RESUMO
Caveolae are an abundant, but enigmatic, plasma membrane feature of vertebrate cells. In this brief commentary, the authors attempt to answer some key questions related to the formation and function of caveolae based on round-table discussions at the first EMBO Workshop on Caveolae held in France in May 2019.
Assuntos
Cavéolas , Caveolinas , Animais , Membrana CelularRESUMO
RATIONALE: Metabolic syndrome (MetS) is a cluster of interrelated risk factors for cardiovascular diseases and atherosclerosis. Circulating levels of large extracellular vesicles (lEVs), submicrometer-sized vesicles released from plasma membrane, from MetS patients were shown to induce endothelial dysfunction, but their role in early stage of atherosclerosis and on vascular smooth muscle cells (SMC) remain to be fully elucidated. OBJECTIVE: To determine the mechanisms by which lEVs lead to the progression of atherosclerosis in the setting of MetS. METHODS AND RESULTS: Proteomic analysis revealed that the small GTPase, Rap1 was overexpressed in lEVs from MetS patients compared with those from non-MetS subjects. Rap1 was in GTP-associated active state in both types of lEVs, and Rap1-lEVs levels correlated with increased cardiovascular risks, including stenosis. MetS-lEVs, but not non-MetS-lEVs, increased Rap1-dependent endothelial cell permeability. MetS-lEVs significantly promoted migration and proliferation of human aortic SMC and increased expression of proinflammatory molecules and activation of ERK (extracellular signal-regulated kinase) 5/p38 pathways. Neutralization of Rap1 by specific antibody or pharmacological inhibition of Rap1 completely prevented the effects of lEVs from MetS patients. High-fat diet-fed ApoE-/- mice displayed an increased expression of Rap1 both in aortas and circulating lEVs. lEVs accumulated in plaque atherosclerotic lesions depending on the progression of atherosclerosis. lEVs from high-fat diet-fed ApoE-/- mice, but not those from mice fed with a standard diet, enhanced SMC proliferation. Human atherosclerotic lesions were enriched in lEVs expressing Rap1. CONCLUSIONS: These data demonstrate that Rap1 carried by MetS-lEVs participates in the enhanced SMC proliferation, migration, proinflammatory profile, and activation of ERK5/p38 pathways leading to vascular inflammation and remodeling, and atherosclerosis. These results highlight that Rap1 carried by MetS-lEVs may be a novel determinant of diagnostic value for cardiometabolic risk factors and suggest Rap1 as a promising therapeutic target against the development of atherosclerosis. Graphical Abstract: A graphical abstract is available for this article.
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Aterosclerose/metabolismo , Células Endoteliais/metabolismo , Vesículas Extracelulares/metabolismo , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/metabolismo , Placa Aterosclerótica , Proteínas rap1 de Ligação ao GTP/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Aterosclerose/sangue , Aterosclerose/patologia , Estudos de Casos e Controles , Movimento Celular , Proliferação de Células , Células Cultivadas , Modelos Animais de Doenças , Células Endoteliais/patologia , Feminino , Humanos , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout para ApoE , Pessoa de Meia-Idade , Proteína Quinase 7 Ativada por Mitógeno/metabolismo , Músculo Liso Vascular/patologia , Miócitos de Músculo Liso/patologia , Permeabilidade , Fosforilação , Prognóstico , Proteômica , Medição de Risco , Fatores de Risco , Transdução de Sinais , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Proteínas rap de Ligação ao GTPRESUMO
White adipose tissue (WAT) is involved in long-term energy storage and represents 10-15% of total body weight in healthy humans. WAT secretes many peptides (adipokines), hormones and steroids involved in its homeostatic role, especially in carbohydrate-lipid metabolism regulation. Recently, adipocyte-derived extracellular vesicles (AdEVs) have been highlighted as important actors of intercellular communication that participate in metabolic responses to control energy flux and immune response. In this review, we focus on the role of AdEVs in the cross-talks between the different cellular types composing WAT with regard to their contribution to WAT homeostasis and metabolic complications development. We also discuss the AdEV cargoes (proteins, lipids, RNAs) which may explain AdEV's biological effects and demonstrate that, in terms of proteins, AdEV has a very specific signature. Finally, we list and suggest potential therapeutic strategies to modulate AdEV release and composition in order to reduce their deleterious effects during the development of metabolic complications associated with obesity.
