RESUMEN
BACKGROUND: Critical limb-threatening ischemia (CLTI) and type 2 diabetes (T2D) frequently co-exist and often with less favorable outcome after revascularization. The objective was to evaluate the effects of revascularization on muscle function, perfusion and mitochondrial respiration in patients with combined CLTI and T2D. METHODS: A prospective translational observational study. Two groups of patients facing unilateral peripheral revascularization was included: Patients suffering from combined disease with CLTI+T2D (N.=14) and patients suffering from CLTI (N.=15). During pedal exercise testing, calf muscle perfusion was monitored with near-infrared spectroscopy (NIRS) and leg arterial volume flow in the common femoral artery with duplex ultrasound. Calf muscle biopsy and subsequent assessment of mitochondrial respiratory capacity on isolated muscle fibers was performed. Tests was performed before and six weeks after revascularization. RESULTS: After revascularization, patients CLTI+T2D improved in muscle force from 8.48 kg (CI: 7.49-9.46) to 13.11 kg (CI: 11.58-14.63), (P<0.001). Conversely, muscle force in patients suffering from non-diabetic CLTI decreased from 10.03 kg (CI: 9.1-10.96) to 9.73 kg (CI: 8.77-10.69), (P=0.042). Muscle oxygenation during exercise improved more in the CLTI+T2D group (6.36 µM/kg/s [CI: 5.71-7.01] compared to 2.11 µM/kg/s [CI:1.38-2.83] in the CLTI group; P=0.002). No improvement or difference between groups regarding mitochondrial function was detected. CONCLUSIONS: Patients with combined CLTI+T2D, had an unsuspected better effect of revascularization compared to patients with non-diabetic CLTI, in terms of increased muscle force and improved muscle perfusion. Further studies are needed to elucidate the apparent interaction of the CLTI and T2D syndromes.
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Diabetes Mellitus Tipo 2 , Procedimientos Endovasculares , Enfermedad Arterial Periférica , Enfermedad Crónica , Diabetes Mellitus Tipo 2/complicaciones , Diabetes Mellitus Tipo 2/diagnóstico , Humanos , Isquemia/diagnóstico por imagen , Isquemia/cirugía , Recuperación del Miembro , Extremidad Inferior/cirugía , Músculos , Enfermedad Arterial Periférica/diagnóstico por imagen , Enfermedad Arterial Periférica/cirugía , Estudios Prospectivos , Factores de Riesgo , Factores de Tiempo , Resultado del TratamientoRESUMEN
Utilizing the thermogenic capacity of brown adipose tissue is a potential anti-obesity strategy; therefore, the mechanisms controlling expression of thermogenesis-related genes are of interest. Pyruvate kinase (PK) catalyzes the last step of glycolysis and exists as four isoenzymes: PK, liver, PK, red blood cell, PK, muscle (PKM1 and PKM2). PKM2 has both glycolytic and nuclear functions. Here, we report that PKM2 is enriched in brown adipose compared with white adipose tissue. Specific knockdown of PKM2 in mature brown adipocytes demonstrates that silencing of PKM2 does not lead to a decrease in PK activity, but causes a robust upregulation of thermogenic uncoupling protein 1 (Ucp1) and fibroblast growth factor 21 (Fgf21) gene expression. This increase is not mediated by any of the known mechanisms for PKM2-regulated gene expression, thus implying the existence of a novel mechanism for PKM2-dependent effects on gene expression.
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Adipocitos Marrones/enzimología , Adipocitos Marrones/metabolismo , Regulación hacia Abajo , Piruvato Quinasa/metabolismo , Termogénesis/genética , Animales , Línea Celular , Femenino , Factores de Crecimiento de Fibroblastos/genética , Ratones , Piruvato Quinasa/deficiencia , Piruvato Quinasa/genética , Proteína Desacopladora 1/genéticaRESUMEN
Designing effective and safe tuberculosis (TB) subunit vaccines for inhalation requires identification of appropriate antigens and adjuvants and definition of the specific areas to target in the lungs. Magnetic resonance imaging (MRI) enables high spatial resolution, but real-time anatomical and functional MRI of lungs is challenging. Here, we describe the design of a novel gadoteridol-loaded cationic adjuvant formulation 01 (CAF01) for MRI-guided vaccine delivery of the clinically tested TB subunit vaccine candidate H56/CAF01. Gadoteridol-loaded CAF01 liposomes were engineered by using a quality-by-design approach to (i) increase the mechanistic understanding of formulation factors governing the loading of gadoteridol and (ii) maximize the loading of gadoteridol in CAF01, which was confirmed by cryotransmission electron microscopy. The encapsulation efficiency and loading of gadoteridol were highly dependent on the buffer pH due to strong attractive electrostatic interactions between gadoteridol and the cationic lipid component. Optimal gadoteridol loading of CAF01 liposomes showed good in vivo stability and safety upon intrapulmonary administration into mice while generating 1.5-fold MRI signal enhancement associated with approximately 30% T1 relaxation change. This formulation principle and imaging approach can potentially be used for other mucosal nanoparticle-based formulations, species, and lung pathologies, which can readily be translated for clinical use.
