RESUMO
Activation of energy-dissipating brown/beige adipocytes represents an attractive therapeutic strategy against metabolic disorders. While lactate is known to induce beiging through the regulation of Ucp1 gene expression, the role of lactate transporters on beige adipocytes' ongoing metabolic activity remains poorly understood. To explore the function of the lactate-transporting monocarboxylate transporters (MCTs), we used a combination of primary cell culture studies, 13C isotopic tracing, laser microdissection experiments, and in situ immunofluorescence of murine adipose fat pads. Dissecting white adipose tissue heterogeneity revealed that the MCT1 is expressed in inducible beige adipocytes as the emergence of uncoupling protein 1 after cold exposure was restricted to a subpopulation of MCT1-expressing adipocytes suggesting MCT1 as a marker of inducible beige adipocytes. We also observed that MCT1 mediates bidirectional and simultaneous inward and outward lactate fluxes, which were required for efficient utilization of glucose by beige adipocytes activated by the canonical ß3-adrenergic signaling pathway. Finally, we demonstrated that significant lactate import through MCT1 occurs even when glucose is not limiting, which feeds the oxidative metabolism of beige adipocytes. These data highlight the key role of lactate fluxes in finely tuning the metabolic activity of beige adipocytes according to extracellular metabolic conditions and reinforce the emerging role of lactate metabolism in the control of energy homeostasis.
Assuntos
Adipócitos Bege/metabolismo , Regulação da Expressão Gênica , Ácido Láctico/metabolismo , Células-Tronco Mesenquimais/metabolismo , Transportadores de Ácidos Monocarboxílicos/metabolismo , Simportadores/metabolismo , Adipócitos Bege/citologia , Animais , Masculino , Células-Tronco Mesenquimais/citologia , Camundongos , Camundongos Endogâmicos C57BL , Transportadores de Ácidos Monocarboxílicos/genética , Transdução de Sinais , Simportadores/genética , TermogêneseRESUMO
Human adipose-derived stem/stromal cells (hASCs) can differentiate into specialized cell types and thereby contribute to tissue regeneration. As such, hASCs have drawn increasing attention in cell therapy and regenerative medicine, not to mention the ease to isolate them from donors. Culture conditions are critical for expanding hASCs while maintaining optimal therapeutic capabilities. Here, we identified a role for transforming growth factor ß1 (TGFß1) in culture medium in influencing the fate of hASCs during in vitro cell expansion. Human ASCs obtained after expansion in standard culture medium (Standard-hASCs) and in endothelial cell growth medium 2 (EGM2-hASCs) were characterized by high-throughput transcriptional studies, gene set enrichment analysis and functional properties. EGM2-hASCs exhibited enhanced multipotency capabilities and an immature phenotype compared with Standard-hASCs. Moreover, the adipogenic potential of EGM2-hASCs was enhanced, including toward beige adipogenesis, compared with Standard-hASCs. In these conditions, TGFß1 acts as a critical factor affecting the immaturity and multipotency of Standard-hASCs, as suggested by small mother of decapentaplegic homolog 3 (SMAD3) nuclear localization and phosphorylation in Standard-hASCs vs EGM2-hASCs. Finally, the typical priming of Standard-hASCs into osteoblast, chondroblast, and vascular smooth muscle cell (VSMC) lineages was counteracted by pharmacological inhibition of the TGFß1 receptor, which allowed retention of SMAD3 into the cytoplasm and a decrease in expression of osteoblast and VSMC lineage markers. Overall, the TGFß1 pathway appears critical in influencing the commitment of hASCs toward osteoblast, chondroblast, and VSMC lineages, thus reducing their adipogenic potential. These effects can be counteracted by using EGM2 culture medium or chemical inhibition of the TGFß1 pathway.
