Numerical 3D modeling of heat transfer in human tissues for microwave radiometry monitoring of Brown fat metabolismo

Background: Brown adipose tissue (BAT) plays an important role in whole body metabolism and could potentially mediate weight gain and insulin sensitivity. Although some imaging techniques allow BAT detection, there are currently no viable methods for continuous acquisition of BAT energy expenditure. We present a non-invasive technique for long term monitoring of BAT metabolism using microwave radiometry. Methods: A multilayer 3D computational model was created in HFSS™ with 1.5 mm skin, 3-10 mm subcutaneous fat, 200 mm muscle and a BAT region (2-6 cm3) located between fat and muscle. Based on this model, a log-spiral antenna was designed and optimized to maximize reception of thermal emissions from the target (BAT). The power absorption patterns calculated in HFSS™ were combined with simulated thermal distributions computed in COMSOL® to predict radiometric signal measured from an ultra-low-noise microwave radiometer. The power received by the antenna was characterized as a function of different levels of BAT metabolism under cold and noradrenergic stimulation. Results: The optimized frequency band was 1.5-2.2 GHz, with averaged antenna efficiency of 19%. The simulated power received by the radiometric antenna increased 2-9 mdBm (noradrenergic stimulus) and 4-15 mdBm (cold stimulus) corresponding to increased 15-fold BAT metabolism. Conclusions: Results demonstrated the ability to detect thermal radiation from small volumes (2-6 cm3) of BAT located up to 12 mm deep and to monitor small changes (0.5°C) in BAT metabolism. As such, the developed miniature radiometric antenna sensor appears suitable for non-invasive long term monitoring of BAT metabolism.

Plac8 is an inducer of C/EBPβ required for brown fat differentiation, thermoregulation, and control of body weight.

Brown adipocytes oxidize fatty acids to produce heat in response to cold or to excessive energy intake; stimulation of brown fat development and function may thus counteract obesity. Brown adipogenesis requires activation of the transcription factor C/EBPβ and recruitment of the zinc finger protein Prdm16, but upstream inducers of these proteins are incompletely defined. Here, we show that genetic inactivation of Plac8, a gene encoding an evolutionarily conserved protein, induces cold intolerance, and late-onset obesity, as well as abnormal morphology and impaired function of brown adipocytes. Using brown preadipocyte lines we show that Plac8 is required for brown fat differentiation, that its overexpression induces C/EBPβ and Prdm16, and that upon induction of differentiation Plac8 associates with C/EBPβ and binds to the C/EBPβ promoter to induce its transcription. Thus, Plac8 is a critical upstream regulator of brown fat differentiation and function that acts, at least in part, by inducing C/EBPβ expression.

Thermogenic and metabolic consequences of thyroid hormone treatment in brown and white adipose tissue

Male rats were treated with triiodothyronine in the drinking water for 12 days. In vitro rates of isoprenaline stimulated lipolysis were significantly greater in brown but not white adipose tissue. Rates of [14C]glucose incorporation into triacylglycerols were significantly reduced in BAT (brown adipose tissue) and WAT (white adipose tissue) under basal and isoprenaline stimulated conditions. In a second experiment, hyperthyroid animals showed impaired weight gain, despite increased food intake during 19 days' treatment. Energy expenditure on days 5 and 12, and BAT core temperature differences (TBAT - TCORE) on day 19, were significantly greater than in control animals. Epididymal white fat pad weight was reduced and interscapular brown fat pad weight increased by triiodothyronine treatment.

Orexin restores aging-related brown adipose tissue dysfunction in male mice

The aging process causes an increase in percent body fat, but the mechanism remains unclear. In the present study we examined the impact of aging on brown adipose tissue (BAT) thermogenic activity as potential cause for the increase in adiposity. We show that aging is associated with interscapular BAT morphologic abnormalities and thermogenic dysfunction. In vitro experiments revealed that brown adipocyte differentiation is defective in aged mice. Interscapular brown tissue in aged mice is progressively populated by adipocytes bearing white morphologic characteristics. Aged mice fail to mobilize intracellular fuel reserves from brown adipocytes and exhibit deficiency in homeothermy. Our results suggest a role for orexin (OX) signaling in the regulation of thermogenesis during aging. Brown fat dysfunction and age-related assimilation of fat mass were accelerated in mice in which OX-producing neurons were ablated. Conversely, OX injections in old mice increased multilocular morphology, increased core body temperature, improved cold tolerance, and reduced adiposity. These results argue that BAT can be targeted for interventions to reverse age-associated increase in fat mass.