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Cationes/química , Compuestos Heterocíclicos/administración & dosificación , Compuestos Heterocíclicos/química , Liposomas/química , Pulmón/efectos de los fármacos , Compuestos Organometálicos/administración & dosificación , Compuestos Organometálicos/química , Adyuvantes Inmunológicos/química , Adyuvantes Farmacéuticos , Animales , Química Farmacéutica/métodos , Femenino , Gadolinio/administración & dosificación , Gadolinio/química , Lípidos/química , Imagen por Resonancia Magnética/métodos , Ratones , Ratones Endogámicos BALB C , Nanopartículas/química , Tuberculosis/tratamiento farmacológico , Vacunas contra la Tuberculosis/química , Vacunas de Subunidad/químicaRESUMEN
PURPOSE: We evaluated muscle proton elimination following similar exercise in the same muscle group following two exercise modalities. METHODS: Seven rowers performed handgrip or rowing exercise for ~ 5 min. The intracellular response of the wrist flexor muscles was evaluated by 31P nuclear magnetic resonance spectroscopy, while arterial and venous forearm blood was collected. RESULTS: Rowing and handgrip reduced intracellular pH to 6.3 ± 0.2 and 6.5 ± 0.1, arterial pH to 7.09 ± 0.03 and 7.40 ± 0.03 and venous pH to 6.95 ± 0.06 and 7.20 ± 0.04 (P < 0.05), respectively. Arterial and venous lactate increased to 17.5 ± 1.6 and 20.0 ± 1.6 mM after rowing while only to 2.6 ± 0.8 and 6.8 ± 0.8 mM after handgrip exercise. Arterio-venous concentration difference of bicarbonate and phosphocreatine recovery kinetics (T50% rowing 1.5 ± 0.7 min; handgrip 1.4 ± 1.0 min) was similar following the two exercise modalities. Yet, intramuscular pH recovery in the forearm flexor muscles was 3.5-fold slower after rowing than after handgrip exercise (T50% rowing of 2 ± 0.1 vs. 7 ± 0.3 min for handgrip). CONCLUSION: Rowing delays intracellular-pH recovery compared with handgrip exercise most likely because rowing, as opposed to handgrip exercise, increases systemic lactate concentration. Thus the intra-to-extra-cellular lactate gradient is small after rowing. Since this lactate gradient is the main driving force for intracellular lactate removal in muscle and, since pHi normalization is closely related to intracellular lactate removal, rowing results in a slower pHi recovery compared to handgrip exercise.
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Ejercicio Físico/fisiología , Antebrazo/irrigación sanguínea , Ácido Láctico/sangre , Músculo Esquelético/metabolismo , Adulto , Fuerza de la Mano/fisiología , Humanos , Concentración de Iones de Hidrógeno , Espectroscopía de Resonancia Magnética , Masculino , Contracción Muscular/fisiología , Flujo Sanguíneo Regional/fisiología , Adulto JovenRESUMEN
AIMS: Poor myometrial contractility has been demonstrated in women at term with diabetes and decreased muscular mitochondrial content and/or function has been extensively implicated in the progression of type 2 diabetes. Alterations of the uterine mitochondrial phenotype in pregnant women with diabetes have yet to be investigated as a causal link to decreased myometrial contractility. METHODS: Observational study of 18 women with diabetes (type 2 and gestational) scheduled for an elective Caesarean section at term with matching controls. A uterine biopsy and fasting blood samples were taken on the day of delivery. RESULTS: Respiration rates in isolated mitochondria and myometrial mRNA levels of genes related to mitochondrial biogenesis were unaffected by diabetes. Mitochondrial quantity examined by quantification of the complexes of the respiratory chain and histology did not indicate alterations in mitochondrial quantity. Citrate syntase activity was higher (0.31 ± 0.02 vs. 0.24 ± 0.02 U/mg protein, P = 0.008), whereas protein content was lower in women with diabetes compared with the control group (94.6 ± 6.9 vs. 118.6 ± 7.4 mg/g wet wt, P = 0.027). Histological examinations did not support any structural alterations in the myometrium or its mitochondria. CONCLUSION: No indication of decreased mitochondrial function, content, morphology, or localization in the myometrium at term in women with diabetes compared with controls was observed. The increase in citrate syntase activity in the myometrium could be explained by the lower protein content in the myometrium, which we suggest is due to alterations in tissue or cellular composition.