Assuntos
Adipócitos Bege/metabolismo , Adipócitos Brancos/metabolismo , Tecido Adiposo/metabolismo , Células Estromais/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Proliferação de Células , Células Cultivadas , Meios de Cultura , HumanosRESUMO
FGF21 (fibroblast growth factor 21), first described as a main fasting-responsive molecule in the liver, has been shown to act as a true metabolic regulator in additional tissues, including muscle and adipose tissues. In the present study, we found that the expression and secretion of FGF21 was very rapidly increased following lactate exposure in adipocytes. Using different pharmacological and knockout mice models, we demonstrated that lactate regulates Fgf21 expression through a NADH/NAD-independent pathway, but requires active p38-MAPK (mitogen activated protein kinase) signalling. We also demonstrated that this effect is not restricted to lactate as additional metabolites including pyruvate and ketone bodies also activated the FGF21 stress response. FGF21 release by adipose cells in response to an excess of intermediate metabolites may represent a physiological mechanism by which the sensing of environmental metabolic conditions results in the release of FGF21 to improve metabolic adaptations.
Assuntos
Adipócitos/metabolismo , Fatores de Crescimento de Fibroblastos/metabolismo , Lactatos/metabolismo , Sistema de Sinalização das MAP Quinases/fisiologia , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Fator 4 Ativador da Transcrição/genética , Fator 4 Ativador da Transcrição/metabolismo , Adipócitos/fisiologia , Animais , Fatores de Crescimento de Fibroblastos/genética , Regulação da Expressão Gênica/fisiologia , Canais Iônicos/genética , Canais Iônicos/metabolismo , Camundongos , Camundongos Knockout , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , PPAR alfa/genética , PPAR alfa/metabolismo , PPAR gama/genética , PPAR gama/metabolismo , Sirtuína 3/genética , Sirtuína 3/metabolismo , Sirtuínas/genética , Sirtuínas/metabolismo , Proteína Desacopladora 1 , Regulação para Cima , Proteínas Quinases p38 Ativadas por Mitógeno/genéticaRESUMO
Identification of molecular mechanisms involved in generation of different types of adipocytes is progressing substantially in mice. However, much less is known regarding characterization of brown (BAP) and white adipocyte progenitors (WAPs) in humans, highlighting the need for an in vitro model of human adipocyte development. Here, we report a procedure to selectively derive BAP and WAPs from human-induced pluripotent stem cells. Molecular characterization of APs of both phenotypes revealed that BMP4, Hox8, Hoxc9, and HoxA5 genes were specifically expressed in WAPs, whereas expression of PRDM16, Dio2, and Pax3 marked BAPs. We focused on Pax3 and we showed that expression of this transcription factor was enriched in human perirenal white adipose tissue samples expressing UCP1 and in human classical brown fat. Finally, functional experiments indicated that Pax3 was a critical player of human AP fate as its ectopic expression led to convert WAPs into brown-like APs. Together, these data support a model in which Pax3 is a new marker of human BAPs and a molecular mediator of their fate. The findings of this study could lead to new anti-obesity therapies based on the recruitment of APs and constitute a platform for investigating in vitro the developmental origins of human white and brown adipocytes.