Sensing of glucose in the brain.

The brain, and in particular the hypothalamus and brainstem, have been recognized for decades as important centers for the homeostatic control of feeding, energy expenditure, and glucose homeostasis. These structures contain neurons and neuronal circuits that may be directly or indirectly activated or inhibited by glucose, lipids, or amino acids. The detection by neurons of these nutrient cues may become deregulated, and possibly cause metabolic diseases such as obesity and diabetes. Thus, there is a major interest in identifying these neurons, how they respond to nutrients, the neuronal circuits they form, and the physiological function they control. Here I will review some aspects of glucose sensing by the brain. The brain is responsive to both hyperglycemia and hypoglycemia, and the glucose sensing cells involved are distributed in several anatomical sites that are connected to each other. These eventually control the activity of the sympathetic or parasympathetic nervous system, which regulates the function of peripheral organs such as liver, white and brown fat, muscle, and pancreatic islets alpha and beta cells. There is now evidence for an extreme diversity in the sensing mechanisms used, and these will be reviewed.

Nonshivering thermogenesis capacity associated to mitochondrial DNA haplotypes and gender in the greater white-toothed shrew, Crocidura russula.

A selection gradient was recently suggested as one possible cause for a clinal distribution of mitochondrial DNA (mtDNA) haplotypes along an altitudinal transect in the greater white-toothed shrew, Crocidura russula (Ehinger et al. 2002). One mtDNA haplotype (H1) rare in lowland, became widespread when approaching the altitudinal margin of the distribution. As H1 differs from the main lowland haplotype by several nonsynonymous mutations (including on ATP6), and as mitochondria play a crucial role in metabolism and thermogenesis, distribution patterns might stem from differences in the thermogenic capacity of different mtDNA haplotypes. In order to test this hypothesis, we measured the nonshivering thermogenesis (NST) associated with different mtDNA haplotypes. Sixty-two shrews, half of which had the H1 haplotype, were acclimated in November at semioutdoor conditions and measured for NST throughout winter. Our results showed the crucial role of NST for winter survival in C. russula. The individuals that survived winter displayed a higher significant increase in NST during acclimation, associated with a significant gain in body mass, presumably from brown fat accumulation. The NST capacity (ratio of NST to basal metabolic rate) was exceptionally high for such a small species. NST was significantly affected by a gender x haplotype interaction after winter-acclimation: females bearing the H1 haplotype displayed a better thermogenesis at the onset of the breeding season, while the reverse was true for males. Altogether, our results suggest a sexually antagonistic cyto-nuclear selection on thermogenesis.

Plac8 is required for white adipocyte differentiation in vitro and cell number control in vivo.

Plac8 belongs to an evolutionary conserved family of proteins, mostly abundant in plants where they control fruit weight through regulation of cell number. In mice, Plac8 is expressed both in white and brown adipose tissues and we previously showed that Plac8(-/-) mice develop late-onset obesity, with abnormal brown fat differentiation and reduced thermogenic capacity. We also showed that in brown adipocytes, Plac8 is an upstream regulator of C/EBPβ expression. Here, we first assessed the role of Plac8 in white adipogenesis in vitro. We show that Plac8 is induced early after induction of 3T3-L1 adipocytes differentiation, a process that is prevented by Plac8 knockdown; similarly, embryonic fibroblasts obtained from Plac8 knockout mice failed to form adipocytes upon stimulation of differentiation. Knockdown of Plac8 in 3T3-L1 was associated with reduced expression of C/EBPβ, Krox20, and Klf4, early regulators of the white adipogenic program, and we show that Plac8 could transactivate the C/EBPβ promoter. In vivo, we show that absence of Plac8 led to increased white fat mass with enlarged adipocytes but reduced total number of adipocytes. Finally, even though Plac8(-/-) mice showed impaired thermogenesis due to brown fat dysfunction, this was not associated with changes in glycemia or plasma free fatty acid and triglyceride levels. Collectively, these data indicate that Plac8 is an upstream regulator of C/EBPβ required for adipogenesis in vitro. However, in vivo, Plac8 is dispensable for the differentiation of white adipocytes with preserved fat storage capacity but is required for normal fat cell number regulation.

