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1.
Am J Physiol Endocrinol Metab ; 325(6): E682-E687, 2023 12 01.
Artículo en Inglés | MEDLINE | ID: mdl-37877798

RESUMEN

Adipocyte dysfunction is a hallmark of systemic insulin resistance. Insulin-responsive glucose transporter 4 (GLUT4) is downregulated in the insulin resistant state, and cellular insulin responsiveness varies depending on fat depot origin and degree of adipose expansion. Here, we have resolved factors limiting cellular insulin responsiveness, by examining adipocyte function and traits related to glucose transport at the cellular level. Subcutaneous (inguinal) and visceral (epididymal) adipocytes were isolated from C57BL/6J mice fed either chow or high-fat diet. Cell size was determined using coulter counter method, glucose uptake and cytosolic volume were assessed using glucose tracer assays. Total and GLUT4 protein content expressions were determined by Western blot. We found that basal glucose uptake per cell was preserved independent of diet or fat depot origin. Insulin-stimulated glucose uptake per cell was sustained in visceral adipocytes but decreased with adipose expansion in subcutaneous adipocytes. In parallel, the cytosolic space and total protein increased proportionally to total cellular volumetric expansion in visceral, but not in subcutaneous, adipocytes, whereas GLUT4 content decreased exclusively in expanding subcutaneous adipocytes. Together, these data support the existence of distinct phenotypic adipocyte traits that could limit cellular insulin responsiveness. Potentially, these characteristics account for fat depot-specific differences related to glucose transport capacity.NEW & NOTEWORTHY This work illustrates that adipocyte characteristics related to fat depot origin rather than adipocyte size per se limit cellular insulin responsiveness and glucose uptake in male C57BL/6J mice. These findings contribute to the overall understanding of factors limiting adipocyte function and how adipose progression affects insulin response and glucose transport capacity differently in diverse fat depots. Future studies examining whether the proposed characteristics hold true in adipocytes derived from female mice or human origin are needed.


Asunto(s)
Resistencia a la Insulina , Insulina , Humanos , Masculino , Femenino , Ratones , Animales , Insulina/farmacología , Insulina/metabolismo , Ratones Endogámicos C57BL , Adipocitos/metabolismo , Glucosa/metabolismo , Resistencia a la Insulina/fisiología , Obesidad/metabolismo , Tejido Adiposo/metabolismo
2.
Cytokine ; 161: 156057, 2023 01.
Artículo en Inglés | MEDLINE | ID: mdl-36208532

RESUMEN

During inflammation, cellular glucose uptake and glycolysis are upregulated to meet an increased energy demand. For example, keratinocyte glycolysis is essential for progression of psoriasis. Therefore, understanding the regulation of glucose metabolism in keratinocytes is of importance. Here, we show that the pro-inflammatory cytokines IFNγ and TNF together rapidly induce glucose uptake, glycolysis, and glycolytic capacity in cultured keratinocytes. Furthermore, we found that acute IFNγ and TNF stimulation induces glucose transporter 4 (GLUT4) translocation to the plasma membrane and engages AMPK-dependent intracellular signaling. Together, these findings suggest acute cytokine-induced glucose metabolism in keratinocytes could contribute to inflammation in psoriatic disease, and that GLUT4 is involved in these processes.


Asunto(s)
Citocinas , Queratinocitos , Humanos , Citocinas/metabolismo , Queratinocitos/metabolismo , Glucosa/metabolismo , Glucólisis , Inflamación/metabolismo
3.
Cell Mol Life Sci ; 79(8): 459, 2022 Aug 01.
Artículo en Inglés | MEDLINE | ID: mdl-35913515

RESUMEN

Differentiation of smooth muscle cells (SMCs) depends on serum response factor (SRF) and its co-activator myocardin (MYOCD). The role of MYOCD for the SMC program of gene transcription is well established. In contrast, the role of MYOCD in control of SMC-specific alternative exon usage, including exon splicing, has not been explored. In the current work we identified four splicing factors (MBNL1, RBPMS, RBPMS2, and RBFOX2) that correlate with MYOCD across human SMC tissues. Forced expression of MYOCD family members in human coronary artery SMCs in vitro upregulated expression of these splicing factors. For global profiling of transcript diversity, we performed RNA-sequencing after MYOCD transduction. We analyzed alternative transcripts with three different methods. Exon-based analysis identified 1637 features with differential exon usage. For example, usage of 3´ exons in MYLK that encode telokin increased relative to 5´ exons, as did the 17 kDa telokin to 130 kDa MYLK protein ratio. Dedicated event-based analysis identified 239 MYOCD-driven splicing events. Events involving MBNL1, MCAM, and ACTN1 were among the most prominent, and this was confirmed using variant-specific PCR analyses. In support of a role for RBPMS and RBFOX2 in MYOCD-driven splicing we found enrichment of their binding motifs around differentially spliced exons. Moreover, knockdown of either RBPMS or RBFOX2 antagonized splicing events stimulated by MYOCD, including those involving ACTN1, VCL, and MBNL1. Supporting an in vivo role of MYOCD-SRF-driven splicing, we demonstrate altered Rbpms expression and splicing in inducible and SMC-specific Srf knockout mice. We conclude that MYOCD-SRF, in part via RBPMS and RBFOX2, induce a program of differential exon usage and alternative splicing as part of the broader program of SMC differentiation.