Assuntos
Adipócitos/metabolismo , Tecido Adiposo Branco/metabolismo , Comunicação Celular , Vesículas Extracelulares/metabolismo , Obesidade/metabolismo , Obesidade/terapia , Adipócitos/patologia , Adipocinas/metabolismo , Tecido Adiposo Branco/patologia , Animais , Vesículas Extracelulares/patologia , Humanos , Obesidade/patologiaRESUMO
Lipin-1 is a Mg2+-dependent phosphatidic acid phosphatase (PAP) that in mice is necessary for normal glycerolipid biosynthesis, controlling adipocyte metabolism, and adipogenic differentiation. Mice carrying inactivating mutations in the Lpin1 gene display the characteristic features of human familial lipodystrophy. Very little is known about the roles of lipin-1 in human adipocyte physiology. Apparently, fat distribution and weight is normal in humans carrying LPIN1 inactivating mutations, but a detailed analysis of adipose tissue appearance and functions in these patients has not been available so far. In this study, we performed a systematic histopathological, biochemical, and gene expression analysis of adipose tissue biopsies from human patients harboring LPIN1 biallelic inactivating mutations and affected by recurrent episodes of severe rhabdomyolysis. We also explored the adipogenic differentiation potential of human mesenchymal cell populations derived from lipin-1 defective patients. White adipose tissue from human LPIN1 mutant patients displayed a dramatic decrease in lipin-1 protein levels and PAP activity, with a concomitant moderate reduction of adipocyte size. Nevertheless, the adipose tissue develops without obvious histological signs of lipodystrophy and with normal qualitative composition of storage lipids. The increased expression of key adipogenic determinants such as SREBP1, PPARG, and PGC1A shows that specific compensatory phenomena can be activated in vivo in human adipocytes with deficiency of functional lipin-1.
Assuntos
Adipócitos/metabolismo , Tecido Adiposo Branco/metabolismo , Mutação , Fosfatidato Fosfatase/genética , Rabdomiólise/genética , Adipócitos/citologia , Tecido Adiposo Branco/citologia , Adolescente , Alelos , Distribuição da Gordura Corporal , Peso Corporal , Estudos de Casos e Controles , Diferenciação Celular , Criança , Pré-Escolar , Feminino , Regulação da Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , PPAR gama/genética , PPAR gama/metabolismo , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/genética , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Fosfatidato Fosfatase/deficiência , Rabdomiólise/metabolismo , Rabdomiólise/patologia , Proteína de Ligação a Elemento Regulador de Esterol 1/genética , Proteína de Ligação a Elemento Regulador de Esterol 1/metabolismoRESUMO
Adipose extracellular vesicles (AdEVs) transport lipids that could participate in the development of obesity-related metabolic dysfunctions. This study aims to define mouse AdEV lipid signature by a targeted LC-MS/MS approach in either healthy or obesity context. Distinct clustering of AdEV and visceral adipose tissue (VAT) lipidomes by principal component analysis reveals specific AdEV lipid sorting when compared with secreting VAT. Comprehensive analysis identifies enrichment of ceramides, sphingomyelins, and phosphatidylglycerols species in AdEVs compared with source VAT whose lipid content closely relates to the obesity status and is influenced by the diet. Obesity moreover impacts AdEV lipidome, mirroring lipid alterations retrieved in plasma and VAT. Overall, our study identifies specific lipid fingerprints for plasma, VAT, and AdEVs that are informative of the metabolic status. Lipid species enriched in AdEVs in the obesity context may constitute biomarker candidates or mediators of the obesity-associated metabolic dysfunctions.
