worms. Moreover, the effects and potential mechanisms of CF on delaying C. elegans senescence at the mRNA and metabolite levels were investigated. The results showed that CF treatment significantly increased the lifespan and stress resistance, decreased the levels of lipofuscin and reactive oxygen species (ROS), and improved the antioxidant system of C. elegans. The extension of the lifespan of C. elegans was remarkably correlated with the upregulation of mtl-1 and Hsp-16.2, along with the downregulation of age-1, daf-2, and akt-1. Metabolomics analysis revealed that purine metabolism is a key regulatory pathway for CF to exert anti-aging effects. The present study suggests that C. lancifolius fruit has potential for use as a functional food to enhance antioxidant capacity and delay aging.


Assuntos
Proteínas de Caenorhabditis elegans , Caenorhabditis elegans , Animais , Caenorhabditis elegans/metabolismo , Longevidade , Antioxidantes/farmacologia , Antioxidantes/metabolismo , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Frutas/metabolismo , Lipofuscina/metabolismo , Estresse Oxidativo , Espécies Reativas de Oxigênio/metabolismo , Purinas/metabolismo , Fatores de Transcrição Forkhead/metabolismo
9.
Genes (Basel) ; 15(2)2024 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-38397235

RESUMO

A progressive neurological disorder was observed in a male neutered Weimaraner. Clinical signs included fecal incontinence, lethargy, moderate paraparesis, proprioceptive pelvic limb ataxia, falling, cognitive decline, incoordination, decreased interest in food, changes in posture, and episodes of trance-like behavior. Neurologic signs were first observed at approximately 4 years, 10 months of age and progressed slowly. Magnetic resonance imaging showed generalized brain atrophy with areas of white matter pathology. Humane euthanasia was elected at 6 years, 7 months of age due to increasing severity of the neurological signs. Autofluorescent intracellular granules were observed in the cerebral and cerebellar cortexes, optic nerve, and cardiac muscle of the affected dog. These abnormal inclusions in the cerebral cortex and cardiac muscle immunolabeled with antibodies to mitochondrial ATP synthase subunit c protein, like that observed in the neuronal ceroid lipofuscinosis group of lysosomal storage diseases. Immunolabeling also demonstrated pronounced neuroinflammation in brain tissues. The ultrastructural appearances of the disease-related inclusion bodies in the brain and optic nerve were quite variable. The ultrastructure and locations of many of the inclusions in the nervous tissues suggested that they were derived, at least in part, from the myelin surrounding axons. The storage bodies in the cardiac muscle were located in mitochondria-rich regions and consisted of parallel arrays of membrane-like components interspersed with electron-dense flocculent material. The disease was characterized by pronounced abnormalities in the myelin of the brain and optic nerve consisting of distinctive areas of ballooning between the layers of myelin. The whole genome sequence generated from the affected dog contained a homozygous G-to-A missense mutation in CNP, which encodes proteins with CNPase enzyme activity and a structural role in myelin. The mutation predicts a Thr42Met amino acid sequence substitution. Genotyping of archived Weimaraner DNA samples identified an additional G > A variant homozygote with a clinical history and brain lesions similar to those of the proband. Of 304 Weimaraners and over 4000 other dogs of various breeds, the proband and the other Weimaraner that exhibited similar signs were the only two that were homozygous for the CNP missense variant. CNPase immunolabeling was widespread in brain tissues from normal dogs but was undetectable in the same tissues from the proband. Based on the clinical history, fluorescence and electron-microscopy, immunohistochemistry, and molecular genetic findings, the late-onset Weimaraner disorder likely results from the missense mutation that results in CNPase deficiency, leading to myelin abnormalities, accumulation of lysosomal storage bodies, and brain atrophy. Similar disorders have been associated with different CNP variants in Dalmatians and in human subjects.