Asunto(s)
Empalme Alternativo , Miocitos del Músculo Liso , Empalme Alternativo/genética , Animales , Diferenciación Celular/genética , Exones/genética , Humanos , Ratones , Miocitos del Músculo Liso/metabolismo , Proteínas Nucleares , Factores de Empalme de ARN/genética , Factores de Empalme de ARN/metabolismo , Proteínas Represoras/metabolismo , Transactivadores
4.
Lipids Health Dis ; 21(1): 68, 2022 Aug 04.
Artículo en Inglés | MEDLINE | ID: mdl-35927727

RESUMEN

PURPOSE: Previous studies have shown that at a similar body mass index, Middle Eastern immigrants are more insulin resistant and at higher risk for type 2 diabetes (T2D) than native Europeans. Insulin resistance is strongly associated with disturbed fat metabolism and cardiovascular disease (CVD). However, fat metabolism is poorly investigated comparing Middle Eastern and European ethnicities. METHODS: This observational study included 26 Iraqi and 16 Swedish-born men without T2D or clinical risk factors for CVD. An oral fat tolerance test (OFTT) was performed, where plasma triglycerides (p-TG) were measured for 6 h. mRNA expression and adipocyte size were measured in subcutaneous adipose tissue biopsies collected prior to OFTT, and magnetic resonance imaging was conducted to assess body fat distribution. RESULTS: The median p-TG accumulation was higher and the clearance slower among Iraqis than Swedes. None of the groups reached their fasting p-TG (Iraqis 1.55 mmol/l; Swedes 0.95 mmol/l) after 6 h (Iraqis p-TG 3.10 mmol/l; Swedes p-TG 1.50 mmol/l). Adipocyte size, mRNA expression, and fat accumulation in the liver, muscle and abdomen were similar in both groups. CONCLUSION: Postprandial p-TG levels rather than fat distribution may reflect early signs of disturbed fat metabolism in Iraqi immigrants without CVD risk factors.


Asunto(s)
Enfermedades Cardiovasculares , Diabetes Mellitus Tipo 2 , Emigrantes e Inmigrantes , Antígenos CD36 , Humanos , Irak , Masculino , Periodo Posprandial , ARN Mensajero , Suecia , Triglicéridos
5.
Int J Mol Sci ; 22(17)2021 Sep 03.
Artículo en Inglés | MEDLINE | ID: mdl-34502470

RESUMEN

Bacterial endotoxin is a potent inflammatory antigen abundant in the human intestine. Endotoxins circulate in the blood at low concentrations in all healthy individuals. Elevated levels of circulatory endotoxins may cause inflammation with the development of chronic disease, either affecting metabolism, neurological disease, or resistance to viral and bacterial infections. The most important endotoxin is LPS, being a superantigen. In this narrative review, the effect of various food components to postprandially elevate circulating LPS and inflammatory markers is described. There is evidence that the intake of food enriched in fat, in particular saturated fat, may elevate LPS and pro-inflammatory markers. This occurs in both normal-weight and obese subjects. In obese subjects, inflammatory markers are already elevated before meal consumption. The importance of food choice for endotoxemia and inflammatory response is discussed.


Asunto(s)
Grasas de la Dieta , Endotoxemia/sangre , Contaminación de Alimentos , Lipopolisacáridos/toxicidad , Obesidad/sangre , Biomarcadores/sangre , Endotoxemia/etiología , Humanos , Inflamación/sangre , Inflamación/etiología
6.
Biochem Biophys Res Commun ; 533(1): 64-69, 2020 11 26.
Artículo en Inglés | MEDLINE | ID: mdl-32921413