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Diabetes Mellitus Tipo 2/metabolismo , Mitocondrias/metabolismo , Miometrio/metabolismo , Complicaciones del Embarazo/fisiopatología , Adulto , Glucemia/metabolismo , Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/patología , Diabetes Mellitus Tipo 2/fisiopatología , Femenino , Humanos , Contracción Muscular , Miometrio/patología , Miometrio/fisiopatología , Fenotipo , Embarazo , Complicaciones del Embarazo/metabolismo , Complicaciones del Embarazo/patología , ARN Mensajero/genética , ARN Mensajero/metabolismo , Útero/metabolismo , Útero/patología , Útero/fisiopatología , Adulto JovenRESUMEN
Nicotinamide adenine dinucleotide (NAD+) can be synthesized by nicotinamide phosphoribosyltransferase (NAMPT). We aimed to determine the role of NAMPT in maintaining NAD+ levels, mitochondrial function, and metabolic homeostasis in skeletal muscle cells. We generated stable Nampt knockdown (sh Nampt KD) C2C12 cells using a shRNA lentiviral approach. Moreover, we applied gene electrotransfer to express Cre recombinase in tibialis anterior muscle of floxed Nampt mice. In sh Nampt KD C2C12 myoblasts, Nampt and NAD+ levels were reduced by 70% and 50%, respectively, and maximal respiratory capacity was reduced by 25%. Moreover, anaerobic glycolytic flux increased by 55%, and 2-deoxyglucose uptake increased by 25% in sh Nampt KD cells. Treatment with the NAD+ precursor nicotinamide riboside restored NAD+ levels in sh Nampt cells and increased maximal respiratory capacity by 18% and 32% in control and sh Nampt KD cells, respectively. Expression of Cre recombinase in muscle of floxed Nampt mice reduced NAMPT and NAD+ levels by 38% and 43%, respectively. Glucose uptake increased by 40%, and mitochondrial complex IV respiration was compromised by 20%. Hypoxia-inducible factor (HIF)-1α-regulated genes and histone H3 lysine 9 (H3K9) acetylation, a known sirtuin 6 (SIRT6) target, were increased in shNampt KD cells. Thus, we propose that the shift toward glycolytic metabolism observed, at least in part, is mediated by the SIRT6/HIF1α axis. Our findings suggest that NAMPT plays a key role for maintaining NAD+ levels in skeletal muscle and that NAMPT deficiency compromises oxidative phosphorylation capacity and alters energy homeostasis in this tissue.
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Citocinas/genética , Metabolismo Energético/genética , Mitocondrias Musculares/fisiología , Músculo Esquelético/metabolismo , Mioblastos/metabolismo , NAD/metabolismo , Nicotinamida Fosforribosiltransferasa/genética , Animales , Metabolismo de los Hidratos de Carbono/genética , Células Cultivadas , Citocinas/metabolismo , Homeostasis/genética , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Nicotinamida Fosforribosiltransferasa/metabolismo , Fosforilación Oxidativa , Transducción de Señal/genéticaRESUMEN
During thermogenic activation, brown adipocytes take up large amounts of glucose. In addition, cold stimulation leads to an upregulation of glycolytic enzymes. Here we have investigated the importance of glycolysis for brown adipocyte glucose consumption and thermogenesis. Using siRNA-mediated knockdown in mature adipocytes, we explored the effect of glucose transporters and glycolytic enzymes on brown adipocyte functions such as consumption of glucose and oxygen. Basal oxygen consumption in brown adipocytes was equally dependent on glucose and fatty acid oxidation, whereas isoproterenol (ISO)-stimulated respiration was fueled mainly by fatty acids, with a significant contribution from glucose oxidation. Knockdown of glucose transporters in brown adipocytes not only impaired ISO-stimulated glycolytic flux but also oxygen consumption. Diminishing glycolytic flux by knockdown of the first and final enzyme of glycolysis, i.e., hexokinase 2 (HK2) and pyruvate kinase M (PKM), respectively, decreased glucose uptake and ISO-stimulated oxygen consumption. HK2 knockdown had a more severe effect, which, in contrast to PKM knockdown, could not be rescued by supplementation with pyruvate. Hence, brown adipocytes rely on glucose consumption and glycolytic flux to achieve maximum thermogenic output, with glycolysis likely supporting thermogenesis not only by pyruvate formation but also by supplying intermediates for efferent metabolic pathways.