Assuntos
Lipofuscina , Bainha de Mielina , Humanos , Masculino , Animais , Cães , Bainha de Mielina/genética , Homozigoto , Mutação , 2',3'-Nucleotídeo Cíclico Fosfodiesterases , Atrofia
10.
Geroscience ; 46(3): 3445-3455, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38358579

RESUMO

Senescent cell number increases with age in different tissues, leading to greater senescent cell load, proinflammatory stress, and tissue dysfunction. In the current study, we tested the efficacy of senolytic drugs to reduce ovarian senescence and improve fertility in reproductive age female mice. In the first experiment, 1-month-old C57BL/6 female mice were treated every other week with D + Q (n = 24) or placebo (n = 24). At 3 and 6 months of age, female mice were mated with untreated males to evaluate pregnancy rate and litter size. In the second experiment, 6-month-old C57BL/6 female mice were treated monthly with D + Q (n = 30), fisetin (n = 30), or placebo (n = 30). Females were treated once a month until 11 months of age, then they were mated with untreated males for 30 days to evaluate pregnancy rate and litter size. In the first experiment, D + Q treatment did not affect pregnancy rate (P = 0.68), litter size (P = 0.58), or ovarian reserve (P > 0.05). Lipofuscin staining was lower in females treated with D + Q (P = 0.04), but expression of senescence genes in ovaries was similar. In the second experiment, D + Q or fisetin treatment also did not affect pregnancy rate (P = 0.37), litter size (P = 0.20), or ovarian reserve (P > 0.05). Lipofuscin staining (P = 0.008) and macrophage infiltration (P = 0.002) was lower in fisetin treated females. Overall, treatment with D + Q or fisetin did not affect ovarian reserve or fertility but did decrease some senescence markers in the ovary.


Assuntos
Reserva Ovariana , Gravidez , Masculino , Camundongos , Feminino , Animais , Senoterapia , Lipofuscina , Camundongos Endogâmicos C57BL , Fertilidade
11.
Physiol Rep ; 12(3): e15922, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38296333

RESUMO

Lipofuscin (LF) is an intracellular aggregate associated with proteostatic impairments, especially prevalent in nondividing skeletal muscle fibers. Reactive oxygen species (ROS) drive LF-formation. Resistance training (RT) improves muscle performance but also increases ROS production, potentially promoting LF-formation. Thus, we aimed to investigate if RT of a mesocycle duration increases LF-formation in type-I and II muscle fibers and whether RT increases the antioxidant capacity (AOC) in terms of SOD1 and SOD2 content. An intervention group (IG) performed 14 eccentrically accented RT-sessions within 7 weeks. Vastus lateralis muscle biopsies were collected before and after the intervention from IG as well as from a control group (CG) which refrained from RT for the same duration. LF was predominantly found near nuclei, followed by membrane-near and a minor amount in the fiber core, with corresponding spot sizes. Overall, LF-content was higher in type-I than type-II fibers (p < 0.05). There was no increase in LF-content in type-I or IIA fibers, neither for the IG following RT nor for the CG. The same is valid for SOD1/2. We conclude that, in healthy subjects, RT can be safely performed, without adverse effects on increased LF-formation.


Assuntos
Lipofuscina , Treinamento Resistido , Masculino , Humanos , Projetos Piloto , Músculo Esquelético/fisiologia , Espécies Reativas de Oxigênio , Superóxido Dismutase-1 , Fibras Musculares Esqueléticas/fisiologia
12.
Neurotoxicol Teratol ; 101: 107320, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38199312

