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A clear relationship exists between visceral obesity and type 2 diabetes, whereas subcutaneous obesity is comparatively benign. Here, we show that adipocyte-specific deletion of the coregulatory protein PRDM16 caused minimal effects on classical brown fat but markedly inhibited beige adipocyte function in subcutaneous fat following cold exposure or ß3-agonist treatment. These animals developed obesity on a high-fat diet, with severe insulin resistance and hepatic steatosis. They also showed altered fat distribution with markedly increased subcutaneous adiposity. Subcutaneous adipose tissue in mutant mice acquired many key properties of visceral fat, including decreased thermogenic and increased inflammatory gene expression and increased macrophage accumulation. Transplantation of subcutaneous fat into mice with diet-induced obesity showed a loss of metabolic benefit when tissues were derived from PRDM16 mutant animals. These findings indicate that PRDM16 and beige adipocytes are required for the "browning" of white fat and the healthful effects of subcutaneous adipose tissue.
Assuntos
Tecido Adiposo Marrom/metabolismo , Tecido Adiposo/metabolismo , Proteínas de Ligação a DNA/metabolismo , Obesidade/metabolismo , Fatores de Transcrição/metabolismo , Adipócitos/metabolismo , Animais , Proteínas de Ligação a DNA/genética , Dieta Hiperlipídica , Resistência à Insulina , Camundongos , Camundongos Knockout , Fatores de Transcrição/genéticaRESUMO
The HtrA serine peptidase 1 (HTRA1) is a multidomain secretory protein with serine-protease activity involved in the regulation of many cellular processes in both physiological and pathological conditions. HTRA1 is normally expressed in the human placenta, and its expression is higher in the first trimester compared to the third trimester, suggesting an important role of this serine protease in the early phases of human placenta development. The aim of this study was to evaluate the functional role of HTRA1 in in vitro models of human placenta in order to define the role of this serine protease in preeclampsia (PE). BeWo and HTR8/SVneo cells expressing HTRA1 were used as syncytiotrophoblast and cytotrophoblast models, respectively. Oxidative stress was induced by treating BeWo and HTR8/SVneo cells with H2O2 to mimic PE conditions in order to evaluate its effect on HTRA1 expression. In addition, HTRA1 overexpression and silencing experiments were performed to evaluate the effects on syncytialization, cell mobility, and invasion processes. Our main data showed that oxidative stress significantly increased HTRA1 expression in both BeWo and HTR8/SVneo cells. In addition, we demonstrated that HTRA1 has a pivotal role in cell motility and invasion processes. In particular, HTRA1 overexpression increased while HTRA1 silencing decreased cell motility and invasion in HTR8/SVneo cell model. In conclusion, our results suggest an important role of HTRA1 in regulating extravillous cytotrophoblast invasion and motility during the early stage of placentation in the first trimester of gestation, suggesting a key role of this serine protease in PE onset.
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Dysregulated immunity and widespread metabolic dysfunctions are the most relevant hallmarks of the passing of time over the course of adult life, and their combination at midlife is strongly related to increased vulnerability to diseases; however, the causal connection between them remains largely unclear. By combining multi-omics and functional analyses of adipose-derived stromal cells established from young (1â month) and midlife (12â months) mice, we show that an increase in expression of interferon regulatory factor 7 (IRF7) during adult life drives major metabolic changes, which include impaired mitochondrial function, altered amino acid biogenesis and reduced expression of genes involved in branched-chain amino acid (BCAA) degradation. Our results draw a new paradigm of aging as the 'sterile' activation of a cell-autonomous pathway of self-defense and identify a crucial mediator of this pathway, IRF7, as driver of metabolic dysfunction with age.