Brain glucose sensing and neural regulation of insulin and glucagon secretion.

Glucose homeostasis requires the tight regulation of glucose utilization by liver, muscle and white or brown fat, and glucose production and release in the blood by liver. The major goal of maintaining glycemia at ∼ 5 mM is to ensure a sufficient flux of glucose to the brain, which depends mostly on this nutrient as a source of metabolic energy. This homeostatic process is controlled by hormones, mainly glucagon and insulin, and by autonomic nervous activities that control the metabolic state of liver, muscle and fat tissue but also the secretory activity of the endocrine pancreas. Activation or inhibition of the sympathetic or parasympathetic branches of the autonomic nervous systems are controlled by glucose-excited or glucose-inhibited neurons located at different anatomical sites, mainly in the brainstem and the hypothalamus. Activation of these neurons by hyper- or hypoglycemia represents a critical aspect of the control of glucose homeostasis, and loss of glucose sensing by these cells as well as by pancreatic β-cells is a hallmark of type 2 diabetes. In this article, aspects of the brain-endocrine pancreas axis are reviewed, highlighting the importance of central glucose sensing in the control of counterregulation to hypoglycemia but also mentioning the role of the neural control in β-cell mass and function. Overall, the conclusions of these studies is that impaired glucose homeostasis, such as associated with type 2 diabetes, but also defective counterregulation to hypoglycemia, may be caused by initial defects in glucose sensing.

Regulation of lipid metabolism in adipocytes and hepatocytes by hexarelin through scavenger receptor CD36