RESUMO

INTRODUCTION: Methylmercury (MeHg) is an environmental contaminant that is of particular concern in Northern Arctic Canadian populations. Specifically, organic mercury compounds such as MeHg are potent toxicants that affect multiple bodily systems including the nervous system. Developmental exposure to MeHg is a major concern, as the developing fetus and neonate are thought to be especially vulnerable to the toxic effects of MeHg. The objective of this study was to examine developmental exposure to low doses of MeHg and effects upon the adult central nervous system (CNS). The doses of MeHg chosen were scaled to be proportional to the concentrations of MeHg that have been reported in human maternal blood samples in Northern Arctic Canadian populations. METHOD: Offspring were exposed to MeHg maternally where pregnant Sprague Dawley rats were fed cookies that contained MeHg or vehicle (vehicle corn oil; MeHg 0.02 mg/kg/body weight or 2.0 mg/kg/body weight) daily, throughout gestation (21 days) and lactation (21 days). Offspring were not exposed to MeHg after the lactation period and were euthanized on postnatal day 450. Brains were extracted, fixed, frozen, and sectioned for immunohistochemical analysis. A battery of markers of brain structure and function were selected including neuronal GABAergic enzymatic marker glutamic acid decarboxylase-67 (GAD67), apoptotic/necrotic marker cleaved caspase-3 (CC3), catecholamine marker tyrosine hydroxylase (TH), immune inflammatory marker microglia (Cd11b), endothelial cell marker rat endothelial cell antigen-1 (RECA-1), doublecortin (DCX), Bergmann glia (glial fibrillary acidic protein (GFAP)), and general nucleic acid and cellular stains Hoechst, and cresyl violet, respectively. Oxidative stress marker lipofuscin (autofluorescence) was also assessed. Both male and female offspring were included in analysis. Two-way analysis of variance (ANOVA) was utilized where sex and treatment were considered as between-subject factors (p* <0.05). ImageJ was used to assess immunohistochemical results. RESULTS: In comparison with controls, adult rat offspring exposed to both doses of MeHg were observed to have (1) increased GAD67 in the cerebellum; (2) decreased lipofuscin in the locus coeruleus; and (3) decreased GAD67 in the anterior CA1 region. Furthermore, in the substantia nigra and periaqueductal gray, adult male offspring consistently had a larger endothelial cell and capillary perimeter in comparison to females. The maternal high dose of MeHg influenced RECA-1 immunoreactivity in both the substantia nigra and periaqueductal gray of adult rat offspring, where the latter neuronal region also showed statistically significant decreases in RECA-1 immunoreactivity at the maternal low dose exposure level. Lastly, males exposed to high doses of MeHg during development exhibited a statistically significant increase in the perimeter of endothelial cells and capillaries (RECA-1) in the cerebellum, in comparison to male controls. CONCLUSION: Findings suggest that in utero and early postnatal exposure to MeHg at environmentally relevant doses leads to long-lasting and selective changes in the CNS. Exposure to MeHg at low doses may affect GABAergic homeostasis and vascular integrity of the CNS. Such changes may contribute to neurological disturbances in learning, cognition, and memory that have been reported in epidemiological studies.


Assuntos
Compostos de Metilmercúrio , Efeitos Tardios da Exposição Pré-Natal , Gravidez , Ratos , Animais , Masculino , Feminino , Humanos , Compostos de Metilmercúrio/toxicidade , Ratos Sprague-Dawley , Glutamato Descarboxilase/metabolismo , Glutamato Descarboxilase/farmacologia , Efeitos Tardios da Exposição Pré-Natal/induzido quimicamente , Efeitos Tardios da Exposição Pré-Natal/metabolismo , Capilares/metabolismo , Células Endoteliais/metabolismo , Lipofuscina/metabolismo , Lipofuscina/farmacologia , Canadá , Cerebelo , Mesencéfalo/metabolismo , Peso Corporal
13.
Gerontology ; 70(4): 408-417, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38228128

RESUMO

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/genética
14.
STAR Protoc ; 4(4): 102730, 2023 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-37980568

RESUMO

The procedures used routinely for collagen and lipofuscin evaluation are, in many cases, qualitative, observer dependent, and disregard spatial distribution. Here, we present a protocol for automatic quantification and spatial characterization of collagen and lipofuscin from label-free microscopy images of human ventricular tissues. We describe the steps for sample collection, tissue processing, image acquisition, and quantification of collagen and lipofuscin. This protocol avoids discrepancies between observers and can be adapted to other tissues and species. For complete details on the use and execution of this protocol, please refer to García-Mendívil et al. (2022).1.