Assuntos
Vesículas Extracelulares , Lipidômica , Animais , Camundongos , Cromatografia Líquida , Espectrometria de Massas em Tandem , Obesidade/metabolismo , Esfingomielinas/metabolismo , Vesículas Extracelulares/metabolismoRESUMO
Recent evidence supporting that adipose tissue (AT)-derived extracellular vesicles (EVs) carry an important part of the AT secretome led us to characterize the EV-adipokine profile. In addition to evidencing a high AT-derived EV secretion ability that is further increased by obesity, we identify enrichment of oligomeric forms of adiponectin in small EVs (sEVs). This adipokine is mainly distributed at the EV external surface as a result of nonspecific adsorption of soluble adiponectin. EVs also constitute stable conveyors of adiponectin in the blood circulation. Adiponectin-enriched sEVs display in vitro insulin-sensitizing effects by binding to regular adiponectin receptors. Adoptive transfer of adiponectin-enriched sEVs in high-fat-diet-fed mice prevents animals from gaining weight and ameliorated insulin resistance and tissue inflammation, with major effects observed in the AT and liver. Our results therefore provide information regarding adiponectin-related metabolic responses by highlighting EVs as delivery platforms of metabolically active forms of adiponectin molecules.
RESUMO
Nutrient excess induces mitochondrial dysfunction, which participates in obesity-related complications. Obesity also associates with high cardiac oxidative stress, which contributes to myocardial dysfunction. Crewe et al. recently evidenced the pivotal role of adipocyte-derived extracellular vesicles (EVs) in cardiac oxidative stress responses and revealed their unexpected protective effect against ischemia/reperfusion injury.
Assuntos
Vesículas Extracelulares , Mitocôndrias , Adipócitos/metabolismo , Animais , Modelos Animais de Doenças , Vesículas Extracelulares/metabolismo , Humanos , Obesidade/metabolismoRESUMO
Lipodystrophies belong to the heterogenous group of syndromes in which the primary defect is a generalized or partial absence of adipose tissue, which may be congenital or acquired in origin. Lipodystrophy should be considered in patients manifesting the combination of insulin resistance (with or without overt diabetes), dyslipidemia and fatty liver. Lipodystrophies are classified according to the etiology of the disease (genetic or acquired) and to the anatomical distribution of adipose tissue (generalized or partial). The mechanism of adipose tissue loss is specific to each syndrome, depending on the biological function of the mutated gene. Mice models, together with cellular studies have permitted clarification of the mechanisms by which human mutations deeply compromise adipocyte homeostasis. In addition, rodent models have proven to be crucial in deciphering the cardiometabolic consequences of the lack of adipose tissue such as NAFLD, muscle insulin resistance and cardiomyopathy. More precisely, tissue-specific transgenic and knockout mice have brought new tools to distinguish phenotypic traits that are the consequences of lipodystrophy from those that are cell-autonomous. In this review, we discuss the mice models of lipodystrophy including those of inherited human syndromes of generalized and partial lipodystrophy. We present how these models have demonstrated the central role of white adipose tissue in energetic homeostasis in general, including insulin sensitivity and lipid handling in particular. We underscore the differences reported with the human phenotype and discuss the limit of rodent models in recapitulating adipose tissue primary default. Finally, we present how these mice models have highlighted the function of the causative-genes and brought new insights into the pathophysiology of the cardiometabolic complications associated with lipodystrophy.
Assuntos
Doenças Cardiovasculares , Resistência à Insulina , Lipodistrofia , Hepatopatia Gordurosa não Alcoólica , Tecido Adiposo , Animais , Doenças Cardiovasculares/complicações , Modelos Animais de Doenças , Humanos , Resistência à Insulina/genética , Lipodistrofia/genética , Camundongos , Camundongos Knockout , Hepatopatia Gordurosa não Alcoólica/complicações , SíndromeRESUMO
Deficiency of the endoplasmic reticulum (ER) protein seipin results in generalized lipodystrophy by incompletely understood mechanisms. Here, we report mitochondrial abnormalities in seipin-deficient patient cells. A subset of seipin is enriched at ER-mitochondria contact sites (MAMs) in human and mouse cells and localizes in the vicinity of calcium regulators SERCA2, IP3R, and VDAC. Seipin association with MAM calcium regulators is stimulated by fasting-like stimuli, while seipin association with lipid droplets is promoted by lipid loading. Acute seipin removal does not alter ER calcium stores but leads to defective mitochondrial calcium import accompanied by a widespread reduction in Krebs cycle metabolites and ATP levels. In mice, inducible seipin deletion leads to mitochondrial dysfunctions preceding the development of metabolic complications. Together, these data suggest that seipin controls mitochondrial energy metabolism by regulating mitochondrial calcium influx at MAMs. In seipin-deficient adipose tissue, reduced ATP production compromises adipocyte properties, contributing to lipodystrophy pathogenesis.