Assuntos
Adipócitos Marrons/citologia , Adipócitos Brancos/citologia , Diferenciação Celular , Células-Tronco Pluripotentes Induzidas/citologia , Fatores de Transcrição Box Pareados/metabolismo , Adipócitos Marrons/efeitos dos fármacos , Adipócitos Marrons/metabolismo , Adipócitos Brancos/efeitos dos fármacos , Adipócitos Brancos/metabolismo , Adipogenia/efeitos dos fármacos , Idoso de 80 Anos ou mais , Animais , Diferenciação Celular/efeitos dos fármacos , Feminino , Humanos , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Células-Tronco Pluripotentes Induzidas/metabolismo , Masculino , Camundongos , Fator de Transcrição PAX3 , Fenótipo , Tretinoína/farmacologiaRESUMO
Alzheimer's disease is strongly linked to metabolic abnormalities. We aimed to distinguish amyloid-positive people who progressed to cognitive decline from those who remained cognitively intact. We performed untargeted metabolomics of blood samples from amyloid-positive individuals, before any sign of cognitive decline, to distinguish individuals who progressed to cognitive decline from those who remained cognitively intact. A plasma-derived metabolite signature was developed from Supercritical Fluid chromatography coupled with high-resolution mass spectrometry (SFC-HRMS) and nuclear magnetic resonance (NMR) metabolomics. The 2 metabolomics data sets were analyzed by Data Integration Analysis for Biomarker discovery using Latent approaches for Omics studies (DIABLO), to identify a minimum set of metabolites that could describe cognitive decline status. NMR or SFC-HRMS data alone cannot predict cognitive decline. However, among the 320 metabolites identified, a statistical method that integrated the 2 data sets enabled the identification of a minimal signature of 9 metabolites (3-hydroxybutyrate, citrate, succinate, acetone, methionine, glucose, serine, sphingomyelin d18:1/C26:0 and triglyceride C48:3) with a statistically significant ability to predict cognitive decline more than 3 years before decline. This metabolic fingerprint obtained during this exploratory study may help to predict amyloid-positive individuals who will develop cognitive decline. Due to the high prevalence of brain amyloid-positivity in older adults, identifying adults who will have cognitive decline will enable the development of personalized and early interventions.
Assuntos
Doença de Alzheimer , Disfunção Cognitiva , Humanos , Idoso , Vida Independente , Doença de Alzheimer/metabolismo , Amiloide/metabolismo , Disfunção Cognitiva/metabolismo , Encéfalo/metabolismo , Metabolômica , Proteínas Amiloidogênicas , Peptídeos beta-Amiloides/metabolismo , BiomarcadoresRESUMO
The extent and duration of MAPK (mitogen-activated protein kinase) signalling govern a diversity of normal and aberrant cellular outcomes. Genetic and pharmacological disruption of the MAPK-activated kinase RSK (ribosomal S6 kinase) leads to elevated MAPK activity indicative of a RSK-dependent negative feedback loop. Using biochemical, pharmacological and quantitative MS approaches we show that RSK phosphorylates the Ras activator SOS1 (Son of Sevenless homologue 1) in cultured cells on two C-terminal residues, Ser(1134) and Ser(1161). Furthermore, we find that RSK-dependent SOS1 phosphorylation creates 14-3-3-binding sites. We show that mutating Ser(1134) and Ser(1161) disrupts 14-3-3 binding and modestly increases and extends MAPK activation. Together these data suggest that one mechanism whereby RSK negatively regulates MAPK activation is via site-specific SOS1 phosphorylation.
Assuntos
Proteínas 14-3-3/metabolismo , Sistema de Sinalização das MAP Quinases , Proteínas Quinases S6 Ribossômicas/metabolismo , Proteína SOS1/metabolismo , Substituição de Aminoácidos , Animais , Sítios de Ligação , Células COS , Células Cultivadas , Chlorocebus aethiops , Células HEK293 , Humanos , Camundongos , Modelos Biológicos , Mutagênese Sítio-Dirigida , Células NIH 3T3 , Fosforilação , Ratos , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Proteína SOS1/química , Proteína SOS1/genética , Serina/químicaRESUMO
In mammals, typically two types of adipose tissues are described: the white and the brown adipose tissue (WAT and BAT respectively). Whereas WAT represents the main energy storage in the organism, BAT dissipates energy as heat through the expression of the uncoupling protein UCP1 (uncoupling protein-1) that uncouples the functioning of the respiratory chain from ATP synthase. While both white and brown adipocytes have been considered for a long time as two very close cellular types sharing a common precursor, recent data challenge these conclusions and propose the existence of a new possible type of adipocyte, the BRITE (brown-in-white) adipocyte. In parallel, the recent discovery of significant amounts of BAT in human adults has renewed the interest of the scientific community for this tissue. Given its considerable capacity to dissipate substrates, BAT appears again as a therapeutic target against metabolic diseases such as diabetes and obesity. This review's objective is to discuss recent literature and to highlight elements to be clarified.