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Aminoácidos de Cadeia Ramificada , Fator Regulador 7 de Interferon , Tecido Adiposo/metabolismo , Envelhecimento/genética , Animais , Fator Regulador 7 de Interferon/metabolismo , Camundongos , Células Estromais/metabolismoRESUMO
The brown adipose tissue (BAT) is a thermogenic organ that plays a major role in energy balance, obesity, and diabetes due to the potent glucose and lipid clearance that fuels its thermogenesis, which is largely mediated via sympathetic nervous system activation. However, thus far there has been little experimental validation of the hypothesis that selective neuromodulation of the sympathetic nerves innervating the BAT is sufficient to elicit thermogenesis in mice. We generated mice expressing blue light-activated channelrhodopsin-2 (ChR2) in the sympathetic nerves innervating the BAT using two different strategies: injecting the BAT of C57Bl/6J mice with AAV6-hSyn-ChR2 (H134R)-EYFP; crossbreeding tyrosine hydroxylase-Cre mice with floxed-stop ChR2-EYFP mice. The nerves in the BAT expressing ChR2 were selectively stimulated with a blue LED light positioned underneath the fat pad of anesthetized mice, while the BAT and core temperatures were simultaneously recorded. Using immunohistochemistry we confirmed the selective expression of EYFP in TH positive nerves fibers. In addition, local optogenetic stimulation of the sympathetic nerves induced significant increase in the BAT temperature followed by an increase in core temperature in mice expressing ChR2, but not in the respective controls. The BAT activation was also paralleled by increased levels of pre-UCP1 transcript. Our results demonstrate that local optogenetic stimulation of the sympathetic nerves is sufficient to elicit BAT and core thermogenesis, thus suggesting that peripheral neuromodulation has the potential to be exploited as an alternative to pharmacotherapies to elicit organ activation and thus ameliorate type 2 diabetes and/or obesity.
Assuntos
Tecido Adiposo Marrom/metabolismo , Metabolismo Energético/fisiologia , Optogenética , Termogênese/fisiologia , Animais , Diabetes Mellitus Tipo 2/metabolismo , Glucose/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Obesidade/metabolismo , Optogenética/métodos , Sistema Nervoso Simpático/fisiologiaRESUMO
During pregnancy and lactation, subcutaneous white adipocytes in the mouse mammary gland transdifferentiate reversibly to milk-secreting epithelial cells. In this study, we demonstrate by transmission electron microscopy that in the post-lactating mammary gland interscapular multilocular adipocytes found close to the mammary alveoli contain milk protein granules. Use of the Cre-loxP recombination system allowed showing that the involuting mammary gland of whey acidic protein-Cre/R26R mice, whose secretory alveolar cells express the lacZ gene during pregnancy, contains some X-Gal-stained and uncoupling protein 1-positive interscapular multilocular adipocytes. These data suggest that during mammary gland involution some milk-secreting epithelial cells in the anterior subcutaneous depot may transdifferentiate to brown adipocytes, highlighting a hitherto unappreciated feature of mouse adipose organ plasticity.
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Adipócitos Marrons/fisiologia , Transdiferenciação Celular , Células Epiteliais/fisiologia , Lactação , Glândulas Mamárias Animais/citologia , Desmame , Adipócitos Marrons/metabolismo , Adipócitos Marrons/ultraestrutura , Animais , Linhagem da Célula , Plasticidade Celular , Células Epiteliais/metabolismo , Células Epiteliais/ultraestrutura , Feminino , Genótipo , Integrases/genética , Glândulas Mamárias Animais/metabolismo , Glândulas Mamárias Animais/ultraestrutura , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Microscopia Eletrônica de Transmissão , Proteínas do Leite/genética , Proteínas do Leite/metabolismo , Fenótipo , Gravidez , RNA não Traduzido/genética , Proteína Desacopladora 1/metabolismo , beta-Galactosidase/genética , beta-Galactosidase/metabolismoRESUMO
Brown adipose tissue (BAT) is essential for adaptive thermogenesis and dissipation of caloric excess through the activity of uncoupling protein (UCP)-1. BAT in humans is of great interest for the treatment of obesity and related diseases. In this study, the expression of Twik-related acid-sensitive K(+) channel (TASK)-1 [a pH-sensitive potassium channel encoded by the potassium channel, 2-pore domain, subfamily K, member 3 (Kcnk3) gene] correlated highly with Ucp1 expression in obese and cold-exposed mice. In addition, Task1-null mice, compared with their controls, became overweight, mainly because of an increase in white adipose tissue mass and BAT whitening. Task1(-/-)-mouse-derived brown adipocytes, compared with wild-type mouse-derived brown adipocytes, displayed an impaired ß3-adrenergic receptor response that was characterized by a decrease in oxygen consumption, Ucp1 expression, and lipolysis. This phenotype was thought to be caused by an exacerbation of mineralocorticoid receptor (MR) signaling, given that it was mimicked by corticoids and reversed by an MR inhibitor. We concluded that the K(+) channel TASK1 controls the thermogenic activity in brown adipocytes through modulation of ß-adrenergic receptor signaling.