RESUMEN

Obesity is the main risk factor behind insulin resistance and type 2 diabetes. Still, the mechanism behind adipocyte dysfunction is not yet resolved. Recently, we reported that rapid actin remodeling correlates with adipose cell size changes after short-term overfeeding. Therefore, we hypothesized that the actin-driven myocardin-related transcription factor (MRTF-A) contributes to impaired mature adipocyte function. Primary human adipocytes were subjected to adenoviral overexpression of MRTF-A or MRTF-B, followed by Western blot analysis and tracer glucose uptake assay. Further, we assessed cell size distribution, insulin response, MRTF-A localization, actin organization and degree of polymerization in adipocytes isolated from Ob/Ob mice. Overexpression of MRTF-A, but not MRTF-B, markedly suppressed PPARγ expression. Further, MRTF-A expression resulted in decreased IRS-1 level, shifted phosphorylation of Akt (pS473/pT308), IRS-1 (pS302) and AS160 (pT642), and lowered insulin-stimulated glucose uptake. Hypertrophic adipocytes from Ob/Ob mice displayed an increased proportion of polymerized actin, and increased nuclear translocation of MRTF-A compared with control (Ob/+). Similar with human adipocytes overexpressing MRTF-A, adipocytes isolated from Ob/Ob mice had reduced expression of IRS-1 and PPARγ, as well as impaired insulin response. Together, these data demonstrate that MRTF-A negatively influences insulin sensitivity and the expression of key targets in fully mature human adipocytes. This suggests that MRTF-A is poised to exert a transcriptional response in hypertrophic adipocytes, contributing to adipocyte dysfunction and insulin resistance.


Asunto(s)
Adipocitos/metabolismo , Resistencia a la Insulina , PPAR gamma/metabolismo , Transactivadores/metabolismo , Animales , Células Cultivadas , Regulación hacia Abajo , Glucosa/metabolismo , Humanos , Insulina/metabolismo , Ratones Obesos , PPAR gamma/genética , Transactivadores/genética , Regulación hacia Arriba
7.
Diabetologia ; 62(7): 1257-1267, 2019 07.
Artículo en Inglés | MEDLINE | ID: mdl-31069401

RESUMEN

AIMS/HYPOTHESIS: Finding new treatment alternatives for individuals with diabetes with severe insulin resistance is highly desired. To identify novel mechanisms that improve glucose uptake in skeletal muscle, independently from insulin levels and signalling, we have explored the therapeutic potential of a short peptide sequence, RG54, derived from apolipoprotein A-I (ApoA-I). METHODS: INS-1E rat clonal beta cells, C2C12 rat muscle myotubes and J774 mouse macrophages were used to study the impact of RG54 peptide on glucose-stimulated insulin secretion, glucose uptake and cholesterol efflux, respectively. GTTs were carried out on diet-induced insulin-resistant and Leprdb diabetic mouse models treated with RG54 peptide, and the impact of RG54 peptide on atherosclerosis was evaluated in Apoe-/- mice. Control mice received ApoA-I protein, liraglutide or NaCl. RESULTS: The synthetic RG54 peptide induced glucose uptake in cultured muscle myotubes by a similar amount as insulin, and also primed pancreatic beta cells for improved glucose-stimulated insulin secretion. The findings were verified in diet-induced insulin-resistant and Leprdb diabetic mice, jointly confirming the physiological effect. The RG54 peptide also efficiently catalysed cholesterol efflux from macrophages and prevented the formation of atherosclerotic plaques in Apoe-/- mice. CONCLUSIONS/INTERPRETATION: The RG54 peptide exhibits good prospects for providing glucose control and reducing the risk of cardiovascular disease in individuals with severe insulin resistance.


Asunto(s)
Apolipoproteína A-I/química , Aterosclerosis/prevención & control , Glucosa/metabolismo , Péptidos/química , Péptidos/uso terapéutico , Animales , Aterosclerosis/metabolismo , Glucemia/efectos de los fármacos , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/prevención & control , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/prevención & control , Resistencia a la Insulina/fisiología , Masculino , Ratones , Ratones Endogámicos C57BL
8.
J Cell Biochem ; 120(1): 343-356, 2019 01.
Artículo en Inglés | MEDLINE | ID: mdl-30171718

RESUMEN

The mechanism of how patatin-like phospholipase domain-containing protein 3 (PNPLA3) variant M148 is associated with increased risk of development of hepatic steatosis is still debated. Here, we propose a novel role of PNPLA3 as a key player during autophagosome formation in the process of lipophagy. A human hepatocyte cell line, HepG2 cells, expressing recombinant I148 or 148M, was used to study lipophagy under energy deprived conditions, and lipid droplet morphology was investigated using florescence microscopy, image analysis and biochemical assays. Autophagic flux was studied using the golden-standard of LC3-II turnover in combination with the well characterized GFP-RFP-LC3 vector. To discriminate between, perturbed autophagic initiation and lysosome functionality, lysosomes were characterized by Lysotracker staining and LAMP1 protein levels as well as activity and activation of cathepsin B. For validation, human liver biopsies genotyped for I148 and 148M were analyzed for the presence of LC3-II and PNPLA3 on lipid droplets. We show that the M148-PNPLA3 variant is associated with lipid droplets that are resistant to starvation-mediated degradation. M148 expressing hepatocytes reveal decreased autophagic flux and reduced lipophagy. Both I148-PNPLA3 and M148-PNPLA3 colocalize and interact with LC3-II, but the M148-PNPLA3 variant has lower ability to bind LC3-II. Together, our data indicate that PNPLA3 might play an essential role in lipophagy in hepatocytes and furthermore that the M148-PNPLA3 variant appears to display a loss in this activity, leading to decreased lipophagy.