Assuntos
Adipócitos Marrons/fisiologia , Adipócitos Brancos/fisiologia , Doenças Metabólicas/terapia , Animais , Diabetes Mellitus/terapia , Metabolismo Energético , Humanos , Canais Iônicos/fisiologia , Proteínas Mitocondriais/fisiologia , Obesidade/terapia , Proteína Desacopladora 1RESUMO
Obesity and type 2 diabetes are becoming a global sociobiomedical burden. Beige adipocytes are emerging as key inducible actors and putative relevant therapeutic targets for improving metabolic health. However, in vitro models of human beige adipose tissue are currently lacking and hinder research into this cell type and biotherapy development. Unlike traditional bottom-up engineering approaches that aim to generate building blocks, here a scalable system is proposed to generate pre-vascularized and functional human beige adipose tissue organoids using the human stromal vascular fraction of white adipose tissue as a source of adipose and endothelial progenitors. This engineered method uses a defined biomechanical and chemical environment using tumor growth factor ß (TGFß) pathway inhibition and specific gelatin methacryloyl (GelMA) embedding parameters to promote the self-organization of spheroids in GelMA hydrogel, facilitating beige adipogenesis and vascularization. The resulting vascularized organoids display key features of native beige adipose tissue including inducible Uncoupling Protein-1 (UCP1) expression, increased uncoupled mitochondrial respiration, and batokines secretion. The controlled assembly of spheroids allows to translate organoid morphogenesis to a macroscopic scale, generating vascularized centimeter-scale beige adipose micro-tissues. This approach represents a significant advancement in developing in vitro human beige adipose tissue models and facilitates broad applications ranging from basic research to biotherapies.
Assuntos
Diabetes Mellitus Tipo 2 , Humanos , Diabetes Mellitus Tipo 2/metabolismo , Obesidade/metabolismo , Adipogenia , Tecido Adiposo Branco/metabolismo , Organoides/metabolismoRESUMO
Identifying mechanisms underlying relapse is a major clinical issue for effective cancer treatment. The emerging understanding of the importance of metastasis in hematologic malignancies suggests that it could also play a role in drug resistance and relapse in acute myeloid leukemia (AML). In a cohort of 1,273 AML patients, we uncovered that the multifunctional scavenger receptor CD36 was positively associated with extramedullary dissemination of leukemic blasts, increased risk of relapse after intensive chemotherapy, and reduced event-free and overall survival. CD36 was dispensable for lipid uptake but fostered blast migration through its binding with thrombospondin-1. CD36-expressing blasts, which were largely enriched after chemotherapy, exhibited a senescent-like phenotype while maintaining their migratory ability. In xenograft mouse models, CD36 inhibition reduced metastasis of blasts and prolonged survival of chemotherapy-treated mice. These results pave the way for the development of CD36 as an independent marker of poor prognosis in AML patients and a promising actionable target to improve the outcome of patients. SIGNIFICANCE: CD36 promotes blast migration and extramedullary disease in acute myeloid leukemia and represents a critical target that can be exploited for clinical prognosis and patient treatment.