Assuntos
Adipócitos Marrons/metabolismo , Tecido Adiposo Marrom/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Canais de Potássio de Domínios Poros em Tandem/metabolismo , Receptores Adrenérgicos beta 3/metabolismo , Receptores de Mineralocorticoides/metabolismo , Transdução de Sinais/fisiologia , Adipócitos Marrons/citologia , Tecido Adiposo Marrom/citologia , Animais , Feminino , Camundongos , Camundongos Knockout , Proteínas do Tecido Nervoso/genética , Consumo de Oxigênio/fisiologia , Canais de Potássio de Domínios Poros em Tandem/genética , Receptores de Mineralocorticoides/genética , Termogênese/fisiologiaRESUMO
Adipose tissue contains thermogenic adipocytes (i.e., brown and brite/beige) that oxidize nutrients at exceptionally high rates via nonshivering thermogenesis. Its recent discovery in adult humans has opened up new avenues to fight obesity and related disorders such as diabetes. Here, we identified miR-26a and -26b as key regulators of human white and brite adipocyte differentiation. Both microRNAs are upregulated in early adipogenesis, and their inhibition prevented lipid accumulation while their overexpression accelerated it. Intriguingly, miR-26a significantly induced pathways related to energy dissipation, shifted mitochondrial morphology toward that seen in brown adipocytes, and promoted uncoupled respiration by markedly increasing the hallmark protein of brown fat, uncoupling protein 1. By combining in silico target prediction, transcriptomics, and an RNA interference screen, we identified the sheddase ADAM metallopeptidase domain 17 (ADAM17) as a direct target of miR-26 that mediated the observed effects on white and brite adipogenesis. These results point to a novel, critical role for the miR-26 family and its downstream effector ADAM17 in human adipocyte differentiation by promoting characteristics of energy-dissipating thermogenic adipocytes.
Assuntos
Adipócitos Marrons/metabolismo , Adipogenia/genética , MicroRNAs/metabolismo , Proteínas ADAM/metabolismo , Proteína ADAM17 , Adipócitos Marrons/citologia , Adipócitos Marrons/ultraestrutura , Tecido Adiposo Branco/metabolismo , Tecido Adiposo Branco/ultraestrutura , Adulto , Diferenciação Celular/genética , Pré-Escolar , Temperatura Baixa , Simulação por Computador , Humanos , Lactente , Canais Iônicos , Masculino , MicroRNAs/genética , Mitocôndrias/metabolismo , Mitocôndrias/ultraestrutura , Proteínas Mitocondriais , Transdução de Sinais/genética , Transcriptoma/genética , Proteína Desacopladora 1 , Regulação para Cima/genéticaRESUMO
The morphological and functional differences between lipid droplets (LDs) in brown (BAT) and white (WAT) adipose tissues will largely be determined by their associated proteins. Analysing mRNA expression in mice fat depots we have found that most LD protein genes are expressed at higher levels in BAT, with the greatest differences observed for Cidea and Plin5. Prolonged cold exposure, which induces the appearance of brown-like adipocytes in mice WAT depots, was accompanied with the potentiation of the lipolytic machinery, with changes in ATGL, CGI-58 and G0S2 gene expression. However the major change detected in WAT was the enhancement of Cidea mRNA. Together with the increase in Cidec, it indicates that LD enlargement through LD-LD transference of fat is an important process during WAT browning. To study the dynamics of this phenotypic change, we have applied 4D confocal microscopy in differentiated 3T3-L1 cells under sustained ß-adrenergic stimulation. Under these conditions the cells experienced a LD remodelling cycle, with progressive reduction on the LD size by lipolysis, followed by the formation of new LDs, which were subjected to an enlargement process, likely to be CIDE-triggered, until the cell returned to the basal state. This transformation would be triggered by the activation of a thermogenic futile cycle of lipolysis/lipogenesis and could facilitate the molecular mechanism for the unilocular to multilocular transformation during WAT browning. This article is part of a Special Issue entitled Brown and White Fat: From Signaling to Disease.