Asunto(s)
Autofagia , Variación Genética , Hepatocitos/metabolismo , Lipasa/genética , Gotas Lipídicas/metabolismo , Proteínas de la Membrana/genética , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Autofagosomas/metabolismo , Biopsia , Catepsina B/metabolismo , Estudios de Cohortes , Genotipo , Células Hep G2 , Humanos , Lipasa/metabolismo , Metabolismo de los Lípidos , Hígado/patología , Proteínas de Membrana de los Lisosomas/metabolismo , Lisosomas/metabolismo , Proteínas de la Membrana/metabolismo , Microscopía Fluorescente , Proteínas Asociadas a Microtúbulos/metabolismo , Enfermedad del Hígado Graso no Alcohólico/patología , Transfección
9.
J Biol Chem ; 292(27): 11206-11217, 2017 07 07.
Artículo en Inglés | MEDLINE | ID: mdl-28495883

RESUMEN

Type 2 diabetes is characterized by insulin resistance, which arises from malfunctions in the intracellular insulin signaling network. Knowledge of the insulin signaling network is fragmented, and because of the complexity of this network, little consensus has emerged for the structure and importance of the different branches of the network. To help overcome this complexity, systems biology mathematical models have been generated for predicting both the activation of the insulin receptor (IR) and the redistribution of glucose transporter 4 (GLUT4) to the plasma membrane. Although the insulin signal transduction between IR and GLUT4 has been thoroughly studied with modeling and time-resolved data in human cells, comparable analyses in cells from commonly used model organisms such as rats and mice are lacking. Here, we combined existing data and models for rat adipocytes with new data collected for the signaling network between IR and GLUT4 to create a model also for their interconnections. To describe all data (>140 data points), the model needed three distinct pathways from IR to GLUT4: (i) via protein kinase B (PKB) and Akt substrate of 160 kDa (AS160), (ii) via an AS160-independent pathway from PKB, and (iii) via an additional pathway from IR, e.g. affecting the membrane constitution. The developed combined model could describe data not used for training the model and was used to generate predictions of the relative contributions of the pathways from IR to translocation of GLUT4. The combined model provides a systems-level understanding of insulin signaling in rat adipocytes, which, when combined with corresponding models for human adipocytes, may contribute to model-based drug development for diabetes.


Asunto(s)
Adipocitos/metabolismo , Transportador de Glucosa de Tipo 4/metabolismo , Receptor de Insulina/metabolismo , Transducción de Señal/fisiología , Adipocitos/citología , Animales , Proteínas Activadoras de GTPasa/genética , Proteínas Activadoras de GTPasa/metabolismo , Transportador de Glucosa de Tipo 4/genética , Transporte de Proteínas/fisiología , Proteínas Proto-Oncogénicas c-akt/genética , Proteínas Proto-Oncogénicas c-akt/metabolismo , Ratas , Receptor de Insulina/genética , Biología de Sistemas/métodos
10.
J Cell Physiol ; 233(9): 7195-7205, 2018 09.
Artículo en Inglés | MEDLINE | ID: mdl-29574754

RESUMEN

Diabetes is a major risk factor for cardiovascular disease and this is in part due to the effects of hyperglycemia on vascular smooth muscle cells. Small non-coding microRNAs are known to control smooth muscle phenotype and arterial contractility and are dysregulated in diabetes. The effect of microRNAs on smooth muscle differentiation is in part mediated by the transcription factor KLF4 but the role of this mechanism in diabetic vascular disease is not fully understood. Herein, we have investigated the importance of hyperglycemia and diabetes for the expression of KLF4 in vascular smooth muscle and the involvement of miRNAs in this regulation. Hyperglycemia down-regulated KLF4 in vascular smooth muscle cells and similar results were found in arteries of diabetic mice and patients. This correlated with a Foxa2-dependent up-regulation of miR-29c, which targeted KLF4 in vascular smooth muscle cells. Importantly, by preventing downregulation of KLF4, the induction of smooth muscle contractile protein markers by glucose was inhibited. In conclusion, miR-29 mediated inhibition of KLF4 in hyperglycemic conditions contributes to increased expression of contractile markers in vascular smooth muscle cells. Further studies are warranted to determine the therapeutic implications of miR-29 inhibition in diabetic vascular disease.


