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Podeu consultar dades primàries associades a l'article a: http://hdl.handle.net/2445/111074, Background. Food selection and ingestion both in humans and rodents, often is a critical factor in determining excess energy intake and its related disorders. Methods. Two different concepts of high-fat diets were tested for their obesogenic effects in rats; in both cases, lipids constituted about 40% of their energy intake. The main difference with controls fed standard lab chow, was, precisely, the lipid content. Cafeteria diets (K) were self-selected diets devised to be desirable to the rats, mainly because of its diverse mix of tastes, particularly salty and sweet. This diet was compared with another, more classical high-fat (HF) diet, devised not to be as tasty as K, and prepared by supplementing standard chow pellets with fat. We also analysed the influence of sex on the effects of the diets. Results. K rats grew faster because of a high lipid, sugar and protein intake, especially the males, while females showed lower weight but higher proportion of body lipid. In contrast, the weight of HF groups were not different from controls. Individual nutrient's intake were analysed, and we found that K rats ingested large amounts of both disaccharides and salt, with scant differences of other nutrients' proportion between the three groups. The results suggest that the key differential factor of the diet eliciting excess energy intake was the massive presence of sweet and salty tasting food. Conclusions. The significant presence of sugar and salt appears as a powerful inducer of excess food intake, more effective than a simple (albeit large) increase in the diet's lipid content. These effects appeared already after a relatively short treatment. The differential effects of sex agree with their different hedonic and obesogenic response to diet.

White adipose tissue (WAT) produces large amounts of lactate and glycerol from glucose. We used mature epididymal adipocytes to analyse the relative importance of glycolytic versus lipogenic glycerol in adipocytes devoid of external stimuli. Cells were incubated (24/48 h) with 7/14 mM glucose; half of the wells contained 14C-glucose. We analysed glucose label fate, medium metabolites, and the expression of key genes coding for proteins controlling glycerol metabolism. The effects of initial glucose levels were small, but time of incubation increased cell activity and modified its metabolic focus. The massive efflux of lactate was uniform with time and unrelated to glucose concentration; however, glycerol-3P synthesis was higher in the second day of incubation, being largely incorporated into the glycerides-glycerol fraction. Glycerophosphatase expression was not affected by incubation. The stimulation of glycerogenic enzymes’ expression was mirrored in lipases. The result was a shift from medium glycolytic to lipolytic glycerol released as a consequence of increased triacylglycerol turnover, in which most fatty acids were recycled. Production of glycerol seems to be an important primary function of adipocytes, maintained both by glycerogenesis and acyl-glycerol turnover. Production of 3C fragments may also contribute to convert excess glucose into smaller, more readily usable, 3C metabolites.

Podeu consultar dades primàries associades a l'article a: http://hdl.handle.net/2445/96861, Background. A ``cafeteria'' diet is a self-selected high-fat diet, providing an excess of energy, which can induce obesity. Excess of lipids in the diet hampers glucose utilization eliciting insulin resistance, which, further limits amino acid oxidation for energy. Methods. Male Wistar rats were exposed for a month to ``cafeteria'' diet. Rats were cannulated and fluorescent microspheres were used to determine blood flow. Results. Exposure to the cafeteria diet did not change cardiac output, but there was a marked shift in organ irrigation. Skin blood flow decreased to compensate increases in lungs and heart. Blood flow through adipose tissue tended to increase in relation to controls, but was considerably increased in brown adipose tissue (on a weight basis). Discussion. The results suggest that the cafeteria diet-induced changes were related to heat transfer and disposal.

Female and male adult Wistar rats were fed standard chow or a simplified cafeteria diet for one month. Then, the rats were killed and the white adipose tissue (WAT) in four sites: perigonadal, retroperitoneal, mesenteric and subcutaneous (inguinal) were sampled and frozen. The complete WAT weight in each site was measured. Gene expression analysis of key lipid and glucose metabolism enzymes were analyzed, as well as tissue and plasma lactate and the activity of lactate dehydrogenase. Lactate gradients between WAT and plasma were estimated. The influence of sex and diet (and indirectly WAT mass) on lactate levels and their relationships with lactate dehydrogenase activity and gene expressions were also measured. A main conclusion is the high production of lactate by WAT, practically irrespective of site, diet or sex. Lactate production is a direct correlate of lactate dehydrogenase activity in the tissue. Furthermore, lactate dehydrogenase activity is again directly correlated with the expression of the genes Ldha and Ldhb for this enzyme. In sum, the ability to produce lactate by WAT is not directly dependent of WAT metabolic state. We postulate that, in WAT, a main function of the lactate dehydrogenase path may be that of converting excess available glucose to 3C fragments, as a way to limit tissue self-utilization as substrate, to help control glycaemia and/or providing short chain substrates for use as energy source elsewhere. More information must be gathered before a conclusive role of WAT in the control of glycaemia, and the full existence of a renewed glucose-lactate-fatty acid cycle is definitely established.