Assuntos
Lipofuscina , Microscopia , Humanos , Ventrículos do Coração/diagnóstico por imagem , Miocárdio , Colágeno
15.
J Comp Pathol ; 207: 66-82, 2023 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-37977048

RESUMO

Lipofuscin accumulation has been observed in human coronary arteries but whether or not myocardial tissue can release lipofuscin generated within cardiomyocytes must be clarified, as this may provide indicators for future anti-ageing research. The hearts of Sprague Dawley rats, aged 6-24 months, were embedded in resin and ultrathin sections cut for electron microscopy. Lipofuscin granules were abundant in cardiomyocytes. Cardiomyocytes were seen to release lipofuscin granules into the myocardial interstitium as cytoplasmic fragments with irregular protrusions on the sarcolemma surface. The cytoplasmic fragments entering the stroma fused directly with the endothelial cells of adjacent capillaries, delivering lipofuscin to the vessel wall. These fragments were also seen to be engulfed by stromal macrophages or fused with fibroblasts, which then combined with capillary endothelial cells to deliver lipofuscin to the vessel wall. Some cytoplasmic fragments disaggregated and formed membrane-like waste, which travelled to the vessel wall from the myocardial stroma as soluble fine particles via diffusion or pinocytosis of capillary endothelial cells. Lipofuscin entered the vascular wall and endothelial cells, forming large and small protrusions or folds that transported the lipofuscin to the vascular lumen and bloodstream.


Assuntos
Células Endoteliais , Lipofuscina , Ratos , Humanos , Animais , Ratos Sprague-Dawley , Miocárdio , Microscopia Eletrônica/veterinária
16.
Aging (Albany NY) ; 15(21): 11764-11781, 2023 11 09.
Artigo em Inglês | MEDLINE | ID: mdl-37950722

RESUMO

Aging is a continuous degenerative process caused by a progressive decline of cell and tissue functions in an organism. It is induced by the accumulation of damage that affects normal cellular processes, ultimately leading to cell death. It has been speculated for many years that mitochondria play a key role in the aging process. In the aim of characterizing the implications of mitochondria in aging, here we used Caenorhabditis elegans (C. elegans) as an organismal model treated a panel of mitochondrial inhibitors and assessed for survival. In our study, we assessed survival by evaluating worm lifespan, and we assessed aging markers by evaluating the pharyngeal muscle contraction, the accumulation of lipofuscin pigment and ATP levels. Our results show that treatment of worms with either doxycycline, azithromycin (inhibitors of the small and the large mitochondrial ribosomes, respectively), or a combination of both, significantly extended median lifespan of C. elegans, enhanced their pharyngeal pumping rate, reduced their lipofuscin content and their energy consumption (ATP levels), as compared to control untreated worms, suggesting an aging-abrogating effect for these drugs. Similarly, DPI, an inhibitor of mitochondrial complex I and II, was capable of prolonging the median lifespan of treated worms. On the other hand, subjecting worms to vitamin C, a pro-oxidant, failed to extend C. elegans lifespan and upregulated its energy consumption, revealing an increase in ATP level. Therefore, our longevity study reveals that mitochondrial inhibitors (i.e., mitochondria-targeting antibiotics) could abrogate aging and extend lifespan in C. elegans.


Assuntos
Proteínas de Caenorhabditis elegans , Longevidade , Animais , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Antibacterianos/farmacologia , Lipofuscina/metabolismo , Mitocôndrias/metabolismo , Trifosfato de Adenosina/metabolismo
17.
Nat Commun ; 14(1): 7060, 2023 11 03.
Artigo em Inglês | MEDLINE | ID: mdl-37923732

RESUMO

Engulfment of cellular material and proteins is a key function for microglia, a resident macrophage of the central nervous system (CNS). Among the techniques used to measure microglial engulfment, confocal light microscopy has been used the most extensively. Here, we show that autofluorescence (AF) likely due to lipofuscin (lipo-AF) and typically associated with aging, can also be detected within microglial lysosomes in the young mouse brain by light microscopy. This lipo-AF signal accumulates first within microglia and it occurs earliest in white versus gray matter. Importantly, in gray matter, lipo-AF signal can confound the interpretation of antibody-labeled synaptic material within microglia in young adult mice. We further show that there is an age-dependent accumulation of lipo-AF inside and outside of microglia, which is not affected by amyloid plaques. We finally implement a robust and cost-effective strategy to quench AF in mouse, marmoset, and human brain tissue.