Asunto(s)
Regulación de la Expresión Génica/efectos de los fármacos , Glucosa/farmacología , Factores de Transcripción de Tipo Kruppel/genética , MicroARNs/metabolismo , Músculo Liso Vascular/metabolismo , Anciano , Animales , Biomarcadores/metabolismo , Diabetes Mellitus/genética , Diabetes Mellitus/patología , Factor Nuclear 3-beta del Hepatocito/metabolismo , Humanos , Hiperglucemia/genética , Hiperglucemia/patología , Factor 4 Similar a Kruppel , Factores de Transcripción de Tipo Kruppel/metabolismo , Ratones Endogámicos C57BL , MicroARNs/genética , Contracción Muscular/efectos de los fármacos , Miocitos del Músculo Liso/efectos de los fármacos , Miocitos del Músculo Liso/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo
11.
Am J Physiol Regul Integr Comp Physiol ; 315(2): R284-R295, 2018 08 01.
Artículo en Inglés | MEDLINE | ID: mdl-29641234

RESUMEN

Adipose tissue is necessary to harbor energy. To handle excess energy, adipose tissue expands by increasing adipocyte size (hypertrophy) and number (hyperplasia). Here, we have summarized the different experimental techniques used to study adipocyte cell size and describe adipocyte size in relation to insulin resistance, type 2 diabetes, and diet interventions. Hypertrophic adipocytes have an impaired cellular function, and inherent mechanisms restrict their expansion to protect against cell breakage and subsequent inflammation. Reduction of large fat cells by diet restriction, physical activity, or bariatric surgery therefore is necessary to improve cellular function and health. Small fat cells may also be dysfunctional and unable to expand. The distribution and function of the entire cell size range of fat cells, from small to very large fat cells, are an important but understudied aspect of adipose tissue biology. To prevent dysmetabolism, therapeutic strategies to expand small fat cells, recruit new fat cells, and reduce large fat cells are needed.


Asunto(s)
Adipocitos/patología , Tejido Adiposo/patología , Tamaño de la Célula , Diabetes Mellitus Tipo 2/patología , Obesidad/patología , Adipocitos/metabolismo , Tejido Adiposo/metabolismo , Tejido Adiposo/fisiopatología , Adiposidad , Animales , Cirugía Bariátrica , Restricción Calórica , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/fisiopatología , Diabetes Mellitus Tipo 2/terapia , Metabolismo Energético , Ejercicio Físico , Humanos , Hipertrofia , Resistencia a la Insulina , Obesidad/metabolismo , Obesidad/fisiopatología , Obesidad/terapia
12.
J Pathol ; 241(5): 578-582, 2017 04.
Artículo en Inglés | MEDLINE | ID: mdl-28139834

RESUMEN

Tumours displaying differentiation towards normal fat constitute the most common subgroup of soft tissue neoplasms. A series of such tumours was investigated by whole-exome sequencing followed by targeted ultra-deep sequencing. Eighty per cent of angiolipomas, but not any other tumour type, displayed mutations in the protein kinase D2 (PRKD2) gene, typically in the part encoding the catalytic domain. The absence of other aberrations at the chromosome or RNA level suggests that PRKD2 mutations are critical for angiolipoma development. Consistently, the mutated PRKD2 alleles were present at low (3-15%) frequencies, indicating that only a subset of the tumour cells is affected. Indeed, by sequencing mature fat cells and other cells separately, the former typically showed the highest mutation frequencies. Thus, we hypothesize that altered PRKD2 signalling in the adipocytic cells drives tumourigenesis and, in agreement with its pivotal role in angiogenesis, induces the vessel formation that is characteristic for angiolipoma. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.


Asunto(s)
Angiolipoma/genética , Proteína Quinasa Activada por ADN/genética , Proteínas Nucleares/genética , Neoplasias de los Tejidos Blandos/genética , Adipocitos , Secuencia de Aminoácidos , Angiolipoma/irrigación sanguínea , Angiolipoma/patología , Carcinogénesis , Exoma/genética , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Modelos Moleculares , Mutación , Neovascularización Patológica , Alineación de Secuencia , Análisis de Secuencia de ARN , Transducción de Señal , Neoplasias de los Tejidos Blandos/irrigación sanguínea , Neoplasias de los Tejidos Blandos/patología
13.
Diabetologia ; 60(2): 314-323, 2017 02.
Artículo en Inglés | MEDLINE | ID: mdl-27807598