Assuntos
Leucemia Mieloide Aguda , Humanos , Animais , Camundongos , Leucemia Mieloide Aguda/patologia , Resultado do Tratamento , Prognóstico , Recidiva , Crise Blástica/patologia , Doença CrônicaRESUMO
The Ras/mitogen-activated protein kinase (MAPK) pathway regulates a variety of cellular processes by activating specific transcriptional and translational programs. Ras/MAPK signaling promotes mRNA translation and protein synthesis, but the exact molecular mechanisms underlying this regulation remain poorly understood. Increasing evidence suggests that the mammalian target of rapamycin (mTOR) plays an essential role in this process. Here, we show that Raptor, an essential scaffolding protein of the mTOR complex 1 (mTORC1), becomes phosphorylated on proline-directed sites following activation of the Ras/MAPK pathway. We found that ERK1 and ERK2 interact with Raptor in cells and mediate its phosphorylation in vivo and in vitro. Using mass spectrometry and phosphospecific antibodies, we found three proline-directed residues within Raptor, Ser(8), Ser(696), and Ser(863), which are directly phosphorylated by ERK1/2. Expression of phosphorylation-deficient alleles of Raptor revealed that phosphorylation of these sites by ERK1/2 normally promotes mTORC1 activity and signaling to downstream substrates, such as 4E-BP1. Our data provide a novel regulatory mechanism by which mitogenic and oncogenic activation of the Ras/MAPK pathway promotes mTOR signaling.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Proteínas/metabolismo , Proteínas ras/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/química , Sequência de Aminoácidos , Sítios de Ligação , Linhagem Celular , Proliferação de Células , Humanos , Sistema de Sinalização das MAP Quinases , Alvo Mecanístico do Complexo 1 de Rapamicina , Dados de Sequência Molecular , Complexos Multiproteicos , Fosforilação , Prolina/metabolismo , Proteína Regulatória Associada a mTOR , Serina-Treonina Quinases TORRESUMO
Compared to cell suspensions or monolayers, 3D cell aggregates provide cellular interactions organized in space and heterogeneity that better resume the real organization of native tissues. They represent powerful tools to narrow down the gap between in vitro and in vivo models, thanks to their self-evolving capabilities. Recent strategies have demonstrated their potential as building blocks to generate microtissues. Developing specific methodologies capable of organizing these cell aggregates into 3D architectures and environments has become essential to convert them into functional microtissues adapted for regenerative medicine or pharmaceutical screening purposes. Although the techniques for producing individual cell aggregates have been on the market for over a decade, the methodology for engineering functional tissues starting from them is still a young and quickly evolving field of research. In this review, we first present a panorama of emerging cell aggregates microfabrication and assembly technologies. We further discuss the perspectives opened in the establishment of functional tissues with a specific focus on controlled architecture and heterogeneity to favor cell differentiation and proliferation.
Assuntos
Medicina Regenerativa , Engenharia Tecidual , Ciclo Celular , Diferenciação Celular , Microtecnologia , Engenharia Tecidual/métodosRESUMO
The rapamycin-sensitive mTOR complex 1 (mTORC1) promotes protein synthesis, cell growth, and cell proliferation in response to growth factors and nutritional cues. To elucidate the poorly defined mechanisms underlying mTORC1 regulation, we have studied the phosphorylation of raptor, an mTOR-interacting partner. We have identified six raptor phosphorylation sites that lie in two centrally localized clusters (cluster 1, Ser(696)/Thr(706) and cluster 2, Ser(855)/Ser(859)/Ser(863)/Ser(877)) using tandem mass spectrometry and generated phosphospecific antibodies for each of these sites. Here we focus primarily although not exclusively on raptor Ser(863) phosphorylation. We report that insulin promotes mTORC1-associated phosphorylation of raptor Ser(863) via the canonical PI3K/TSC/Rheb pathway in a rapamycin-sensitive manner. mTORC1 activation by other stimuli (e.g. amino acids, epidermal growth factor/MAPK signaling, and cellular energy) also promote raptor Ser(863) phosphorylation. Rheb overexpression increases phosphorylation on raptor Ser(863) as well as on the five other identified sites (e.g. Ser(859), Ser(855), Ser(877), Ser(696), and Thr(706)). Strikingly, raptor Ser(863) phosphorylation is absolutely required for raptor Ser(859) and Ser(855) phosphorylation. These data suggest that mTORC1 activation leads to raptor multisite phosphorylation and that raptor Ser(863) phosphorylation functions as a master biochemical switch that modulates hierarchical raptor phosphorylation (e.g. on Ser(859) and Ser(855)). Importantly, mTORC1 containing phosphorylation site-defective raptor exhibits reduced in vitro kinase activity toward the substrate 4EBP1, with a multisite raptor 6A mutant more strongly defective that single-site raptor S863A. Taken together, these data suggest that complex raptor phosphorylation functions as a biochemical rheostat that modulates mTORC1 signaling in accordance with environmental cues.