Les sécrétines de l’hormone de croissance (GHRPs) sont de petits peptides synthétiques capables de stimuler la sécrétion de l’hormone de croissance à partir de l’hypophyse via leur liaison au récepteur de la ghréline GHS-R1a. Le GHRP hexaréline a été utilisé afin d’étudier la distribution tissulaire de GHS-R1a et son effet GH-indépendant. Ainsi, par cette approche, il a été déterminé que l’hexaréline était capable de se lier à un deuxième récepteur identifié comme étant le récepteur scavenger CD36. Ce récepteur possède une multitude de ligands dont les particules oxLDL et les acides gras à longue chaîne. CD36 est généralement reconnu pour son rôle dans l’athérogénèse et sa contribution à la formation de cellules spumeuses suite à l’internalisation des oxLDL dans les macrophages/monocytes. Auparavant, nous avions démontré que le traitement des macrophages avec l’hexaréline menait à l’activation de PPARƔ via sa liaison à GHS-R1a, mais aussi à CD36. De plus, une cascade d’activation impliquant LXRα et les transporteurs ABC provoquait également une augmentation de l’efflux du cholestérol. Une stimulation de la voie du transport inverse du cholestérol vers les particules HDL entraînait donc une diminution de l’engorgement des macrophages de lipides et la formation de cellules spumeuses. Puisque CD36 est exprimé dans de multiples tissus et qu’il est également responsable du captage des acides gras à longue chaîne, nous avons voulu étudier l’impact de l’hexaréline uniquement à travers sa liaison à CD36. Dans le but d’approfondir nos connaissances sur la régulation du métabolisme des lipides par CD36, nous avons choisi des types cellulaires jouant un rôle important dans l’homéostasie lipidique n’exprimant pas GHS-R1a, soient les adipocytes et les hépatocytes. L’ensemble de mes travaux démontre qu’en réponse à son interaction avec l’hexaréline, CD36 a le potentiel de réduire le contenu lipidique des adipocytes et des hépatocytes. Dans les cellules adipeuses, l'hexaréline augmente l’expression de plusieurs gènes impliqués dans la mobilisation et l’oxydation des acides gras, et induit également l’expression des marqueurs thermogéniques PGC-1α et UCP-1. De même, hexaréline augmente l’expression des gènes impliqués dans la biogenèse mitochondriale, un effet accompagné de changements morphologiques des mitochondries; des caractéristiques observées dans les types cellulaires ayant une grande capacité oxydative. Ces résultats démontrent que les adipocytes blancs traités avec hexaréline ont la capacité de se transformer en un phénotype similaire aux adipocytes bruns ayant l’habileté de brûler les acides gras plutôt que de les emmagasiner. Cet effet est également observé dans les tissus adipeux de souris et est dépendant de la présence de CD36. Dans les hépatocytes, nous avons démontré le potentiel de CD36 à moduler le métabolisme du cholestérol. En réponse au traitement des cellules avec hexaréline, une phosphorylation rapide de LKB1 et de l’AMPK est suivie d’une phosphorylation inhibitrice de l’HMG-CoA réductase (HMGR), l’enzyme clé dans la synthèse du cholestérol. De plus, la liaison d'hexaréline à CD36 provoque le recrutement d’insig-2 à HMGR, l’étape d’engagement dans sa dégradation. La dégradation de HMGR par hexaréline semble être dépendante de l’activité de PPARƔ et de l’AMPK. Dans le but d’élucider le mécanisme d’activation par hexaréline, nous avons démontré d’une part que sa liaison à CD36 provoque une déphosphorylation de Erk soulevant ainsi l’inhibition que celui-ci exerce sur PPARƔ et d’autre part, un recrutement de l’AMPK à PGC-1α expliquant ainsi une partie du mécanisme d’activation de PPARƔ par hexaréline. Les résultats générés dans cette thèse ont permis d’élucider de nouveaux mécanismes d’action de CD36 et d'approfondir nos connaissances de son influence dans la régulation du métabolisme des lipides.

Monoacylglycerol, alpha/beta-hydrolase domain-6, and the regulation of insulin secretion and energy metabolism