RESUMEN

AIMS/HYPOTHESIS: Salt-inducible kinases (SIKs) are related to the metabolic regulator AMP-activated protein kinase (AMPK). SIK2 is abundant in adipose tissue. The aims of this study were to investigate the expression of SIKs in relation to human obesity and insulin resistance, and to evaluate whether changes in the expression of SIKs might play a causal role in the development of disturbed glucose uptake in human adipocytes. METHODS: SIK mRNA and protein was determined in human adipose tissue or adipocytes, and correlated to clinical variables. SIK2 and SIK3 expression and phosphorylation were analysed in adipocytes treated with TNF-α. Glucose uptake, GLUT protein levels and localisation, phosphorylation of protein kinase B (PKB/Akt) and the SIK substrate histone deacetylase 4 (HDAC4) were analysed after the SIKs had been silenced using small interfering RNA (siRNA) or inhibited using a pan-SIK-inhibitor (HG-9-91-01). RESULTS: We demonstrate that SIK2 and SIK3 mRNA are downregulated in adipose tissue from obese individuals and that the expression is regulated by weight change. SIK2 is also negatively associated with in vivo insulin resistance (HOMA-IR), independently of BMI and age. Moreover, SIK2 protein levels and specific kinase activity display a negative correlation to BMI in human adipocytes. Furthermore, SIK2 and SIK3 are downregulated by TNF-α in adipocytes. Silencing or inhibiting SIK1-3 in adipocytes results in reduced phosphorylation of HDAC4 and PKB/Akt, less GLUT4 at the plasma membrane, and lower basal and insulin-stimulated glucose uptake in adipocytes. CONCLUSION/INTERPRETATION: This is the first study to describe the expression and function of SIKs in human adipocytes. Our data suggest that SIKs might be protective in the development of obesity-induced insulin resistance, with implications for future treatment strategies.


Asunto(s)
Adipocitos/metabolismo , Tejido Adiposo/metabolismo , Glucosa/metabolismo , Insulina/metabolismo , Obesidad/metabolismo , Proteínas Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Células 3T3-L1 , Adipocitos/efectos de los fármacos , Adulto , Anciano , Animales , Western Blotting , Femenino , Humanos , Resistencia a la Insulina/genética , Resistencia a la Insulina/fisiología , Masculino , Ratones , Persona de Mediana Edad , Fosforilación/efectos de los fármacos , Proteínas Quinasas/genética , Proteínas Serina-Treonina Quinasas/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transducción de Señal/efectos de los fármacos , Transducción de Señal/genética , Factor de Necrosis Tumoral alfa/farmacología
14.
J Cell Sci ; 128(3): 472-86, 2015 Feb 01.
Artículo en Inglés | MEDLINE | ID: mdl-25472719

RESUMEN

Salt-inducible kinase 2 (SIK2) is an AMP-activated protein kinase (AMPK) related kinase abundantly expressed in adipose tissue. Our aim was to identify molecular targets and functions of SIK2 in adipocytes, and to address the role of PKA-mediated phosphorylation of SIK2 on Ser358. Modulation of SIK2 in adipocytes resulted in altered phosphorylation of CREB-regulated transcription co-activator 2 (CRTC2), CRTC3 and class IIa histone deacetylase 4 (HDAC4). Furthermore, CRTC2, CRTC3, HDAC4 and protein phosphatase 2A (PP2A) interacted with SIK2, and the binding of CRTCs and PP2A to wild-type but not Ser358Ala SIK2, was reduced by cAMP elevation. Silencing of SIK2 resulted in reduced GLUT4 (also known as SLC2A4) protein levels, whereas cells treated with CRTC2 or HDAC4 siRNA displayed increased levels of GLUT4. Overexpression or pharmacological inhibition of SIK2 resulted in increased and decreased glucose uptake, respectively. We also describe a SIK2­CRTC2­HDAC4 pathway and its regulation in human adipocytes, strengthening the physiological relevance of our findings. Collectively, we demonstrate that SIK2 acts directly on CRTC2, CRTC3 and HDAC4, and that the cAMP­PKA pathway reduces the interaction of SIK2 with CRTCs and PP2A. Downstream, SIK2 increases GLUT4 levels and glucose uptake in adipocytes.


Asunto(s)
Glucosa/metabolismo , Histona Desacetilasas/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Factores de Transcripción/metabolismo , Células 3T3 , Adipocitos/metabolismo , Tejido Adiposo/metabolismo , Animales , Línea Celular , AMP Cíclico/metabolismo , Transportador de Glucosa de Tipo 4/metabolismo , Células HEK293 , Histona Desacetilasas/genética , Humanos , Ratones , Ratones Noqueados , Fosforilación , Unión Proteica , Proteína Fosfatasa 2/metabolismo , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Proteínas Serina-Treonina Quinasas/genética , Interferencia de ARN , ARN Interferente Pequeño , Ratas , Transducción de Señal , Factores de Transcripción/genética
15.
J Physiol ; 594(17): 4741-52, 2016 09 01.
Artículo en Inglés | MEDLINE | ID: mdl-27060572