Assuntos
Lipofuscina , Microglia , Camundongos , Humanos , Animais , Microglia/metabolismo , Lipofuscina/metabolismo , Sistema Nervoso Central/metabolismo , Macrófagos/metabolismo , Microscopia Confocal
18.
Adv Sci (Weinh) ; 10(35): e2303381, 2023 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-37882348

RESUMO

The study of aging and neurodegenerative processes in the human brain requires a comprehensive understanding of cytoarchitectonic, myeloarchitectonic, and vascular structures. Recent computational advances have enabled volumetric reconstruction of the human brain using thousands of stained slices, however, tissue distortions and loss resulting from standard histological processing have hindered deformation-free reconstruction. Here, the authors describe an integrated serial sectioning polarization-sensitive optical coherence tomography (PSOCT) and two photon microscopy (2PM) system to provide label-free multi-contrast imaging of intact brain structures, including scattering, birefringence, and autofluorescence of human brain tissue. The authors demonstrate high-throughput reconstruction of 4 × 4 × 2cm3 sample blocks and simple registration between PSOCT and 2PM images that enable comprehensive analysis of myelin content, vascular structure, and cellular information. The high-resolution 2PM images provide microscopic validation and enrichment of the cellular information provided by the PSOCT optical properties on the same sample, revealing the densely packed fibers, capillaries, and lipofuscin-filled cell bodies in the cortex and white matter. It is  shown that the imaging system enables quantitative characterization of various pathological features in aging process, including myelin degradation, lipofuscin accumulation, and microvascular changes, which opens up numerous opportunities in the study of neurodegenerative diseases in the future.


Assuntos
Microscopia , Tomografia de Coerência Óptica , Humanos , Tomografia de Coerência Óptica/métodos , Microscopia/métodos , Lipofuscina , Encéfalo/diagnóstico por imagem , Neuroimagem
20.
Int J Mol Sci ; 24(17)2023 Aug 23.
Artigo em Inglês | MEDLINE | ID: mdl-37685907

RESUMO

It is known that during the process of aging, there is a significant decrease in the number of melanosomes in the retinal pigment epithelium (RPE) cells in the human eye. Melanosomes act as screening pigments in RPE cells and are fundamentally important for protection against the free radicals generated by light. A loss or change in the quality of melanin in melanosomes can lead to the development of senile pathologies and aggravation in the development of various retinal diseases. We have previously shown that the interaction between melanin melanosomes and superoxide radicals results in oxidative degradation with the formation of water-soluble fluorescent products. In the present study, we show, using fluorescence analysis, HPLC, and mass spectrometry, that visible light irradiation on melanolipofuscin granules isolated from RPE cells in the human eye results in the formation of water-soluble fluorescent products from oxidative degradation of melanin, which was in contrast to lipofuscin granules and melanosomes irradiation. The formation of these products occurs as a result of the oxidative degradation of melanin by superoxide radicals, which are generated by the lipofuscin part of the melanolipofuscin granule. We identified these products both in the composition of melanolipofuscin granules irradiated with visible light and in the composition of melanosomes that were not irradiated but were, instead, oxidized by superoxide radicals. In the melanolipofuscin granules irradiated by visible light, ions that could be associated with melanin oxidative degradation products were identified by applying the principal component analysis of the time-of-flight secondary ion mass spectrometry (ToF-SIMS) data. Degradation of the intact melanosomes by visible light is also possible; however, this requires significantly higher irradiation intensities than for melanolipofuscin granules. It is concluded that the decrease in the concentration of melanin in RPE cells in the human eye with age is due to its oxidative degradation by reactive oxygen species generated by lipofuscin, as part of the melanolipofuscin granules, under the action of light.


Assuntos
Lipofuscina , Superóxidos , Humanos , Melaninas , Epitélio Pigmentado da Retina , Grânulos Citoplasmáticos , Corantes
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