Le cycle glycérolipides/acides gras libres (GL/FFA) est une voie métabolique clé qui relie le métabolisme du glucose et des acides gras et il est composé de deux processus métaboliques appelés lipogenèse et lipolyse. Le cycle GL/FFA, en particulier la lipolyse des triglycérides, génère diverses molécules de signalisation pour réguler la sécrétion d'insuline dans les cellules bêta pancréatiques et la thermogenèse non-frissonnante dans les adipocytes. Actuellement, les lipides provenant spécifiquement de la lipolyse impliqués dans ce processus sont mal connus. L’hydrolyse des triglycérides dans les cellules β est réalisée par les actions successives de la triglycéride lipase adipocytaire pour produire le diacylglycérol, ensuite par la lipase hormono-sensible pour produire le monoacylglycérol (MAG) et enfin par la MAG lipase (MAGL) qui relâche du glycerol et des acides gras. Dans les cellules bêta, la MAGL classique est très peu exprimée et cette étude a démontré que l’hydrolyse de MAG dans les cellules β est principalement réalisée par l'α/β-Hydrolase Domain-6 (ABHD6) nouvellement identifiée. L’inhibition d’ABHD6 par son inhibiteur spécifique WWL70, conduit à une accumulation des 1-MAG à longues chaines saturées à l'intérieur des cellules, accompagnée d’une augmentation de la sécrétion d'insuline stimulée par le glucose (GSIS). Baisser les niveaux de MAG en surexprimant ABHD6 dans la lignée cellulaire bêta INS832/13 réduit la GSIS, tandis qu’une augmentation des niveaux de MAG par le « knockdown » d’ABHD6 améliore la GSIS. L'exposition aiguë des monoacylglycérols exogènes stimule la sécrétion d'insuline de manière dose-dépendante et restaure la GSIS supprimée par un inhibiteur de lipases appelé orlistat. En outre, les souris avec une inactivation du gène ABHD6 dans tous les tissus (ABHD6-KO) et celles avec une inactivation du gène ABHD6 spécifiquement dans la cellule β présentent une GSIS stimulée, et leurs îlots montrent une augmentation de la production de monoacylglycérol et de la sécrétion d'insuline en réponse au glucose. L’inhibition d’ABHD6 chez les souris diabétiques (modèle induit par de faibles doses de streptozotocine) restaure la GSIS et améliore la tolérance au glucose. De plus, les résultats montrent que les MAGs non seulement améliorent la GSIS, mais potentialisent également la sécrétion d’insuline induite par les acides gras libres ainsi que la sécrétion d’insuline induite par divers agents et hormones, sans altération de l'oxydation et l'utilisation du glucose ainsi que l'oxydation des acides gras. Nous avons démontré que le MAG se lie à la protéine d’amorçage des vésicules appelée Munc13-1 et l’active, induisant ainsi l’exocytose de l'insuline. Sur la base de ces observations, nous proposons que le 1-MAG à chaines saturées agit comme facteur de couplage métabolique pour réguler la sécrétion d'insuline et que ABHD6 est un modulateur négatif de la sécrétion d'insuline. En plus de son rôle dans les cellules bêta, ABHD6 est également fortement exprimé dans les adipocytes et son niveau est augmenté avec l'obésité. Les souris dépourvues globalement d’ABHD6 et nourris avec une diète riche en gras (HFD) montrent une faible diminution de la prise alimentaire, une diminution du gain de poids corporel et de la glycémie à jeun et une amélioration de la tolérance au glucose et de la sensibilité à l'insuline et ont une activité locomotrice accrue. En outre, les souris ABHD6-KO affichent une augmentation de la dépense énergétique et de la thermogenèse induite par le froid. En conformité avec ceci, ces souris présentent des niveaux élevés d’UCP1 dans les adipocytes blancs et bruns, indiquant le brunissement des adipocytes blancs. Le phénotype de brunissement est reproduit dans les souris soit en les traitant de manière chronique avec WWL70 (inhibiteur d’ABHD6) ou des oligonucléotides anti-sense ciblant l’ABHD6. Les tissus adipeux blanc et brun isolés de souris ABHD6-KO montrent des niveaux très élevés de 1-MAG, mais pas de 2-MAG. L'augmentation des niveaux de MAG soit par administration exogène in vitro de 1-MAG ou par inhibition ou délétion génétique d’ABHD6 provoque le brunissement des adipocytes blancs. Une autre évidence indique que les 1-MAGs sont capables de transactiver PPARα et PPARγ et que l'effet de brunissement induit par WWL70 ou le MAG exogène est aboli par les antagonistes de PPARα et PPARγ. L’administration in vivo de l’antagoniste de PPARα GW6471 à des souris ABHD6-KO inverse partiellement les effets causés par l’inactivation du gène ABHD6 sur le gain de poids corporel, et abolit l’augmentation de la thermogenèse, le brunissement du tissu adipeux blanc et l'oxydation des acides gras dans le tissu adipeux brun. L’ensemble de ces observations indique que ABHD6 régule non seulement l’homéostasie de l'insuline et du glucose, mais aussi l'homéostasie énergétique et la fonction des tissus adipeux. Ainsi, 1-MAG agit non seulement comme un facteur de couplage métabolique pour réguler la sécrétion d'insuline en activant Munc13-1 dans les cellules bêta, mais régule aussi le brunissement des adipocytes blancs et améliore la fonction de la graisse brune par l'activation de PPARα et PPARγ. Ces résultats indiquent que ABHD6 est une cible prometteuse pour le développement de thérapies contre l'obésité, le diabète de type 2 et le syndrome métabolique.