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Adipocitos Marrones/efectos de los fármacos , Adipocitos Marrones/metabolismo , Agonistas Adrenérgicos beta/farmacología , Glucosa/metabolismo , Glucólisis/efectos de los fármacos , Consumo de Oxígeno/efectos de los fármacos , Tejido Adiposo Pardo/efectos de los fármacos , Tejido Adiposo Pardo/metabolismo , Animales , Células Cultivadas , Regulación hacia Abajo/efectos de los fármacos , Regulación hacia Abajo/fisiología , Ácidos Grasos/metabolismo , Isoproterenol/farmacología , Metabolismo de los Lípidos/efectos de los fármacos , Masculino , Ratones , Oxidación-Reducción/efectos de los fármacos , Termogénesis/efectos de los fármacosRESUMEN
PURPOSE: We propose a quantitative technique to assess solute uptake into the brain parenchyma based on dynamic contrast-enhanced MRI (DCE-MRI). With this approach, a small molecular weight paramagnetic contrast agent (Gd-DOTA) is infused in the cerebral spinal fluid (CSF) and whole brain gadolinium concentration maps are derived. METHODS: We implemented a 3D variable flip angle spoiled gradient echo (VFA-SPGR) longitudinal relaxation time (T1) technique, the accuracy of which was cross-validated by way of inversion recovery rapid acquisition with relaxation enhancement (IR-RARE) using phantoms. Normal Wistar rats underwent Gd-DOTA infusion into CSF via the cisterna magna and continuous MRI for approximately 130 min using T1-weighted imaging. Dynamic Gd-DOTA concentration maps were calculated and parenchymal uptake was estimated. RESULTS: In the phantom study, T1 discrepancies between the VFA-SPGR and IR-RARE sequences were approximately 6% with a transmit coil inhomogeneity correction. In the in vivo study, contrast transport profiles indicated maximal parenchymal retention of approximately 19% relative to the total amount delivered into the cisterna magna. CONCLUSION: Imaging strategies for accurate 3D contrast concentration mapping at 9.4T were developed and whole brain dynamic concentration maps were derived to study solute transport via the glymphatic system. The newly developed approach will enable future quantitative studies of the glymphatic system in health and disease states. Magn Reson Med 79:1568-1578, 2018. © 2017 International Society for Magnetic Resonance in Medicine.
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Encéfalo/diagnóstico por imagen , Encéfalo/metabolismo , Compuestos Heterocíclicos/líquido cefalorraquídeo , Compuestos Heterocíclicos/farmacocinética , Imagen por Resonancia Magnética/métodos , Compuestos Organometálicos/líquido cefalorraquídeo , Compuestos Organometálicos/farmacocinética , Algoritmos , Animales , Química Encefálica/efectos de los fármacos , Mapeo Encefálico/métodos , Compuestos Heterocíclicos/farmacología , Procesamiento de Imagen Asistido por Computador , Masculino , Compuestos Organometálicos/farmacología , Fantasmas de Imagen , Ratas , Ratas WistarRESUMEN
KEY POINTS: Obesity during pregnancy and childbirth is associated with labour dystocia leading to instrumental or operative delivery, but the underlying pathophysiological mechanisms remain unclear and insufficient uterine contractility has been suggested. This study examined whether reduced myometrial mitochondrial capacity or quantity could contribute as a pathophysiological mechanism to labour dystocia. Data did not support reduced myometrial mitochondrial capacity or quantity in the myometrium at term in obese women, but a reduced myocyte density with increased triglyceride content was demonstrated, which could lead to poorer uterine contractility. These results add to the understanding of systemic effects of obesity, placing also the myometrium at term as an affected non-adipose tissue. ABSTRACT: Obesity is known to increase the risk of labour dystocia and insufficient energy supply, due to reduced mitochondrial capacity or quantity, could be a possible mechanism leading to reduced efficiency of uterine contractility during labour. In the present study of 36 women having an elective Caesarean section at term, obesity did not change mitochondrial phenotype in the myometrial myocyte obtained from uterine biopsies taken at delivery. Respiration rates in isolated mitochondria were unaffected by obesity. No indication of reduced content, investigated by quantification of the complexes of the respiratory chain, or altered regulation, examined by myometrial mRNA levels of genes related to mitochondrial biogenesis and inflammation, was detected. Yet we found increased myometrial triglyceride content in the obese group (2.39 ± 0.26 vs. 1.56 ± 0.20 mm, P = 0.024), while protein content and citrate synthase activity per gram wet weight myometrium were significantly lower in the obese (109.2 ± 7.2 vs. 139.4 ± 5.6 mg g-1 , P = 0.002, and 24.8 ± 1.0 vs. 29.6 ± 1.4 U g-1 wet wt, P = 0.008, respectively). These differences were substantiated by our histological findings where staining for nuclei, cytoplasm, glycogen and collagen supported the idea of a smaller muscle content in the myometrium in obese women. In conclusion no indication of myometrial mitochondrial dysfunction in the isolated state was found, but the observed increase of lipid content might play a role in the pathophysiological mechanisms behind labour dystocia in obese women.