RESUMEN

Members of the myocardin family bind to the transcription factor serum response factor (SRF) and act as coactivators controlling genes of relevance for myogenic differentiation and motile function. Binding of SRF to DNA is mediated by genetic elements called CArG boxes, found often but not exclusively in muscle and growth controlling genes. Studies aimed at defining the full spectrum of these CArG elements in the genome (i.e. the CArGome) have in recent years, unveiled unexpected roles of the myocardin family proteins in lipid and glucose homeostasis. This coactivator family includes the protein myocardin (MYOCD), the myocardin-related transcription factors A and B (MRTF-A/MKL1 and MRTF-B/MKL2) and MASTR (MAMSTR). Here we discuss growing evidence that SRF-driven transcription is controlled by extracellular glucose through activation of the Rho-kinase pathway and actin polymerization. We also describe data showing that adipogenesis is influenced by MLK activity through actions upstream of peroxisome proliferator-activated receptor γ with consequences for whole body fat mass and insulin sensitivity. The recently demonstrated involvement of myocardin coactivators in the biogenesis of caveolae, Ω-shaped membrane invaginations of importance for lipid and glucose metabolism, is finally discussed. These novel roles of myocardin proteins may open the way for new unexplored strategies to combat metabolic diseases such as diabetes, which, at the current incidence, is expected to reach 333 million people worldwide by 2025. This review highlights newly discovered roles of myocardin-related transcription factors in lipid and glucose metabolism as well as novel insights into their well-established role as mediators of stretch-dependent effects in smooth muscle. As co-factors for serum response factor (SRF), MKLs regulates transcription of genes involved in the contractile function of smooth muscle cells. In addition to mechanical stimuli, this regulation has now been found to be promoted by extracellular glucose levels in smooth muscle. Recent reports also suggest that MKLs can regulate a subset of genes involved in the formation of lipid-rich invaginations in the cell membrane called caveolae. Finally, a potential role of MKLs in non-muscle cells has been discovered as they negatively influence adipocyte differentiation.


Asunto(s)
Glucosa/metabolismo , Metabolismo de los Lípidos , Proteínas Nucleares/metabolismo , Transactivadores/metabolismo , Adipogénesis , Animales , Caveolas , Humanos , Proteínas Nucleares/química , Dominios Proteicos , Transactivadores/química
16.
Biochem Biophys Res Commun ; 474(2): 357-363, 2016 05 27.
Artículo en Inglés | MEDLINE | ID: mdl-27109474

RESUMEN

Serotonin (5-HT) is a biogenic monoamine that functions both as a neurotransmitter and a circulating hormone. Recently, the metabolic effects of 5-HT have gained interest and peripheral 5-HT has been proposed to influence lipid metabolism in various ways. Here, we investigated the metabolic effects of 5-HT in isolated, primary rat adipose cells. Incubation with 5-HT suppressed ß-adrenergically stimulated glycerol release and decreased phosphorylation of protein kinase A (PKA)-dependent substrates, hormone sensitive lipase (Ser563) and perilipin (Ser522). The inhibitory effect of 5-HT on lipolysis enhanced the anti-lipolytic effect of insulin, but sustained in the presence of phosphodiesterase inhibitors, OPC3911 and isobuthylmethylxanthine (IBMX). The relative expression of 5-HT1A, -2B and -4 receptor class family were significantly higher in adipose tissue compared to adipose cells, whereas 5-HT1D, -2A and -7 were highly expressed in isolated adipose cells. Similar to 5-HT, 5-HT2 receptor agonists reduced lipolysis while 5-HT1 receptor agonists rather decreased non-stimulated and insulin-stimulated glucose uptake. Together, these data provide evidence of a direct effect of 5-HT on adipose cells, where 5-HT suppresses lipolysis and glucose uptake, which could contribute to altered systemic lipid- and glucose metabolism.


Asunto(s)
Adipocitos/metabolismo , Lipólisis/fisiología , Receptor de Serotonina 5-HT2A/metabolismo , Agonistas de Receptores de Serotonina/administración & dosificación , Serotonina/metabolismo , Adipocitos/efectos de los fármacos , Animales , Células Cultivadas , Relación Dosis-Respuesta a Droga , Lipólisis/efectos de los fármacos , Masculino , Ratas , Ratas Sprague-Dawley
18.
J Lipid Res ; 56(12): 2248-59, 2015 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-26504176