Assuntos
1-Acilglicerol-3-Fosfato O-Aciltransferase/metabolismo , Tecido Adiposo Marrom/metabolismo , Tecido Adiposo Branco/metabolismo , Proteínas de Ciclo Celular/metabolismo , Lipase/metabolismo , Lipídeos/química , 1-Acilglicerol-3-Fosfato O-Aciltransferase/genética , 1-Metil-3-Isobutilxantina/farmacologia , Células 3T3-L1 , Adaptação Fisiológica , Tecido Adiposo Marrom/citologia , Tecido Adiposo Marrom/efeitos dos fármacos , Tecido Adiposo Branco/citologia , Tecido Adiposo Branco/efeitos dos fármacos , Agonistas Adrenérgicos beta/farmacologia , Animais , Proteínas de Ciclo Celular/genética , Diferenciação Celular/efeitos dos fármacos , Feminino , Processamento de Imagem Assistida por Computador , Técnicas Imunoenzimáticas , Isoproterenol/farmacologia , Lipase/genética , Lipólise , Camundongos , Microscopia Confocal , Inibidores de Fosfodiesterase/farmacologia , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais , TemperaturaRESUMO
In all mammals, white adipose tissue (WAT) and brown adipose tissue (BAT) are found together in several fat depots, forming a multi-depot organ. Adrenergic stimulation induces an increase in BAT usually referred to as "browning". This phenomenon is important because of its potential use in curbing obesity and related disorders; thus, understanding its cellular mechanisms in humans may be useful for the development of new therapeutic strategies. Data in rodents have supported the direct transformation of white into brown adipocytes. Biopsies of pure white omental fat were collected from 12 patients affected by the catecholamine-secreting tumor pheochromocytoma (pheo-patients) and compared with biopsies from controls. Half of the omental fat samples from pheo-patients contained uncoupling protein 1 (UCP1)-immunoreactive-(ir) multilocular cells that were often arranged in a BAT-like pattern endowed with noradrenergic fibers and dense capillary network. Many UCP1-ir adipocytes showed the characteristic morphology of paucilocular cells, which we have been described as cytological marker of transdifferentiation. Electron microscopy showed increased mitochondrial density in multi- and paucilocular cells and disclosed the presence of perivascular brown adipocyte precursors. Brown fat genes, such as UCP1, PR domain containing 16 (PRDM16) and ß3-adrenoreceptor, were highly expressed in the omentum of pheo-patients and in those cases without visible morphologic re-arrangement. Of note, the brown determinant PRDM16 was detected by immunohistochemistry only in nuclei of multi- and paucilocular adipocytes. Quantitative electron microscopy and immunohistochemistry for Ki67 suggest an unlikely contribution of proliferative events to the phenomenon. The data support the idea that, in adult humans, white adipocytes of pure white fat that are subjected to adrenergic stimulation are able to undergo a process of direct transformation into brown adipocytes. This article is part of a Special Issue entitled Brown and White Fat: From Signaling to Disease.