White adipose tissue (WAT) produces lactate in significant amount from circulating glucose, especially in obesity;Under normoxia, 3T3L1 cells secrete large quantities of lactate to the medium, again at the expense of glucose and proportionally to its levels. Most of the glucose was converted to lactate with only part of it being used to synthesize fat. Cultured adipocytes were largely anaerobic, but this was not a Warburg-like process. It is speculated that the massive production of lactate, is a process of defense of the adipocyte, used to dispose of excess glucose. This way, the adipocyte exports glucose carbon (and reduces the problem of excess substrate availability) to the liver, but the process may be also a mechanism of short-term control of hyperglycemia. The in vivo data obtained from adipose tissue of male rats agree with this interpretation.

In the metabolic syndrome, glucocorticoid activity is increased, but circulating levels show little change. Most of blood glucocorticoids are bound to corticosteroid-binding globulin (CBG), which liver expression and circulating levels are higher in females than in males. Since blood hormones are also bound to blood cells, and the size of this compartment is considerable for androgens and estrogens, we analyzed whether sex or eating a cafeteria diet altered the compartmentation of corticosterone in rat blood. The main corticosterone compartment in rat blood is that specifically bound to plasma proteins, with smaller compartments bound to blood cells or free. Cafeteria diet increased the expression of liver CBG gene, binding plasma capacity and the proportion of blood cell-bound corticosterone. There were marked sex differences in blood corticosterone compartmentation in rats, which were unrelated to testosterone. The use of a monoclonal antibody ELISA and a polyclonal Western blot for plasma CBG compared with both specific plasma binding of corticosterone and CBG gene expression suggested the existence of different forms of CBG, with varying affinities for corticosterone in males and females, since ELISA data showed higher plasma CBG for males, but binding and Western blot analyses (plus liver gene expression) and higher physiological effectiveness for females. Good cross- reactivity to the antigen for polyclonal CBG antibody suggests that in all cases we were measuring CBG.The different immunoreactivity and binding affinity may help explain the marked sex-related differences in plasma hormone binding as sex-linked different proportions of CBG forms.

Oleoyl-estrone (OE) is a powerful slimming agent that is also present in plasma and adipose tissue, where it is synthesized. It acts through the formation of a derivative W. OE effects (and W levels) are proportional to the dose. OE reduces food intake but maintains energy expenditure (thermogenesis). The energy gap is fulfilled with adipose tissue fat, sparing body protein and maintaining glycemia (and glycogen) with lower insulin and leptin levels. OE (in fact W) acts through specific receptors, different from those of estrogen. OE increases cholesterol catabolism, reducing hypercholesterolemia in obese rats. The main metabolic effect on adipose tissue is a lowering of lipid synthesis, maintaining unchanged the intracellular lipolytic processes; the imbalance favors the progressive loss of fat, which is largely used by the muscle. OE administration induces additive effects with other antiobesity agents, such as β3-adrenergic agonists, forcing a massive loss of lipid. Corticosteroids, markedly limit OE action by altering the liver control of lipogenesis. OE also inhibits the action of 17β-hydroxysteroid dehydrogenase, decreasing the synthesis of β-estradiol and testosterone. Discontinuous treatment allows for maximal efficacy both in rats and humans. OE has the advantage that the loss of fat is maintained and does not require additional dietary limitations.

Metabolic syndrome developed in consequence of an evolutionary inadequacy: the human body was unprepared for a dietary excess of nutrients, especially lipids (largely in detriment of carbohydrate). This excess awakens metabolic signals akin to those of starvation, in which the main energy staple is the body"s own lipid reserve. Lipid dietary abundance prevents the use of glucose, which in turn limits the oxidation of amino acids. To ward against a subsequent avalanche of substrates, the immune system and hypertrophied tissues (for example, adipose) elicit a series of defence responses. This response is probably the ultimate basis of a disease that is manifested as various pathologies, which were initially defined as distinct entities but which are slowly being seen as a single pathognomic unit in the literature. Based on their common origin of the ample availability of food in our modern society, the cluster of diseases comprising the metabolic syndrome is probably best described as a single multifaceted disease.