RESUMEN

ApoA-I, the main protein component of HDL, is suggested to be involved in metabolic homeostasis. We examined the effects of Milano, a naturally occurring ApoA-I variant, about which little mechanistic information is available. Remarkably, high-fat-fed mice treated with Milano displayed a rapid weight loss greater than ApoA-I WT treated mice, and a significantly reduced adipose tissue mass, without an inflammatory response. Further, lipolysis in adipose cells isolated from mice treated with either WT or Milano was increased. In primary rat adipose cells, Milano stimulated cholesterol efflux and increased glycerol release, independently of ß-adrenergic stimulation and phosphorylation of hormone sensitive lipase (Ser563) and perilipin (Ser522). Stimulation with Milano had a significantly greater effect on glycerol release compared with WT but similar effect on cholesterol efflux. Pharmacological inhibition or siRNA silencing of ABCA1 did not diminish Milano-stimulated lipolysis, although binding to the cell surface was decreased, as analyzed by fluorescence microscopy. Interestingly, methyl-ß-cyclodextrin, a well-described cholesterol acceptor, dose-dependently stimulated lipolysis. Together, these results suggest that decreased fat mass and increased lipolysis following Milano treatment in vivo is partly explained by a novel mechanism at the adipose cell level comprising stimulation of lipolysis independently of the canonical cAMP/protein kinase A signaling pathway.


Asunto(s)
Adipocitos/efectos de los fármacos , Adipocitos/metabolismo , Apolipoproteína A-I/farmacología , Proteínas Quinasas Dependientes de AMP Cíclico/metabolismo , AMP Cíclico/metabolismo , Lipólisis/efectos de los fármacos , Tejido Adiposo/efectos de los fármacos , Tejido Adiposo/metabolismo , Animales , Colesterol/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL
19.
Diabetologia ; 57(4): 797-800, 2014 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-24442447

RESUMEN

AIMS/HYPOTHESIS: Apolipoprotein A-I (apoA-I), the main protein constituent of HDL, has a central role in the reverse cholesterol-transport pathway, which together with the anti-inflammatory properties of apoA-I/HDL provide cardioprotection. Recent findings of direct stimulation of glucose uptake in muscle by apoA-I/HDL suggest that altered apoA-I and HDL functionality may be a contributing factor to the development of diabetes. We have studied the in vivo effects of short treatments with human apoA-I in a high-fat diet fed mouse model. In addition to native apoA-I, we investigated the effects of the cardioprotective Milano variant (Arg173Cys). METHODS: Male C57Bl6 mice on a high-fat diet for 2 weeks that received a single injection of human apoA-I proteins (wild-type and Milano) were analysed for blood glucose and insulin levels during a 3 h incubation followed by glucose tolerance tests. Incorporation of injected human apoA-I protein into HDLs was analysed by native gel electrophoresis. RESULTS: ApoA-I treatment significantly improved insulin secretion and blood glucose clearance in the glucose tolerance test, with an efficiency exceeding that of lean control animals, and led to decreased basal glucose during the 3 h incubation. Notably, the two apoA-I variants triggered insulin secretion and glucose clearance to the same extent. CONCLUSIONS/INTERPRETATION: ApoA-I treatment leads to insulin- and non-insulin-dependent effects on glucose homeostasis. The experimental model of short-term (2 weeks) feeding of a high-fat diet to C57Bl6 mice provides a suitable and time-efficient system to unravel the resulting tissue-specific mechanisms of acute apoA-I treatment that lead to improved glucose homeostasis.


Asunto(s)
Apolipoproteína A-I/administración & dosificación , Apolipoproteína A-I/farmacología , Glucemia/metabolismo , Resistencia a la Insulina/fisiología , Animales , Glucemia/efectos de los fármacos , Dieta Alta en Grasa/efectos adversos , Modelos Animales de Enfermedad , Humanos , Insulina/metabolismo , Lipoproteínas HDL , Masculino , Ratones , Ratones Endogámicos C57BL
20.
Biochem Biophys Res Commun ; 446(4): 1114-9, 2014 Apr 18.
Artículo en Inglés | MEDLINE | ID: mdl-24680680

RESUMEN

The interest in adiponutrin stems from adiponutrin variant I148M, which is strongly associated to non-alcoholic fatty liver disease. Adiponutrin has to date been considered to be solely an intracellular protein, with a role in lipid metabolism in liver and adipose tissue. However, a physiologically relevant role for adiponutrin has not been found. The aim of this study was to investigate the presence of adiponutrin in human plasma, a new facet of adiponutrin research. We demonstrate that adiponutrin is present in plasma as disulfide-bond dependent multimers, estimated to circulate at a concentration of 1.25-4 nM. Experiments reveal that adiponutrin is released from HepG2 cells in the presence of oleate. The presence of adiponutrin in plasma makes it accessible for clinical investigations and use as a potential biomarker for metabolic disease.


Asunto(s)
Proteínas de la Membrana/sangre , Proteínas de la Membrana/metabolismo , Adulto , Hígado Graso/sangre , Hígado Graso/metabolismo , Femenino , Células Hep G2 , Células Endoteliales de la Vena Umbilical Humana , Humanos , Masculino , Proteínas de la Membrana/química , Persona de Mediana Edad , Enfermedad del Hígado Graso no Alcohólico , Ácido Oléico/metabolismo , Multimerización de Proteína
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