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Metabolismo de los Lípidos , Mitocondrias Musculares/metabolismo , Miometrio/metabolismo , Obesidad/metabolismo , Complicaciones del Embarazo/metabolismo , Adulto , Estudios de Casos y Controles , Femenino , Humanos , Mitocondrias Musculares/ultraestructura , Células Musculares/metabolismo , Células Musculares/ultraestructura , Miometrio/patología , Obesidad/patología , Fenotipo , Embarazo , Complicaciones del Embarazo/patologíaRESUMEN
Taurine ameliorates changes occurring in newborn skeletal muscle as a result of gestational protein restriction in C57BL/6 mice, but taurine supplementation effects may be exaggerated in C57BL/6 mice due to their inherent excessive taurinuria.We examined if maternal taurine supplementation could ameliorate changes in gene expression levels, properties of mitochondria, myogenesis, and nutrient transport and sensing, in male newborn skeletal muscle caused by a maternal low protein (LP) diet in Wistar rats.LP diet resulted in an 11% non-significant decrease in birth weight, which was not rescued by taurine supplementation (LP-Tau). LP-Tau offspring had significantly lower birth weight compared to controls. Gene expression profiling revealed 895 significantly changed genes, mainly an LP-induced down-regulation of genes involved in protein translation. Taurine fully or partially rescued 32% of these changes, but with no distinct pattern as to which genes were rescued.Skeletal muscle taurine content in LP-Tau offspring was increased, but no changes in mRNA levels of the taurine synthesis pathway were observed. Taurine transporter mRNA levels, but not protein levels, were increased by LP diet.Nutrient sensing signaling pathways were largely unaffected in LP or LP-Tau groups, although taurine supplementation caused a decrease in total Akt and AMPK protein levels. PAT4 amino acid transporter mRNA was increased by LP, and normalized by taurine supplementation.In conclusion, gestational protein restriction in rats decreased genes involved in protein translation in newborn skeletal muscle and led to changes in nutrient transporters. Taurine partly rescued these changes, hence underscoring the importance of taurine in development.
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Dieta con Restricción de Proteínas/efectos adversos , Músculo Esquelético/efectos de los fármacos , Efectos Tardíos de la Exposición Prenatal , Taurina/farmacología , Transcriptoma/genética , Animales , Femenino , Masculino , Mitocondrias/efectos de los fármacos , Músculo Esquelético/metabolismo , Embarazo , Ratas , Ratas WistarRESUMEN
Brown adipose tissue takes up large amounts of glucose during cold exposure in mice and humans. Here we report an induction of glucose transporter 1 expression and increased expression of several glycolytic enzymes in brown adipose tissue from cold-exposed mice. Accordingly, these genes were also induced after ß-adrenergic activation of cultured brown adipocytes, concomitant with accumulation of hypoxia inducible factor-1α (HIF-1α) protein levels. HIF-1α accumulation was dependent on uncoupling protein 1 and generation of mitochondrial reactive oxygen species. Expression of key glycolytic enzymes was reduced after knockdown of HIF-1α in mature brown adipocytes. Glucose consumption, lactate export and glycolytic capacity were reduced in brown adipocytes depleted of Hif-1α. Finally, we observed a decreased ß-adrenergically induced oxygen consumption in Hif-1α knockdown adipocytes cultured in medium with glucose as the only exogenously added fuel. These data suggest that HIF-1α-dependent regulation of glycolysis is necessary for maximum glucose metabolism in brown adipocytes.