The metabolic syndrome is basically a maturity-onset disease. Typically, its manifestations begin to flourish years after the initial dietary or environmental aggression began. Since most hormonal, metabolic, or defense responses are practically immediate, the procrastinated response do not seem justified. Only in childhood, the damages of the metabolic syndrome appear with minimal delay. Sex affects the incidence of the metabolic syndrome, but this is more an effect of timing than absolute gender differences, females holding better than males up to menopause, when the differences between sexes tend to disappear. The metabolic syndrome is related to an immune response, countered by a permanent increase in glucocorticoids, which keep the immune system at bay but also induce insulin resistance, alter the lipid metabolism, favor fat deposition, mobilize protein, and decrease androgen synthesis. Androgens limit the operation of glucocorticoids, which is also partly blocked by estrogens, since they decrease inflammation (which enhances glucocorticoid release). These facts suggest that the appearance of the metabolic syndrome symptoms depends on the strength (i.e., levels) of androgens and estrogens. The predominance of glucocorticoids and the full manifestation of the syndrome in men are favored by decreased androgen activity. Low androgens can be found in infancy, maturity, advanced age, or because of their inhibition by glucocorticoids (inflammation, stress, medical treatment). Estrogens decrease inflammation and reduce the glucocorticoid response. Low estrogen (infancy, menopause) again allow the predominance of glucocorticoids and the manifestation of the metabolic syndrome. It is postulated that the equilibrium between sex hormones and glucocorticoids may be a critical element in the timing of the manifestation of metabolic syndrome-related pathologies.

Background: The presence of lipid besides muscle fibres facilitates the energy supply for exercise, but it is also indicative of insulin resistance in the untrained. Muscle lipid is associated with increased dietary energy: hyperlipidic diets induce an increase in intramyocellular lipid deposition in skeletal muscle. Methods: In the present study we analyzed the changes in soleus (a red-fibre muscle) intracellular muscle content under a hyperlipidic (cafeteria) diet in Wistar rats. We also analyzed in parallel the mitochondrial content a relative index of energy output capability. Results: Cafeteria diet-fed rats contained more lipid and mitochondria per unit of muscle section area than controls. Conclusions: The correlation found in the increases of muscle lipid and mitochondria hit at this increase as an adaptation of muscle to oxidize excess energy substrates under conditions of excess energy availability, probably contributing to adaptive thermogenesis.

A method for the measurement of carbamoyl-phosphate synthetase I activity in animal tissues has been developed using the livers of rats under normal and hyperproteic diets. The method is based on the incorporation of 14C-ammonium bicarbonate to carbamoyl-phosphate in the presence of ATP-Mg and N-acetyl-glutamate. The reaction is stopped by chilling, lowering the pH and adding ethanol. Excess bicarbonate is flushed out under a gentle stream of cold CO2. The only label remaining in the medium was that incorporated into carbamoyl-phosphate, since all 14C-CO2 from bicarbonate was eliminated. The method is rapid and requires only a low pressure supply of CO2 to remove the excess substrate. The reaction is linear up to 10 min using homogenate dilutions of 1:20 to 1:200 (w/v). Rat liver activity was in the range of 89±8 nkat/g. Hyperproteic diet resulted in a significant 1.4-fold increase. The design of the method allows for the processing of multiple samples at the same time, and incubation medium manipulation is unnecessary, since the plastic incubation vial and its contents are finally counted together.

This review is focused on the fate of dietary glucose under conditions of chronically high energy (largely fat) intake, evolving into the metabolic syndrome. We are adapted to carbohydrate-rich diets similar to those of our ancestors. Glucose is the main energy staple, but fats are our main energy reserves. Starvation drastically reduces glucose availability, forcing the body to shift to fatty acids as main energy substrate, sparing glucose and amino acids. We are not prepared for excess dietary energy, our main defenses being decreased food intake and increased energy expenditure, largely enhanced metabolic activity and thermogenesis. High lipid availability is a powerful factor decreasing glucose and amino acid oxidation. Present-day diets are often hyperenergetic, high on lipids, with abundant protein and limited amounts of starchy carbohydrates. Dietary lipids favor their metabolic processing, saving glucose, which additionally spares amino acids. The glucose excess elicits hyperinsulinemia, which may derive, in the end, into insulin resistance. The available systems of energy disposal could not cope with the excess of substrates, since they are geared for saving not for spendthrift, which results in an unbearable overload of the storage mechanisms. Adipose tissue is the last energy sink, it has to store the energy that cannot be used otherwise. However, adipose tissue growth also has limits, and the excess of energy induces inflammation, helped by the ineffective intervention of the immune system. However, even under this acute situation, the excess of glucose remains, favoring its final conversion to fat. The sum of inflammatory signals and deranged substrate handling induce most of the metabolic syndrome traits: insulin resistance, obesity, diabetes, liver steatosis, hyperlipidemia and their compounded combined effects. Thus, a maintained excess of energy in the diet may result in difficulties in the disposal of glucose, eliciting inflammation and the development of the metabolic syndrome