Assuntos
Adipócitos/citologia , Tecido Adiposo Marrom/citologia , Tecido Adiposo Branco/citologia , Neoplasias das Glândulas Suprarrenais/patologia , Transdiferenciação Celular , Omento/citologia , Feocromocitoma/patologia , Adipócitos/metabolismo , Tecido Adiposo Marrom/metabolismo , Tecido Adiposo Branco/metabolismo , Neoplasias das Glândulas Suprarrenais/genética , Neoplasias das Glândulas Suprarrenais/metabolismo , Adulto , Idoso , Estudos de Casos e Controles , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Feminino , Humanos , Técnicas Imunoenzimáticas , Canais Iônicos/genética , Canais Iônicos/metabolismo , Masculino , Pessoa de Meia-Idade , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Omento/metabolismo , Feocromocitoma/genética , Feocromocitoma/metabolismo , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Receptores Adrenérgicos beta 3/genética , Receptores Adrenérgicos beta 3/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteína Desacopladora 1RESUMO
Brown adipocytes dissipate energy, whereas white adipocytes are an energy storage site. We explored the plasticity of different white adipose tissue depots in acquiring a brown phenotype by cold exposure. By comparing cold-induced genes in white fat to those enriched in brown compared with white fat, at thermoneutrality we defined a "brite" transcription signature. We identified the genes, pathways, and promoter regulatory motifs associated with "browning," as these represent novel targets for understanding this process. For example, neuregulin 4 was more highly expressed in brown adipose tissue and upregulated in white fat upon cold exposure, and cell studies showed that it is a neurite outgrowth-promoting adipokine, indicative of a role in increasing adipose tissue innervation in response to cold. A cell culture system that allows us to reproduce the differential properties of the discrete adipose depots was developed to study depot-specific differences at an in vitro level. The key transcriptional events underpinning white adipose tissue to brown transition are important, as they represent an attractive proposition to overcome the detrimental effects associated with metabolic disorders, including obesity and type 2 diabetes.
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Tecido Adiposo Marrom/metabolismo , Tecido Adiposo Branco/metabolismo , Resposta ao Choque Frio/genética , Regulação da Expressão Gênica , Animais , Células Cultivadas , Feminino , Camundongos , Camundongos Endogâmicos C57BL , Análise em Microsséries , Células PC12 , Ratos , TranscriptomaRESUMO
The use of dental implants for prosthetic rehabilitation in dentistry is based on the concept of osteointegration. This concept enables the clinical stability of the implants and a total absence of inflammatory tissue between the implant surface and the bone tissue. For this reason, it is essential to understand the role of the titanium surface in promoting and maintaining or not maintaining contact between the bone matrix and the surface of the titanium implant. MATERIALS AND METHODS: Five types of titanium discs placed in contact with osteoblast cultures of osteosarcomas were studied. The materials had different roughness. Scanning electron microscopy (SEM) photos were taken before the in vitro culture to analyze the surfaces, and at the end of the culturing time, the different gene expressions of a broad pattern of proteins were evaluated to analyze the osteoblast response, as indicated in the scientific literature. RESULTS: It was demonstrated that the responses of the osteoblasts were different in the five cultures in contact with the five titanium discs with different surfaces; in particular, the response in the production of some proteins was statistically significant. DISCUSSION: The key role of titanium surfaces underlines how it is still possible to carry out increasingly accurate and targeted studies in the search for new surfaces capable of stimulating a better osteoblastic response and the long-term maintenance of osteointegration.
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Asbestos has been widely used due to its unique characteristics. It is known that exposure to asbestos causes serious damage to health but one species, chrysolite, is still used because it is considered less toxic and not biopersistent in some countries. The aim of our study was to investigate if cellular process underlying the proliferation, differentiation and cell death of placental tissues could be modify in presence of asbestos fibres (50 µg/ml final concentration), long chrysolite fibres (CHR-L) and short chrysolite fibres (CHR-S), using BeWo cell line, an in vitro model that mimics the syncytiotrophoblast (STB), the outer layer of placental villi. Our data demonstrated that none of the fibres analysed alter syncytiotrophoblast formation but all of them induce ROS formation and reduced cell proliferation. Moreover, we showed that only CHR-L fibre induced was able to induce irreversible DNA alterations that carried cells to apoptosis. In fact, BeWo cells exposed to CHR-L fibre showed a significant increase in cleaved CASP3 protein, a marker of apoptosis. These data suggest that CHR-L may induce death of the placental villi leading to impaired placental development. The impairment of placental development is the basis of many gestational pathologies such as preeclampsia and intrauterine growth retardation. Since these pathologies are very dangerous for foetal and maternal life, we suggest to the gynaecologists to carefully evaluate the area of maternal residence, the working environment, the food used, and the materials used daily to avoid contact with these fibres as much as possible.