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Adipocitos Marrones/metabolismo , Glucosa/metabolismo , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Adipocitos Marrones/efectos de los fármacos , Agonistas Adrenérgicos beta/farmacología , Animales , Células Cultivadas , Frío , Regulación de la Expresión Génica/efectos de los fármacos , Regulación Enzimológica de la Expresión Génica/efectos de los fármacos , Proteínas Facilitadoras del Transporte de la Glucosa/genética , Proteínas Facilitadoras del Transporte de la Glucosa/metabolismo , Glucólisis/efectos de los fármacos , Glucólisis/genética , Hipoxia , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Masculino , Ratones , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Modelos Biológicos , Termogénesis/efectos de los fármacos , Termogénesis/genéticaRESUMEN
BACKGROUND: This study elucidates the effects on muscle mitochondrial function in patients suffering from combined peripheral arterial disease (PAD) and type 2 diabetes (T2D) and the relation to patient symptoms and treatment. METHODS: Near infrared spectroscopy (NIRS) calf muscle exercise tests were conducted on 40 subjects, 15 (PAD), 15 (PAD+T2D) and 10 healthy age matched controls (CTRL) recruited from the vascular outpatient clinic at Gentofte County Hospital, Denmark. Calf muscle biopsies (~80 mg, gastrocnemius and anterior tibial muscles) were sampled and mitochondrial function tested applying high resolution oxygraphy on isolated muscle fibers. RESULTS: The NIRS exercise tests showed evidence of mitochondrial dysfunction in the PAD+T2D group by a longer recovery of the deoxygenation resulting from exercise in spite of a higher exercise oxygenation level compared to the PAD group. This was confirmed by a ~30% reduction in oxygen consumption in the muscle biopsy tests for the PAD+T2D compared to the PAD group (P<0.05). We claim that this mitochondrial dysfunction partly explains the ~30% reduction in tread mill walking distance for the PAD+T2D group observed in this study. CONCLUSIONS: These findings support the use of early surgical revascularization in the PAD+T2D group, in order to obtain better walking performance and probably reduced risk of permanent mitochondrial damage.
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Diabetes Mellitus Tipo 2/complicaciones , Mitocondrias/patología , Músculo Esquelético/patología , Enfermedad Arterial Periférica/diagnóstico por imagen , Enfermedad Arterial Periférica/fisiopatología , Anciano , Estudios de Casos y Controles , Dinamarca , Prueba de Esfuerzo , Femenino , Humanos , Modelos Lineales , Masculino , Persona de Mediana Edad , Mitocondrias/metabolismo , Músculo Esquelético/irrigación sanguínea , Consumo de Oxígeno , Espectroscopía Infrarroja Corta , CaminataRESUMEN
Extreme diets consisting of either high fat (HF) or high sucrose (HS) may lead to insulin resistance in skeletal muscle, often associated with mitochondrial dysfunction. However, it is not known if these diets alter normal interactions of pyruvate and fatty acid oxidation at the level of the mitochondria. Here, we report that rat muscle mitochondria does show the normal Randle-type fat-carbohydrate interaction seen in vivo. The mechanism behind this metabolic flexibility at the level of the isolated mitochondria is a regulation of the flux-ratio: pyruvate dehydrogenase (PDH)/ß-oxidation to suit the actual substrate availability, with the PDH flux as the major point of regulation. We further report that this regulatory mechanism of carbohydrate-fat metabolic interaction surprisingly is lost in mitochondria obtained from animals exposed for 12 weeks to a HF- or a HS diet as compared to rats given a normal chow diet. The mechanism seems to be a loss of the PDH flux decrease seen in controls, when fatty acid is supplied as substrate in addition to pyruvate, and vice versa for the supply of pyruvate as substrate to mitochondria oxidizing fatty acid. Finally, we report that the calculated TCA flux in the isolated mitochondria under these circumstances shows a significant reduction (~50%) after the HF diet and an even larger reduction (~75%) after the HS diet, compared with the chow group. Thus, it appears that obesogenic diets as those applied here have major influence on key metabolic performance of skeletal muscle mitochondria.