Rabies virus in a pregnant naturally infected southern yellow bat (Lasiurus ega)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Efeito da suplementação com ácido linoléico conjugado e do treinamento em natação sobre a composição corporal e os parâmetros bioquímicos de ratos Wistar em crescimento

OBJETIVO: Avaliar os efeitos da suplementação com ácido linoléico conjugado, associada ao treinamento moderado em natação, sobre a composição corporal, o consumo e a eficiência alimentar, a glicemia, o perfil lipídico e o glicogênio muscular e hepático de ratos Wistar. MÉTODOS: Ratos Wistar (30 dias) foram divididos em: sedentário, sedentário suplementado, treinado e treinado suplementado. Permaneceram em gaiolas individuais com comida e água ad libitum, temperatura de 23ºC (com variação de1ºC) e ciclo claro-escuro de 12 horas, durante 8 semanas. A sessão de natação durou 1 hora e foi realizada três vezes/semana, bem como a suplementação com ácido linoléico conjugado a 2%. Após sacrifício, o plasma, os tecidos adiposos brancos e o marrom, o músculo gastrocnêmio e o fígado foram coletados e pesados. RESULTADOS: A suplementação per se não promoveu modificação na ingestão alimentar e na massa corporal dos animais. Houve aumento na glicemia de jejum (p<0,05), nas lipoproteínas de alta densidade (p<0,05), no colesterol total (p<0,05) e redução dos triacilgliceróis. A suplementação associada ao treinamento reduziu a massa corporal (p<0,05) e aumentou o peso relativo do tecido adiposo, do fígado e a glicemia de jejum. CONCLUSÃO: A suplementação com ácido linoléico conjugado associada à prática de exercício físico parece ter influência no balanço energético, mas, por outro lado, o aumento no peso do fígado indica que a ingestão deste ácido graxo pode ter efeitos indesejáveis, aumentando as chances de desenvolvimento do fígado gorduroso. Estes achados apontam perspectivas para novos estudos envolvendo análises histológicas do fígado, expressão gênica de enzimas chaves do metabolismo lipídico e de carboidratos, associados ou não a diferentes protocolos de treinamento físico.

Effects of acute cold exposure on rectal temperature, blood glucose and plasma free fatty acids in alloxan-diabetic rats

These experiments were carried out to study the effects of acute cold exposure (0-2°C/4 hr) on rectal temperature, blood glucose and plasma free fatty acids (FFA) in alloxan-diabetic rats. Male Wistar rats weighing 170-190 g were used and diabetes was induced by i.v. alloxan injection (40 mg/kg body wt). Cold exposure produced severe hypothermia in diabetic rats. After 4 hr of cold, blood glucose of diabetic rats was reduced from 296±16 to 86t±12 mg/dl (P<0.01), and FFA increased slightly, but was not statistically different (P>0.05) from the initial value. As expected, interscapular brown adipose tissue (IBAT) and retroperitoneal and epididymal white adipose tissues were significantly lower in diabetic than in control rats. Cold exposure reduced total IBAT lipids in control but not in diabetic animals. The results of this experiment suggest that diabetic rats were unable to maintain body temperature in the cold, probably because of a failure to generate an adequate amount of heat by nonshivering thermogenesis in brown adipose tissue.

The development of a febrile response to pyrogen in the thyroid-deficient rabbit

The development of the febrile response to E. coli lipopolysaccharide (1.5 μg/kg, i.v.) in thyroid-deficient rabbits has been studied. Twenty-eight New Zealand White rabbits weighing 2.1-2.3 kg were used. Hypothyroidism was induced by treatment with propylthiouracil (100 or 200 mg/kg body wt./15 days). Thyroid-deficient animals showed a reduction in the febrile response to lipopolysaccharide, but the effect was significantly different (p<0.01) from the control only for rabbits treated with 200 mg/kg of propylthiouracil. Propranolol (2 mg/kg, i.p.) given 30 min before lipopolysaccharide also reduced (p<0.01) the fever response in control rabbits. The results of this experiment are consistent with the hypothesis that the reduction in the febrile response of thyroid-deficient rabbits is due to the reduced number of β-adrenergic receptors, or to a change in the availability of neurotransmitter in thermogenically active tissues, such as brown fat.