Assuntos
Proteínas de Transporte/química , Proteínas de Transporte/metabolismo , Proteínas/química , Proteínas/metabolismo , Serina/metabolismo , Fatores de Transcrição/metabolismo , Proteínas Adaptadoras de Transdução de Sinal , Sequência de Aminoácidos , Animais , Linhagem Celular , Fator de Crescimento Epidérmico/farmacologia , Humanos , Insulina/farmacologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Alvo Mecanístico do Complexo 1 de Rapamicina , Camundongos , Modelos Biológicos , Dados de Sequência Molecular , Proteínas Monoméricas de Ligação ao GTP/metabolismo , Complexos Multiproteicos , Neuropeptídeos/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Fosforilação/efeitos dos fármacos , Proteína Enriquecida em Homólogo de Ras do Encéfalo , Ratos , Proteína Regulatória Associada a mTOR , Relação Estrutura-Atividade , Serina-Treonina Quinases TOR , Espectrometria de Massas em Tandem , Termodinâmica , Proteína 1 do Complexo Esclerose Tuberosa , Proteínas Supressoras de Tumor/metabolismoRESUMO
BACKGROUND: The mammalian target of rapamycin (mTOR) is a Ser/Thr kinase that controls cell growth in response to mitogens, as well as amino acid and energy sufficiency. The scaffolding protein Raptor binds to mTOR and recruits substrates to the rapamycin-sensitive mTOR complex 1 (mTORC1). Although Raptor has been shown to be essential for mTORC1 activity, the mechanisms regulating Raptor function remain unknown. RESULTS: Here, we demonstrate that Raptor becomes highly phosphorylated on RXRXXpS/T consensus motifs after activation of the Ras/mitogen-activated protein kinase (MAPK) pathway. Using pharmacological inhibitors and RNA interference, we show that the p90 ribosomal S6 kinases (RSKs) 1 and 2 are required for Raptor phosphorylation in vivo and directly phosphorylate Raptor in vitro. Quantitative mass spectrometry and site-directed mutagenesis revealed that RSK specifically phosphorylates Raptor within an evolutionarily conserved region with no previously known function. Interestingly, expression of oncogenic forms of Ras and MEK that elevate mTORC1 activity induced strong and constitutive phosphorylation of Raptor on these residues. Importantly, we demonstrate that expression of Raptor mutants lacking RSK-dependent phosphorylation sites markedly reduced mTOR phosphotransferase activity, indicating that RSK-mediated phosphorylation of Raptor is important for mTORC1 activation by the Ras/MAPK pathway. CONCLUSIONS: We propose a unique mode of mTOR regulation in which RSK-mediated phosphorylation of Raptor regulates mTORC1 activity and thus suggest a means by which the Ras/MAPK pathway might promote rapamycin-sensitive signaling independently of the PI3K/Akt pathway.
Assuntos
Sistema de Sinalização das MAP Quinases/fisiologia , Proteínas/metabolismo , Proteínas Quinases S6 Ribossômicas 90-kDa/fisiologia , Fatores de Transcrição/metabolismo , Proteínas Adaptadoras de Transdução de Sinal , Animais , Linhagem Celular , Células HeLa , Humanos , Alvo Mecanístico do Complexo 1 de Rapamicina , Camundongos , Modelos Biológicos , Complexos Multiproteicos , Células NIH 3T3 , Fosforilação , Proteínas Quinases/metabolismo , Proteínas/química , Proteína Regulatória Associada a mTOR , Serina/metabolismo , Especificidade por Substrato , Serina-Treonina Quinases TORRESUMO
Lactate, a metabolite produced when the glycolytic flux exceeds mitochondrial oxidative capacities, is now viewed as a critical regulator of metabolism by acting as both a carbon and electron carrier and a signaling molecule between cells and tissues. In recent years, increasing evidence report its key role in white, beige, and brown adipose tissue biology, and highlights new mechanisms by which lactate participates in the maintenance of whole-body energy homeostasis. Lactate displays a wide range of biological effects in adipose cells not only through its binding to the membrane receptor but also through its transport and the subsequent effect on intracellular metabolism notably on redox balance. This study explores how lactate regulates adipocyte metabolism and plasticity by balancing intracellular redox state and by regulating specific signaling pathways. We also emphasized the contribution of adipose tissues to the regulation of systemic lactate metabolism, their roles in redox homeostasis, and related putative physiopathological repercussions associated with their decline in metabolic diseases and aging.