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Grasas de la Dieta/metabolismo , Sacarosa en la Dieta/metabolismo , Ácidos Grasos/metabolismo , Resistencia a la Insulina/fisiología , Mitocondrias Musculares/metabolismo , Músculo Esquelético/metabolismo , Animales , Dieta Alta en Grasa , Oxidación-Reducción , Complejo Piruvato Deshidrogenasa/metabolismo , Ratas , Ratas WistarRESUMEN
AIMS/HYPOTHESIS: Normal glucose metabolism depends on pancreatic secretion of insulin and glucagon. The bihormonal hypothesis states that while lack of insulin leads to glucose underutilisation, glucagon excess is the principal factor in diabetic glucose overproduction. A recent study reported that streptozotocin-treated glucagon receptor knockout mice have normal glucose tolerance. We investigated the impact of acute disruption of glucagon secretin or action in a mouse model of severe diabetes by three different approaches: (1) alpha cell elimination; (2) glucagon immunoneutralisation; and (3) glucagon receptor antagonism, in order to evaluate the effect of these on glucose tolerance. METHODS: Severe diabetes was induced in transgenic and wild-type mice by streptozotocin. Glucose metabolism was investigated using OGTT in transgenic mice with the human diphtheria toxin receptor expressed in proglucagon producing cells allowing for diphtheria toxin (DT)-induced alpha cell ablation and in mice treated with either a specific high affinity glucagon antibody or a specific glucagon receptor antagonist. RESULTS: Near-total alpha cell elimination was induced in transgenic mice upon DT administration and resulted in a massive decrease in pancreatic glucagon content. Oral glucose tolerance in diabetic mice was neither affected by glucagon immunoneutralisation, glucagon receptor antagonism, nor alpha cell removal, but did not deteriorate further compared with mice with intact alpha cell mass. CONCLUSIONS/INTERPRETATION: Disruption of glucagon action/secretion did not improve glucose tolerance in diabetic mice. Near-total alpha cell elimination may have prevented further deterioration. Our findings support insulin lack as the major factor underlying hyperglycaemia in beta cell-deficient diabetes.
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Diabetes Mellitus Experimental , Glucagón , Intolerancia a la Glucosa , Insulina/deficiencia , Animales , Glucemia/efectos de los fármacos , Glucemia/metabolismo , Diabetes Mellitus Experimental/inducido químicamente , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/patología , Toxina Diftérica , Glucagón/antagonistas & inhibidores , Glucagón/metabolismo , Glucagón/fisiología , Péptido 1 Similar al Glucagón/metabolismo , Células Secretoras de Glucagón/efectos de los fármacos , Células Secretoras de Glucagón/patología , Intolerancia a la Glucosa/sangre , Intolerancia a la Glucosa/tratamiento farmacológico , Intolerancia a la Glucosa/genética , Prueba de Tolerancia a la Glucosa , Insulina/metabolismo , Células Secretoras de Insulina/efectos de los fármacos , Células Secretoras de Insulina/metabolismo , Células Secretoras de Insulina/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Receptores de Glucagón/antagonistas & inhibidores , Receptores de Glucagón/genética , EstreptozocinaRESUMEN
BACKGROUND: Achilles tendinopathy is a painful inflammatory condition characterized by swelling, stiffness and reduced function of the Achilles tendon. Kager's fat pad is an adipose tissue located in the area anterior to the Achilles tendon. Observations reveal a close physical interplay between Kager's fat pad and its surrounding structures during movement of the ankle, suggesting that Kager's fat pad may stabilize and protect the mechanical function of the ankle joint. AIM: The aim of this study was to characterize whether Achilles tendinopathy was accompanied by changes in expression of inflammatory markers and metabolic enzymes in Kager's fat pad. METHODS: A biopsy was taken from Kager's fat pad from 31 patients with chronic Achilles tendinopathy and from 13 healthy individuals. Gene expression was measured by reverse transcription-quantitative PCR. Focus was on genes related to inflammation and lipid metabolism. RESULTS: Expression of the majority of analyzed inflammatory marker genes was increased in patients with Achilles tendinopathy compared to that in healthy controls. Expression patterns of the patient group were consistent with reduced lipolysis and increased fatty acid ß-oxidation. In the fat pad, the pain-signaling neuropeptide substance P was found to be present in one third of the subjects in the Achilles tendinopathy group but in none of the healthy controls. CONCLUSION: Gene expression changes in Achilles tendinopathy patient samples were consistent with Kager's fat pad being more inflamed than in the healthy control group. Additionally, the results indicate an altered lipid metabolism in Kager's fat pad of Achilles tendinopathy patients.