Assuntos
Adipócitos/metabolismo , Subunidades gama da Proteína de Ligação ao GTP/metabolismo , Mitocôndrias/metabolismo , Tecido Adiposo/metabolismo , Animais , Cálcio/metabolismo , Linhagem Celular , Retículo Endoplasmático/metabolismo , Estresse do Retículo Endoplasmático , Metabolismo Energético/fisiologia , Subunidades gama da Proteína de Ligação ao GTP/deficiência , Subunidades gama da Proteína de Ligação ao GTP/fisiologia , Humanos , Gotículas Lipídicas/metabolismo , Metabolismo dos Lipídeos/fisiologia , Lipídeos/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BLRESUMO
The properties of cholesterol-dependent domains (lipid rafts) in cell membranes have been controversial. Because integrin-mediated cell adhesion and caveolin both regulate trafficking of raft components, we investigated the effects of adhesion and caveolin on membrane order. The fluorescent probe Laurdan and two-photon microscopy revealed that focal adhesions are highly ordered; in fact, they are more ordered than caveolae or domains that stain with cholera toxin subunit B (CtxB). Membrane order at focal adhesion depends partly on phosphorylation of caveolin1 at Tyr14, which localizes to focal adhesions. Detachment of cells from the substratum triggers a rapid, caveolin-independent decrease in membrane order, followed by a slower, caveolin-dependent decrease that correlates with internalization of CtxB-stained domains. Endocytosed CtxB domains also become more fluid. Thus, membrane order is highly dependent on caveolae and focal adhesions. These results show that lipid raft properties are conferred by assembly of specific protein complexes. The ordered state within focal adhesions may have important consequences for signaling at these sites.
Assuntos
Membrana Celular/metabolismo , Adesões Focais/metabolismo , Integrinas/metabolismo , Fluidez de Membrana , Animais , Cavéolas/química , Cavéolas/metabolismo , Caveolina 1/genética , Caveolina 1/metabolismo , Adesão Celular , Membrana Celular/química , Células Cultivadas , Células Endoteliais/química , Células Endoteliais/metabolismo , Fibroblastos/metabolismo , Adesões Focais/química , Lipídeos de Membrana/análise , Camundongos , Camundongos Knockout , Mutação , Suínos , TransfecçãoRESUMO
Extracellular vesicles (EVs) correspond to a heterogeneous set of membrane nanovesicles secreted in the extracellular medium and circulating in the various fluids of the body. These EVs convey biological material (proteins, lipids, nucleic acids) that they can transfer to target cells/tissues thus modulating their response and/or phenotype. The metabolic dysfunctions characterizing metabolic diseases associated with obesity are associated with changes in circulating EV concentrations as well as alterations in their content. The growing interest in EVs as new vectors of intercellular communication has led to question about their role in the development of metabolic complications. In this review, we will discuss the literature on circulating EVs as potential markers of metabolic diseases and then detail inter-organ dialogue based on this EV trafficking underlying the development of related obesity. Finally, we will discuss future avenues of research that will help to better understand the link between EVs and metabolic diseases.
TITLE: Vésicules extracellulaires et maladies métaboliques - Des liaisons dangereuses. ABSTRACT: Les vésicules extracellulaires (VE) correspondent à un ensemble hétérogène de nanovésicules membranaires sécrétées dans le milieu extracellulaire et circulant dans les différents fluides de l'organisme. Ces VE véhiculent du matériel biologique (protéines, lipides, acides nucléiques) qu'elles peuvent transférer à des cellules/tissus cibles, modulant ainsi leur réponse et/ou leur phénotype. Les dysfonctions caractérisant les maladies métaboliques liées à l'obésité sont associées à des modifications des concentrations circulantes de VE ainsi qu'à des altérations de leur contenu. L'intérêt grandissant porté aux VE comme nouveaux vecteurs de communication intercellulaire a conduit à s'interroger sur leur rôle dans le développement des complications métaboliques. Dans cette synthèse, nous résumerons la littérature portant sur les VE circulantes comme potentiels marqueurs des maladies métaboliques. Nous détaillerons ensuite le dialogue vésiculaire inter-organes responsable du développement des complications associées à l'obésité. Enfin, nous discuterons les futures pistes de recherche qui contribueront à mieux appréhender le lien entre VE et maladies métaboliques.