RESUMO
OBJECTIVE: Transplantation of adipose-derived stroma cells (ADSCs) stimulates neovascularization after experimental ischemic injury. ADSC proangiogenic potential is likely mediated by their ability to differentiate into endothelial cells and produce a wide array of angiogenic and antiapoptotic factors. Mitochondrial reactive oxygen species (ROS) have been shown to control ADSC differentiation. We therefore hypothesized that mitochondrial ROS production may change the ADSC proangiogenic properties. METHODS AND RESULTS: The use of pharmacological strategies (mitochondrial inhibitors, antimycin, and rotenone, with or without antioxidants) allowed us to specifically and precisely modulate mitochondrial ROS generation in ADSCs. We showed that transient stimulation of mitochondrial ROS generation in ADSCs before their injection in ischemic hindlimb strongly improved revascularization and the number of ADSC-derived CD31-positive cells in ischemic area. Mitochondrial ROS generation increased the secretion of the proangiogenic and antiapoptotic factors, VEGF and HGF, but did not affect ADSC ability to differentiate into endothelial cells, in vitro. Moreover, mitochondrial ROS-induced ADSC preconditioning greatly protect ADSCs against oxidative stress-induced cell death. CONCLUSIONS: Our study demonstrates that in vitro preconditioning by moderate mitochondrial ROS generation strongly increases in vivo ADSC proangiogenic properties and emphasizes the crucial role of mitochondrial ROS in ADSC fate.
Assuntos
Diferenciação Celular/fisiologia , Células Endoteliais/citologia , Células Endoteliais/fisiologia , Mitocôndrias/metabolismo , Neovascularização Fisiológica/fisiologia , Espécies Reativas de Oxigênio/metabolismo , Adipócitos , Animais , Células Cultivadas , Masculino , Camundongos , Traumatismo por Reperfusão/fisiopatologia , Células Estromais/citologia , Células Estromais/metabolismoRESUMO
Thermogenic (brown and beige) adipose tissues improve glucose and lipid homeostasis and therefore represent putative targets to cure obesity and related metabolic diseases including type II diabetes. Beside decades of research and the very well-described role of noradrenergic signaling, mechanisms underlying adipocytes plasticity and activation of thermogenic adipose tissues remain incompletely understood. Recent studies show that metabolites such as lactate control the oxidative capacity of thermogenic adipose tissues. Long time viewed as a metabolic waste product, lactate is now considered as an important metabolic substrate largely feeding the oxidative metabolism of many tissues, acting as a signaling molecule and as an inter-cellular and inter-tissular redox carrier. In this review, we provide an overview of the recent findings highlighting the importance of lactate in adipose tissues, from its production to its role as a browning inducer and its metabolic links with brown adipose tissue. We also discuss additional function(s) than thermogenesis ensured by brown and beige adipose tissues, i.e., their ability to dissipate high redox pressure and oxidative stress thanks to the activity of the uncoupling protein-1, helping to maintain tissue and whole organism redox homeostasis and integrity.