White adipose tissue (WAT) is a disperse organ acting as energy storage depot and endocrine/paracrine controlling factor in the management of energy availability and inflammation. WAT sites response under energy-related stress is not uniform. In the present study we have analyzed how different WAT sites respond to limited food restriction as a way to better understand the role of WAT in the pathogenesis of the metabolic syndrome.

Estrone is a powerful growth-inducing hormone that is present in milk, mainly in the form of fatty acid esters, at concentrations that promote growth in experimental animals. We present here a method useful for the measurement of this natural hormone in foods and applied it to several common dairy products. Samples were frozen, finely powdered, and lyophilized then extracted with trichloromethane/methanol; the dry extract was saponified with potassium hydroxide. The free estrone evolved was extracted with ethyl acetate and was used for the estimation of total estrone content through radioimmunoassay. Application of the method to dairy products showed high relative levels of total estrone (essentially acyl-estrone) in milk, in the range of 1 ¿M, which were halved in skimmed milk. Free estrone levels were much lower, in the nanomolar range. A large proportion of estrone esters was present in all other dairy products, fairly correlated with their fat content. The amount of estrone carried by milk is well within the range, where its intake may exert a physiological response in the sucklings for which it is provided. These growth-inducing and energy expenditure-lowering effects may affect humans ingesting significant amounts of dairy products.

La oleoil-estrona induce la movilización rápida de los líp,dos del tejido adiposo blanco, disminuyendo en paralelo los niveles circulantes de lípidos plasmáticos. En el presente trabajo se ha intentado determinar la implicación del músculo en la oxidación de dichos lípidos.

[eng] Obesity is a widespread crippling and life-shortening disease that can be defined as a pathologic accumulation of fat reserves. In spite of its epidemic distribution, no fully effective treatments are available. The strategies used for the treatment of obesity have relied mainly on the limitation of energy intake or/and increasing energy expenditure. The most widely used method to limit energy intake has been the use of hypocaloric diets. Their effectivity is limited and fade away rapidly with time. Nevertheless, the sound use of hypocaloric diets is yet the mainstay of the fight against overweight. Inhibition of the absorption of nutrients through specific digestive enzyme inhibitors has been also used. Bariatric surgery is now practically the only fairly effective way to treat the morbidly obese. Conductist conditioning has been used to maintain the obese as far as possible from food, but the results are often poor. However, adequate instruction of the obese on basic nutritional knowledge, and nutritional reeducation are a tool not to be neglected. Exercise is the easiest way to increase energy expenditure. but this increase is only transient; in any case it potentiates the slimming effects of dietary restriction. There are a growing number of drugs used for the treatment of obesity, and more are just being under study and development. The main target of these drugs is to diminish the cravings of appetite as a way to help the obese to limit ingestion, but other drugs tend to increase thermogenesis, easing the consumption of fat reserves; often both effects add up. The most widely studied drugs are serotonergic drugs acting on the brain and adrenergic agents acting both on appetite and heat production. Several hormones, metabolites and even poisons have been postulated as antiobesity agents, but now the most promising areas of study rely on hypothalamic control of appetite, thermogenesis and regulatory control of the mass of fat, the latter achieved through signall, [cat] L'obesitat és una malaltia molt estesa que limita l'activitat i escurça la vida, i que es pot definir com un emmagatzemament patològic de reserves de greix. Malgrat la difusió epidèmica, no hi ha cap sistema plenament efectiu disponible per tractar-la. Les estratègies emprades per al tractament de l'obesitat s'han basat principalment en la l'imitació de la ingesta i/o l'increment de la despesa energètica. EI mètode més emprat per l'imitar la ingesta energètica ha estat la utilització de dietes hipocalòriques, però l'efectivitat és limitada i es perd ràpidament amb el temps. Malgrat això, la utilització adequada de dietes hipocalòriques constitueix encara el principal procediment en la lluita contra el sobrepès. També s'ha emprat el bloqueig de l'absorció de nutrients mitjançant la inhibició específica d'enzims digestius. La cirurgia bariàtrica és ara pràcticament l'únic mètode prou efectiu per tractar els obesos mòrbids. S'ha utilitzat el condicionament conductista per mantenir els obesos allunyats del menjar, però els resultats són sovint poc satisfactoris. No obstant això, la informació adequada que reben els obesos sobre els principis elementals de la nutrició, així com la reeducació nutricional són una eina que no s'ha de deixar de costat. L'exercici és Ia forma més senzilla d'augmentar la despesa energètica i, tot i que aquest increment és sols transitori, potencia els efectes aprimadors de la restricció dietètica. Hi ha un nombre considerable de fàrmacs que han estat emprats per al tractament de l'obesitat i encara n'hi ha més que estan essent estudiats i desenvolupats. EI principal objectiu d'aquests fàrmacs és disminuir la gana a fi d'ajudar l'obès a reduir la quantitat de menjar, però altres drogues tendeixen a incrementar la termogènesi, tot facilitant la utilització de les reserves grasses; sovint ambdós efectes tenen lloc a l'hora. les drogues més àmpliament estudiades són les serotoninèrgiques, que actuen sobre el cervell, i els agents adrenèrgics que a tan orientats a trobar solucions parcials al complex problema de l’obesitat. L’esforç coordinat en recerca bàsica i el desenvolupament de fàrmacs efectius, junt amb una adequada informació als pacients i I’actualització dels coneixements del personal sanitari que treballa en aquest camp, són les condicions essencials per poder fer front a aquesta malaltia perillosa que s’estén d’una manera incontrolada: I’obesitat.