Assuntos
Vesículas Extracelulares , Doenças Metabólicas , Humanos , Obesidade/complicaçõesRESUMO
As the largest human energy reservoir, adipocytes drive an intense dialog with other cells/organs throughout the body to regulate the size of adipose tissue and to communicate with other metabolic tissues and the brain to regulate energy supply. Adipokines have long been described as mediators of this crosstalk, participating in obesity-associated complications. Recently, adipocyte-derived extracellular vesicles (Ad-EVs) have emerged as new key actors in this communication due to their powerful capacity to convey complex messages between cells. Ad-EVs convey specific subpopulations of RNA, proteins, and lipids from their parental cells, and can transfer these cargoes into various recipient cells, modulating their metabolism and cell cycle. In healthy individuals, Ad-EVs actively participate in adipose tissue remodeling to compensate energy supply variations by exchanging information between adipocytes or stroma-vascular cells, including immune cells. Besides this, recent evidence points out that Ad-EV secretion and composition from dysfunctional adipocytes are strongly impacted within adipose tissue where they modulate local intercellular communication, contributing to inflammation, fibrosis, abnormal angiogenesis, and at distance with other cells/tissues intrinsically linked to fat (muscle, hepatocytes and even cancer cells). Additionally, some data even suggests that Ad-EVs might have a systemic action. In this review, we will describe the particular properties of Ad-EVs and their involvement in health and diseases, with a particular focus on metabolic and cardiovascular diseases as well as cancer.
RESUMO
BACKGROUND: Metabolic syndrome (MetS) is characterized by a cluster of interconnected risk factors -hyperglycemia, dyslipidemia, hypertension and obesity- leading to an increased risk of cardiovascular events. Small extracellular vesicles (sEVs) can be considered as new biomarkers of different pathologies, and they are involved in intercellular communication. Here, we hypothesize that sEVs are implicated in MetS-associated endothelial dysfunction. METHODS: Circulating sEVs of non-MetS (nMetS) subjects and MetS patients were isolated from plasma and characterized. Thereafter, sEV effects on endothelial function were analyzed by measuring nitric oxide (NO) and reactive oxygen species (ROS) production, and mitochondrial dynamic proteins on human endothelial aortic cells (HAoECs). RESULTS: Circulating levels of sEVs positively correlated with anthropometric and biochemical parameters including visceral obesity, glycaemia, insulinemia, and dyslipidemia. Treatment of HAoECs with sEVs from MetS patients decreased NO production through the inhibition of the endothelial NO-synthase activity. Injection of MetS-sEVs into mice impaired endothelium-dependent relaxation induced by acetylcholine. Furthermore, MetS-sEVs increased DHE and MitoSox-associated fluorescence in HAoECs, reflecting enhanced cytosolic and mitochondrial ROS production which was not associated with mitochondrial biogenesis or dynamic changes. MetS patients displayed elevated circulating levels of LPS in plasma, and, at least in part, it was associated to circulating sEVs. Pharmacological inhibition and down-regulation of TLR4, as well as sEV-carried LPS neutralization, results in a substantial decrease of ROS production induced by MetS-sEVs. CONCLUSION: These results evidence sEVs from MetS patients as potential new biomarkers for this syndrome, and TLR4 pathway activation by sEVs provides a link between the endothelial dysfunction and metabolic disturbances described in MetS.