Assuntos
Amianto , Placenta , Humanos , Gravidez , Feminino , Fibras Minerais/toxicidade , Trofoblastos/metabolismo , Amianto/metabolismo , Amianto/toxicidade , ApoptoseRESUMO
Mature adipocytes are generally considered terminally differentiated because they have lost their proliferative abilities. Here, we studied the gene expression and functional properties of mature adipocytes isolated from human omental and subcutaneous fat tissues. We also focused on dedifferentiated adipocytes in culture and their morphologies and functional changes with respect to mature adipocytes, stromal-vascular fraction (SVF)-derived mesenchymal stem cells (MSCs) and bone marrow (BM)-derived MSCs. Isolated mature adipocytes expressed stem cell and reprogramming genes. They replicated in culture after assuming a fibroblast-like shape and expanded similarly to SVF- and BM-derived MSCs. During the dedifferentiation process, mature adipocytes lost their lineage gene expression profile, assumed the typical mesenchymal morphology and immunophenotype, expressed stem cell genes and differentiated into multilineage cells. Moreover, during the dedifferentiation process, we showed changes in the epigenetic status of mature adipocytes, which led dedifferentiated adipocytes to display a similar DNA methylation condition to BM-derived MSCs. Like SVF- and BM-derived MSCs, dedifferentiated adipocytes were able to inhibit the proliferation of stimulated lymphocytes in coculture while mature adipocytes stimulated their growth. Furthermore, dedifferentiated adipocytes maintained the survival and complete differentiation characteristic of hematopoietic stem cells. This is the first study that in addition to characterizing isolated and dedifferentiated adipocytes also reports on the immunoregulatory and hematopoietic supporting functions of these cells. This structural and functional characterization might have clinical applications of both mature and dedifferentiated adipocytes in such fields, as regenerative medicine.
Assuntos
Adipócitos/metabolismo , Células da Medula Óssea/metabolismo , Desdiferenciação Celular/fisiologia , Células-Tronco Mesenquimais/metabolismo , Adipócitos/ultraestrutura , Idoso , Idoso de 80 Anos ou mais , Células da Medula Óssea/ultraestrutura , Proliferação de Células , Sobrevivência Celular/fisiologia , Células Cultivadas , Metilação de DNA/fisiologia , Epigênese Genética/fisiologia , Feminino , Humanos , Linfócitos/metabolismo , Linfócitos/ultraestrutura , Masculino , Células-Tronco Mesenquimais/ultraestrutura , Pessoa de Meia-IdadeRESUMO
The peptides encoded by the VGF gene are gaining biomedical interest and are increasingly being scrutinized as biomarkers for human disease. An endocrine/neuromodulatory role for VGF peptides has been suggested but never demonstrated. Furthermore, no study has demonstrated so far the existence of a receptor-mediated mechanism for any VGF peptide. In the present study, we provide a comprehensive in vitro, ex vivo and in vivo identification of a novel pro-lipolytic pathway mediated by the TLQP-21 peptide. We show for the first time that VGF-immunoreactivity is present within sympathetic fibres in the WAT (white adipose tissue) but not in the adipocytes. Furthermore, we identified a saturable receptor-binding activity for the TLQP-21 peptide. The maximum binding capacity for TLQP-21 was higher in the WAT as compared with other tissues, and selectively up-regulated in the adipose tissue of obese mice. TLQP-21 increases lipolysis in murine adipocytes via a mechanism encompassing the activation of noradrenaline/ß-adrenergic receptors pathways and dose-dependently decreases adipocytes diameters in two models of obesity. In conclusion, we demonstrated a novel and previously uncharacterized peripheral lipolytic pathway encompassing the VGF peptide TLQP-21. Targeting the sympathetic nerve-adipocytes interaction might prove to be a novel approach for the treatment of obesity-associated metabolic complications.