Asunto(s)
Tendón Calcáneo/metabolismo , Tendón Calcáneo/patología , Tejido Adiposo/metabolismo , Tejido Adiposo/patología , Tendinopatía/metabolismo , Tendinopatía/patología , Adulto , Biomarcadores , Biopsia , Estudios de Casos y Controles , Enfermedad Crónica , Citocinas/genética , Citocinas/metabolismo , Metabolismo Energético , Femenino , Expresión Génica , Humanos , Mediadores de Inflamación/metabolismo , Metabolismo de los Lípidos , Masculino , Persona de Mediana Edad , Factores de Riesgo , Tendinopatía/genética , Adulto JovenAsunto(s)
Dieta Alta en Grasa , Glucosa/metabolismo , Metabolismo de los Lípidos/efectos de los fármacos , Mitocondrias Musculares/efectos de los fármacos , Taurina/farmacología , Animales , Glucemia/efectos de los fármacos , Glucemia/metabolismo , Respiración de la Célula/efectos de los fármacos , Suplementos Dietéticos , Intolerancia a la Glucosa/etiología , Intolerancia a la Glucosa/metabolismo , Lípidos/sangre , Masculino , Mitocondrias Musculares/fisiología , Músculo Esquelético/efectos de los fármacos , Músculo Esquelético/metabolismo , Ratas , Ratas WistarAsunto(s)
Fructosa/farmacología , Homeostasis/efectos de los fármacos , Taurina/metabolismo , Animales , Dieta , Carbohidratos de la Dieta/farmacología , Suplementos Dietéticos , Ingestión de Líquidos/efectos de los fármacos , Ingestión de Alimentos/efectos de los fármacos , Ingestión de Alimentos/fisiología , Hígado/efectos de los fármacos , Hígado/metabolismo , Masculino , Músculo Esquelético/efectos de los fármacos , Músculo Esquelético/metabolismo , Ratas , Ratas Wistar , Taurina/farmacologíaRESUMEN
BACKGROUND: Large mammals are capable of thermoregulation shortly after birth due to the presence of brown adipose tissue (BAT). The majority of BAT disappears after birth and is replaced by white adipose tissue (WAT). RESULTS: We analyzed the postnatal transformation of adipose in sheep with a time course study of the perirenal adipose depot. We observed changes in tissue morphology, gene expression and metabolism within the first two weeks of postnatal life consistent with the expected transition from BAT to WAT. The transformation was characterized by massively decreased mitochondrial abundance and down-regulation of gene expression related to mitochondrial function and oxidative phosphorylation. Global gene expression profiling demonstrated that the time points grouped into three phases: a brown adipose phase, a transition phase and a white adipose phase. Between the brown adipose and the transition phase 170 genes were differentially expressed, and 717 genes were differentially expressed between the transition and the white adipose phase. Thirty-eight genes were shared among the two sets of differentially expressed genes. We identified a number of regulated transcription factors, including NR1H3, MYC, KLF4, ESR1, RELA and BCL6, which were linked to the overall changes in gene expression during the adipose tissue remodeling. Finally, the perirenal adipose tissue expressed both brown and brite/beige adipocyte marker genes at birth, the expression of which changed substantially over time. CONCLUSIONS: Using global gene expression profiling of the postnatal BAT to WAT transformation in sheep, we provide novel insight into adipose tissue plasticity in a large mammal, including identification of novel transcriptional components linked to adipose tissue remodeling. Moreover, our data set provides a useful resource for further studies in adipose tissue plasticity.
Asunto(s)
Tejido Adiposo Pardo/metabolismo , Tejido Adiposo Blanco/metabolismo , Perfilación de la Expresión Génica , Regulación del Desarrollo de la Expresión Génica , Ovinos/genética , Tejido Adiposo Pardo/patología , Tejido Adiposo Blanco/patología , Animales , Citrato (si)-Sintasa/metabolismo , Análisis por Conglomerados , ADN Mitocondrial/genética , ADN Mitocondrial/metabolismo , Regulación hacia Abajo , Canales Iónicos/genética , Canales Iónicos/metabolismo , Mitocondrias/genética , Proteínas Mitocondriales/genética , Proteínas Mitocondriales/metabolismo , Análisis de Componente Principal , Ovinos/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Proteína Desacopladora 1 , Regulación hacia ArribaRESUMEN
It is well known that few weeks of high fat (HF) diet may induce metabolic disturbances and mitochondrial dysfunction in skeletal muscle. However, little is known about the effects of long-term HF exposure and effects on brain mitochondria are unknown. Wistar rats were fed either chow (13E% fat) or HF diet (60E% fat) for 1 year. The HF animals developed obesity, dyslipidemia, insulin resistance, and dysfunction of isolated skeletal muscle mitochondria: state 3 and state 4 were 30% to 50% increased (P<0.058) with palmitoyl carnitine (PC), while there was no effect with pyruvate as substrate. Adding also succinate in state 3 resulted in a higher substrate control ratio (SCR) with PC, but a lower SCR with pyruvate (P<0.05). The P/O2 ratio was lower with PC (P<0.004). However, similar tests on isolated brain mitochondria from the same animal showed no changes with the substrates relevant for brain (pyruvate and 3-hydroxybutyrate). Thus, long-term HF diet was associated with obesity, dyslipidemia, insulin resistance, and significantly altered mitochondrial function in skeletal muscle. Yet, brain mitochondria were unaffected. We suggest that the relative isolation of the brain due to the blood-brain barrier may play a role in this strikingly different phenotype of mitochondria from the two tissues of the same animal.