Assuntos
Endotélio Vascular/patologia , Vesículas Extracelulares/metabolismo , Lipopolissacarídeos/metabolismo , Síndrome Metabólica/metabolismo , Receptor 4 Toll-Like/metabolismo , Animais , Células Cultivadas , Estudos de Coortes , Citosol/metabolismo , Feminino , Humanos , Masculino , Camundongos , Pessoa de Meia-Idade , Mitocôndrias/metabolismo , Óxido Nítrico/metabolismo , Biogênese de Organelas , Estresse Oxidativo , Espécies Reativas de Oxigênio/metabolismoRESUMO
OBJECTIVE: Astrocytes are glial cells proposed as the main Sonic hedgehog (Shh)-responsive cells in the adult brain. Their roles in mediating Shh functions are still poorly understood. In the hypothalamus, astrocytes support neuronal circuits implicated in the regulation of energy metabolism. In this study, we investigated the impact of genetic activation of Shh signaling on hypothalamic astrocytes and characterized its effects on energy metabolism. METHODS: We analyzed the distribution of gene transcripts of the Shh pathway (Ptc, Gli1, Gli2, and Gli3) in astrocytes using single molecule fluorescence in situ hybridization combined with immunohistofluorescence of Shh peptides by Western blotting in the adult mouse hypothalamus. Based on the metabolic phenotype, we characterized Glast-CreERT2-YFP-Ptc-/- (YFP-Ptc-/-) mice and their controls over time and under a high-fat diet (HFD) to investigate the potential effects of conditional astrocytic deletion of the Shh receptor Patched (Ptc) on metabolic efficiency, insulin sensitivity, and systemic glucose metabolism. Molecular and biochemical assays were used to analyze the alteration of key pathways modulating energy metabolism, insulin sensitivity, glucose uptake, and inflammation. Primary astrocyte cultures were used to evaluate a potential role of Shh signaling in astrocytic glucose uptake. RESULTS: Shh peptides were the highest in the hypothalamic extracts of adult mice and a large population of hypothalamic astrocytes expressed Ptc and Gli1-3 mRNAs. Characterization of Shh signaling after conditional Ptc deletion in the YFP-Ptc-/- mice revealed heterogeneity in hypothalamic astrocyte populations. Interestingly, activation of Shh signaling in Glast+ astrocytes enhanced insulin responsiveness as evidenced by glucose and insulin tolerance tests. This effect was maintained over time and associated with lower blood insulin levels and also observed under a HFD. The YFP-Ptc-/- mice exhibited a lean phenotype with the absence of body weight gain and a marked reduction of white and brown adipose tissues accompanied by increased whole-body fatty acid oxidation. In contrast, food intake, locomotor activity, and body temperature were not altered. At the cellular level, Ptc deletion did not affect glucose uptake in primary astrocyte cultures. In the hypothalamus, activation of the astrocytic Shh pathway was associated with the upregulation of transcripts coding for the insulin receptor and liver kinase B1 (LKB1) after 4 weeks and the glucose transporter GLUT-4 after 32 weeks. CONCLUSIONS: Here, we define hypothalamic Shh action on astrocytes as a novel master regulator of energy metabolism. In the hypothalamus, astrocytic Shh signaling could be critically involved in preventing both aging- and obesity-related metabolic disorders.
Assuntos
Astrócitos/metabolismo , Glucose/metabolismo , Proteínas Hedgehog/metabolismo , Receptores Patched/metabolismo , Envelhecimento , Animais , Astrócitos/patologia , Metabolismo Energético/genética , Células HEK293 , Proteínas Hedgehog/genética , Humanos , Hipotálamo/metabolismo , Hipotálamo/patologia , Hibridização in Situ Fluorescente , Camundongos , Camundongos Endogâmicos C57BL , Células NIH 3T3 , Neurônios/metabolismo , Obesidade , Receptores Patched/deficiência , Receptores Patched/genética , Transdução de Sinais , Ativação TranscricionalRESUMO
Caveolins form plasmalemnal invaginated caveolae. They also locate around intracellular lipid droplets but their role in this location remains unclear. By studying primary adipocytes that highly express caveolin-1, we characterized the impact of caveolin-1 deficiency on lipid droplet proteome and lipidome. We identified several missing proteins on the lipid droplet surface of caveolin-deficient adipocytes and showed that the caveolin-1 lipid droplet pool is organized as multi-protein complexes containing cavin-1, with similar dynamics as those found in caveolae. On the lipid side, caveolin deficiency did not qualitatively alter neutral lipids in lipid droplet, but significantly reduced the relative abundance of surface phospholipid species: phosphatidylserine and lysophospholipids. Caveolin-deficient adipocytes can form only small lipid droplets, suggesting that the caveolin-lipid droplet pool might be involved in lipid droplet size regulation. Accordingly, we show that caveolin-1 concentration on adipocyte lipid droplets positively correlated with lipid droplet size in obese rodent models and human adipocytes. Moreover, rescue experiments by caveolin- green fluorescent protein in caveolin-deficient cells exposed to fatty acid overload demonstrated that caveolin-coated lipid droplets were able to grow larger than caveolin-devoid lipid droplets. Altogether, these data demonstrate that the lipid droplet-caveolin pool impacts on phospholipid and protein surface composition of lipid droplets and suggest a functional role on lipid droplet expandability.