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Neuropeptídeos/metabolismo , Fragmentos de Peptídeos/farmacologia , Adipócitos/citologia , Adipócitos/efeitos dos fármacos , Animais , Composição Corporal , Gorduras na Dieta/efeitos adversos , Gorduras na Dieta/metabolismo , Masculino , Camundongos , Células NIH 3T3 , Fatores de Crescimento Neural , Obesidade/induzido quimicamente , Obesidade/metabolismo , Ligação Proteica , Transporte Proteico , Receptores de Superfície CelularRESUMO
One of the main sources of MPs contamination in fish farms is aquafeed. The present study investigated, for the first time through a comparative approach, the effects of different-sized fluorescent MPs included in a diet intended for zebrafish (Danio rerio). A comparison based on fish developmental stage (larval vs. juvenile), exposure time, and dietary MPs' size and concentration was performed. Four experimental diets were formulated, starting from the control, by adding fluorescent polymer A (size range 1-5 µm) and B (size range 40-47 µm) at two different concentrations (50 and 500 mg/kg). Zebrafish were sampled at 20 (larval phase) and 60 dpf (juvenile stage). Whole larvae, intestine, liver and muscles of juveniles were collected for the analyses. Polymer A was absorbed at the intestinal level in both larvae and juveniles, while it was evidenced at the hepatic and muscular levels only in juveniles. Hepatic accumulation caused an increase in oxidative stress markers in juveniles, but at the same time significantly reduced the number of MPs able to reach the muscle, representing an efficient barrier against the spread of MPs. Polymer B simply transited through the gut, causing an abrasive effect and an increase in goblet cell abundance in both stages.
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OBJECTIVE: Pro-peptide precursors are processed into biologically active peptide hormones or neurotransmitters, each playing an essential role in physiology and disease. Genetic loss of function of a pro-peptide precursor results in the simultaneous ablation of all biologically-active peptides within that precursor, often leading to a composite phenotype that can be difficult to align with the loss of specific peptide components. Due to this biological constraint and technical limitations, mice carrying the selective ablation of individual peptides encoded by pro-peptide precursor genes, while leaving the other peptides unaffected, have remained largely unaddressed. METHODS: We developed and characterized a mouse model carrying the selective knockout of the TLQP-21 neuropeptide (ΔTLQP-21) encoded by the Vgf gene. To achieve this goal, we used a knowledge-based approach by mutating a codon in the Vgf sequence leading to the substitution of the C-terminal Arginine of TLQP-21, which is the pharmacophore as well as an essential cleavage site from its precursor, into Alanine (R21âA). RESULTS: We provide several independent validations of this mouse, including a novel in-gel digestion targeted mass spectrometry identification of the unnatural mutant sequence, exclusive to the mutant mouse. ΔTLQP-21 mice do not manifest gross behavioral and metabolic abnormalities and reproduce well, yet they have a unique metabolic phenotype characterized by an environmental temperature-dependent resistance to diet-induced obesity and activation of the brown adipose tissue. CONCLUSIONS: The ΔTLQP-21 mouse line can be a valuable resource to conduct mechanistic studies on the necessary role of TLQP-21 in physiology and disease, while also serving as a platform to test the specificity of novel antibodies or immunoassays directed at TLQP-21. Our approach also has far-reaching implications by informing the development of knowledge-based genetic engineering approaches to generate selective loss of function of other peptides encoded by pro-hormones genes, leaving all other peptides within the pro-protein precursor intact and unmodified.