Assuntos
Tecido Adiposo Bege/metabolismo , Tecido Adiposo Marrom/metabolismo , Ácido Láctico/metabolismo , Proteína Desacopladora 1/metabolismo , Tecido Adiposo Bege/citologia , Tecido Adiposo Marrom/citologia , Animais , Metabolismo Energético , Humanos , Oxirredução , Estresse Oxidativo , TermogêneseRESUMO
Atrial natriuretic peptide (ANP) is a cardiac hormone controlling blood volume and pressure in mammals. It is still unclear whether ANP controls cold-induced thermogenesis in vivo. Here, we show that acute cold exposure induces cardiac ANP secretion in mice and humans. Genetic inactivation of ANP promotes cold intolerance and suppresses half of cold-induced brown adipose tissue (BAT) activation in mice. While white adipocytes are resistant to ANP-mediated lipolysis at thermoneutral temperature in mice, cold exposure renders white adipocytes fully responsive to ANP to activate lipolysis and a thermogenic program, a physiological response that is dramatically suppressed in ANP null mice. ANP deficiency also blunts liver triglycerides and glycogen metabolism, thus impairing fuel availability for BAT thermogenesis. ANP directly increases mitochondrial uncoupling and thermogenic gene expression in human white and brown adipocytes. Together, these results indicate that ANP is a major physiological trigger of BAT thermogenesis upon cold exposure in mammals.
Assuntos
Fator Natriurético Atrial/metabolismo , Termogênese/fisiologia , Animais , Humanos , Masculino , Camundongos , Camundongos KnockoutRESUMO
Cold Atmospheric Plasma (CAP) is a novel promising tool developed in several biomedical applications such as cutaneous wound healing or skin cancer. Nevertheless, in vitro studies are lacking regarding to CAP effects on cellular actors involved in healthy skin healing and regarding to the mechanism of action. In this study, we investigated the effect of a 3 minutes exposure to CAP-Helium on human dermal fibroblasts and Adipose-derived Stromal Cells (ASC) obtained from the same tissue sample. We observed that CAP treatment did not induce cell death but lead to proliferation arrest with an increase in p53/p21 and DNA damages. Interestingly we showed that CAP treated dermal fibroblasts and ASC developed a senescence phenotype with p16 expression, characteristic morphological changes, Senescence-Associated ß-galactosidase expression and the secretion of pro-inflammatory cytokines defined as the Senescence-Associated Secretory Phenotype (SASP). Moreover this senescence phenotype is associated with a glycolytic switch and an increase in mitochondria content. Despite this senescence phenotype, cells kept in vitro functional properties like differentiation potential and immunomodulatory effects. To conclude, we demonstrated that two main skin cellular actors are resistant to cell death but develop a senescence phenotype while maintaining some functional characteristics after 3 minutes of CAP-Helium treatment in vitro.
Assuntos
Ciclo Celular/efeitos dos fármacos , Senescência Celular/efeitos dos fármacos , Fibroblastos/efeitos dos fármacos , Células-Tronco Mesenquimais/efeitos dos fármacos , Gases em Plasma/farmacologia , Tecido Adiposo/citologia , Tecido Adiposo/metabolismo , Ciclo Celular/genética , Senescência Celular/genética , Inibidor p16 de Quinase Dependente de Ciclina/genética , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Inibidor de Quinase Dependente de Ciclina p21/genética , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Fibroblastos/citologia , Fibroblastos/metabolismo , Regulação da Expressão Gênica , Hélio/química , Humanos , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/metabolismo , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Cultura Primária de Células , Transdução de Sinais , Pele/citologia , Pele/metabolismo , Fatores de Tempo , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismoRESUMO
Native human subcutaneous adipose tissue (AT) is well organized into unilocular adipocytes interspersed within dense vascularization. This structure is completely lost under standard culture conditions and may impair the comparison with native tissue. Here, we developed a 3-D model of human white AT reminiscent of the cellular architecture found in vivo. Starting with adipose progenitors derived from the stromal-vascular fraction of human subcutaneous white AT, we generated spheroids in which endogenous endothelial cells self-assembled to form highly organized endothelial networks among stromal cells. Using an optimized adipogenic differentiation medium to preserve endothelial cells, we obtained densely vascularized spheroids containing mature adipocytes with unilocular lipid vacuoles. In vivo study showed that when differentiated spheroids were transplanted in immune-deficient mice, endothelial cells within the spheroids connected to the recipient circulatory system, forming chimeric vessels. In addition, adipocytes of human origin were still observed in transplanted mice. We therefore have developed an in vitro model of vascularized human AT-like organoids that constitute an excellent tool and model for any study of human AT.