Assuntos
Adipócitos/metabolismo , Caveolina 1/deficiência , Fosfolipídeos/química , Fosfolipídeos/metabolismo , Células 3T3-L1 , Adipócitos/citologia , Animais , Caveolina 1/metabolismo , Humanos , Camundongos , Proteoma/química , Proteoma/metabolismo , RatosRESUMO
Caveolins are primarily known as the main constituents of the protein coat of caveolae invaginations at the plasma membrane. They have also been found at the surface of intracellular lipid droplets but their function in this lipid storage organelle remains poorly understood. This paper reviews recent studies in adipocytes, the specialized cell type for fatty acid storage, which suggest a role for caveolins in the formation, maintenance or mobilization of lipid droplet stores. These new functions emerged from studies of fat cells in which caveolin expression was invalidated, highlighting the metabolic phenotype of caveolin-deficient mice or human patients who develop progressive lipoatrophy.
Assuntos
Adipócitos/metabolismo , Caveolina 1/deficiência , Organelas/metabolismo , Animais , Caveolina 1/genética , Genótipo , Humanos , Lipogênese/genética , Lipólise/genética , Camundongos , Camundongos Knockout , FenótipoRESUMO
Repetitive complete or incomplete pharyngeal collapses are leading to chronic intermittent hypoxia (CIH), a hallmark feature of obstructive sleep apnea (OSA) syndrome responsible for many metabolic disorders. In humans, an association between OSA and insulin resistance has been found independently of the degree of obesity. Based on our previous work showing that hypoxia applied to adipocytes led to cellular insulin resistance associated with caveolae flattening, we have investigated the effects of CIH on caveolae structuration in adipose tissue. Original exploratory experiences demonstrate that 6 weeks-exposure of lean mice to CIH is characterized by systemic insulin resistance and translates into adipocyte insulin signaling alterations. Chronic intermittent hypoxia also induces caveolae disassembly in white adipose tissue (WAT) illustrated by reduced plasma membrane caveolae density and enlarged caveolae width, concomitantly to WAT insulin resistance state. We show that CIH downregulates caveolar gene and protein expressions, including cavin-1, cavin-2, and EHD2, underlying molecular mechanisms responsible for such caveolae flattening. Altogether, we provide evidences for adipose tissue caveolae disassembly following CIH exposure, likely linked to cavin protein downregulation. This event may constitute the molecular basis of insulin resistance development in OSA patients.
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Lipid rafts defined as cholesterol- and sphingomyelin-rich domains have been isolated from different cell types that vary greatly in their lipid profiles. Here, we investigated the contribution of the structural protein caveolin-1 (Cav1) to the overall lipid composition and domain abundance in mouse embryonic fibroblasts (MEFs) from wild-type (WT) or Cav1-deficient (Cav1(-/-)) animals. Our findings show that Cav1 expression had no effect on free (membrane-associated) cholesterol levels. However, Cav1(-/-)-deficient cells did have a higher proportion of sphingomyelin, decreased abundance of unsaturated phospholipids, and a trend toward shorter fatty acid chains in phosphatidylcholine. We isolated detergent-resistant membranes (DRMs), nondetergent raft domains (NDR), and cholesterol oxidase (CO)-sensitive domains and assessed the abundance of ordered domains in intact cells using the fluorescent dye Laurdan. Despite differences in phospholipid composition, we found that cholesterol levels in DRMs, NDR, and CO-sensitive domains were similar in both cell types. The data suggest that Cav1 is not required to target cholesterol to lipid rafts and that CO does not specifically oxidize caveolar cholesterol. In contrast, the abundance of ordered domains in adherent cells is reduced in Cav1(-/-) compared with WT MEFs, suggesting that cell architecture is critical in maintaining Cav1-induced lipid rafts.