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Metabolismo Energético , Neuropeptídeos , Hormônios Peptídicos , Animais , Camundongos , Dieta , Homeostase , Neuropeptídeos/genética , Neuropeptídeos/química , Fragmentos de Peptídeos/farmacologia , Metabolismo Energético/genética , Metabolismo Energético/fisiologiaRESUMO
Pro-peptide precursors are processed into biologically active peptide hormones or neurotransmitters, each playing an essential role in physiology and disease. Genetic loss of function of a pro-peptide precursor results in the simultaneous ablation of all biologically-active peptides within that precursor, often leading to a composite phenotype that can be difficult to align with the loss of specific peptide components. Due to this biological constraint and technical limitations, mice carrying the selective ablation of individual peptides encoded by pro-peptide precursor genes, while leaving the other peptides unaffected, have remained largely unaddressed. Here, we developed and characterized a mouse model carrying the selective knockout of the TLQP-21 neuropeptide (ΔTLQP-21) encoded by the Vgf gene. To achieve this goal, we used a knowledge-based approach by mutating a codon in the Vgf sequence leading to the substitution of the C-terminal Arginine of TLQP-21, which is the pharmacophore as well as an essential cleavage site from its precursor, into Alanine (R 21 âA). We provide several independent validations of this mouse, including a novel in-gel digestion targeted mass spectrometry identification of the unnatural mutant sequence, exclusive to the mutant mouse. ΔTLQP-21 mice do not manifest gross behavioral and metabolic abnormalities and reproduce well, yet they have a unique metabolic phenotype characterized by a temperature-dependent resistance to diet-induced obesity and activation of the brown adipose tissue.
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Multipotent stem cells persist within the stromal vascular fraction (SVF) of adipose tissue during adulthood. These cells, commonly referred to as adipose-derived stromal cells (ASC), have been extensively investigated over the past years as a promising therapeutic tool based on their regenerative and immunomodulatory properties. However, how ASC might mirror the age-related alteration of the fat they reside in remains unclear. Herein, we show that inguinal adipose tissue in mice turns from brown/beige- to white-like with age and resident ASC readily mirror these changes both at mRNA and microRNA transcriptional level. Mechanistically, our data suggest that these brown/age-related changes in ASC transcription rely on changes in the activity of E2F1 and NFkB transcription factors.
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Tecido Adiposo , Células Estromais , Animais , CamundongosRESUMO
This study investigated the effect of chronic AMP-kinase (AMPK) activation with 5-aminoimidazole-4-carboxamide-1-ß-D-ribofuranoside (AICAR) on white adipose tissue (WAT) metabolism and the implications for visceral (VC) and subcutaneous (SC) adiposity, whole body-energy homeostasis, and hypothalamic leptin sensitivity. Male Wistar rats received daily single intraperitoneal injections of either saline or AICAR (0.7g/kg body weight) for 4 and 8 weeks and were pair-fed throughout the study. AICAR-treated rats had reduced adiposity with increased mitochondrial density in VC and SC fat pads, which was accompanied by reduced circulating leptin and time-dependent and depot-specific regulation of AMPK phosphorylation and FA oxidation. Interestingly, the anorectic effect to exogenous leptin was more pronounced in AICAR-treated animals than controls. This corresponded to reductions in hypothalamic AMPK phosphorylation and suppressor of cytokine signaling 3 content, whereas signal transducer and activator of transcription 3 phosphorylation was either unchanged or increased at 4 and 8 weeks in AICAR-treated rats. Ambulatory activity and whole-body energy expenditure (EE) were also increased with AICAR treatment. Altogether, chronic AICAR-induced AMPK activation increased WAT oxidative machinery, whole-body EE, and hypothalamic leptin sensitivity. This led to significant reductions in VC and SC adiposity without inducing energy-sparing mechanisms that oppose long-term fat loss.
Assuntos
Adenilato Quinase/metabolismo , Adipócitos/metabolismo , Adiposidade/efeitos dos fármacos , Aminoimidazol Carboxamida/análogos & derivados , Ativação Enzimática/efeitos dos fármacos , Hipoglicemiantes/farmacologia , Leptina/metabolismo , Ribonucleotídeos/farmacologia , Adipócitos/citologia , Tecido Adiposo/citologia , Tecido Adiposo/metabolismo , Aminoimidazol Carboxamida/farmacologia , Animais , Peso Corporal/efeitos dos fármacos , Ingestão de Alimentos/efeitos dos fármacos , Metabolismo Energético/efeitos dos fármacos , Hipotálamo/metabolismo , Masculino , Mitocôndrias/metabolismo , Mitocôndrias/ultraestrutura , Palmitatos/metabolismo , Ratos , Ratos WistarRESUMO
We appreciate very much the interest of Mirabelli et al. [...].