Assuntos
Caveolina 1/fisiologia , Microdomínios da Membrana/química , Fosfolipídeos/análise , Animais , Cavéolas , Caveolina 1/análise , Caveolina 1/deficiência , Caveolina 2/análise , Estruturas da Membrana Celular/química , Células Cultivadas , Colesterol/análise , Ésteres do Colesterol/análise , Colesterol Oxidase/metabolismo , Ácidos Graxos/análise , Feminino , Masculino , Camundongos , Camundongos Knockout , Fosfatidilcolinas/análise , Proteínas Proto-Oncogênicas c-yes/metabolismo , Esfingomielinas/análiseRESUMO
Elevated ceramide concentrations in adipocytes and skeletal muscle impair PKB (protein kinase B; also known as Akt)-directed insulin signalling to key hormonal end points. An important feature of this inhibition involves the ceramide-induced activation of atypical PKCzeta (protein kinase C-zeta), which associates with and negatively regulates PKB. In the present study, we demonstrate that this inhibition is critically dependent on the targeting and subsequent retention of PKCzeta-PKB within CEM (caveolin-enriched microdomains), which is facilitated by kinase interactions with caveolin. Ceramide also recruits PTEN (phosphatase and tensin homologue detected on chromosome 10), a 3'-phosphoinositide phosphatase, thereby creating a repressive membrane microenvironment from which PKB cannot signal. Disrupting the structural integrity of caveolae by cholesterol depletion prevented caveolar targeting of PKCzeta and PKB and suppressed kinase-caveolin association, but, importantly, also ameliorated ceramide-induced inhibition of PKB. Consistent with this, adipocytes from caveolin-1-/- mice, which lack functional caveolae, exhibit greater resistance to ceramide compared with caveolin-1+/+ adipocytes. We conclude that the recruitment and retention of PKB within CEM contribute significantly to ceramide-induced inhibition of PKB-directed signalling.
Assuntos
Ceramidas/farmacologia , Proteína Quinase C/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Células 3T3 , Adipócitos/enzimologia , Animais , Caveolina 1/deficiência , Colesterol/metabolismo , Ativação Enzimática , Humanos , Insulina/fisiologia , Camundongos , Camundongos Knockout , Músculo Esquelético/enzimologia , Transdução de SinaisRESUMO
Despite long traditional utilization and some reports on the antihyperglycemic and antihyperlipidemic action of Cassia siamea, the mechanisms involved have not been investigated yet. Thus, the objective of the present study was to investigate whether and how oral administration of the ethanolic extract of Cassia siamea Lam leaves (LECS) improves glucose and insulin homoeostasis, liver damage, and endothelial dysfunction in an experimental model of type 2 diabetes, the leptin-deficient ob/ob mice. Oxidative stress and protein expression of insulin-dependent and insulin -independent signaling pathways were studied. Obese ( ob/ob) vs. control (ob/+) mice were treated daily with intragastric administration of either vehicle or LECS (200 mg/kg, per day) for 4 weeks. Fasting blood glucose, body weight, food intake, glucose and insulin tolerance, oxidative stress, and liver damage as well as vascular complications with respect to endothelial dysfunction were examined. Administration of LECS in obese mice significantly reduced blood glucose and insulin levels, improved glucose tolerance and insulin sensitivity, and restored the increase of circulating AST and ALT without modification of body weight and food intake. These effects were associated with increased activity of both insulin and AMPK pathways in the liver and skeletal muscles. Of particular interest, administration of LECS in obese mice completely prevented the endothelial dysfunction resulting from an increased NO· and decreased reactive oxygen species (ROS) production in the aorta. Altogether, oral administration of LECS remarkably attenuates features of type 2 diabetes on glucose, hepatic inflammation, insulin resistance, endothelial function, and vascular oxidative stress, being as most of these effects are related to insulin-dependent and insulin-independent mechanisms. Therefore, this study points for the therapeutic potential of Cassia siamea in correcting both metabolic and vascular alterations linked to type 2 diabetes.