L'obesitat es una malaltia mal coneguda queafecta un nombre creixent de persones arreu del mon. L'increment de la incidencia de l'obesitat en alguns paisos s'ha atribu 'it a un increment del consurn de determinats aliments i, sobretot, de les disponibilitats energetiques, malgrat haver-se produit durant els ultims anys un descens significatiu en el consum energetic, en disminuir la ingestio de greixos i de sucre. Aquesta contradiccio es explicada per alguns com una consequencia del sedentarisme, per d'altres per la incidencia de gens que regulen l'obesitat i encara uns altres I'atribueixen a causes desconegudes complementades per tots aquests factors. Sigui com sigui, la morbiditat Iligada a I'obesitat es considerable i tambe ho es la mortalitat addicional que genera.

Conscious female adult lean and obese Zucker rats were injected through the jugular vein with radioactive iodine-labeled murine leptin; in the ensuing 8 min, four blood samples were sequentially extracted from the carotid artery. The samples were used in a modified RIA for leptin, in which paired tubes received the same amount of either labeled or unlabeled leptin, thus allowing us to estimate both leptin levels and specific radioactivity. The data were used to determine the decay curve parameters from which the half-life of leptin (5.46 ± 0.23 min for lean rats and 6.99 ± 0.75 min for obese rats) as well as the size of its circulating pool (32 pmol/kg for lean rats and 267 pmol/kg for obese rats) and the overall degradation rate (96 fkat/kg for lean rats and 645 fkat/kg for obese rats) were estimated. These values are consistent with the hormonal role of leptin and the need for speedy changes in its levels in response to metabolic challenge.

Rats chronically cannulated in the carotid artery and the muscular branch of the femoral vein were subjected to a cold (4 °C) environment for up to 2 h. The changes in blood flow (measured with 46Sc microspheres) and arterio-venous differences in the concentrations of glucose, lactate, triacylglycerols and amino acids allowed the estimation of substrate (and energy) balances across the hindleg. Mean glucose uptake was 0.28mmol min21, mean lactate release was 0.33mmol min21 and the free fatty acid basal release of 0.31mmol min21 was practically zero upon exposure to the cold; the initial uptake of triacylglycerols gave place to a massive release following exposure. The measurement of PO·, PCO· and pH also allowed the estimation of oxygen, CO2 and bicarbonate balances and respiratory quotient changes across the hindleg. The contribution of amino acids to the energy balance of the hindleg was assumed to be low. These data were used to determine the sources of energy used to maintain muscle shivering with time. Three distinct phases were observed in hindleg substrate utilization. (1) The onset of shivering, with the use of glucose/glycogen and an increase in lactate efflux. Lipid oxidation was practically zero (respiratory quotient near 1), but the uptake of triacylglycerols from the blood remained unchanged. (2) A substrate-energy shift, with drastically decreased use of glucose/glycogen, and of lactate efflux; utilization of triacylglycerol as practically the sole source of energy (respiratory quotient approximately 0.7); decreasing uptake of triacylglycerol and increased tissue lipid mobilization. (3) The onset of a new heat-homeostasis setting for prolonged cold-exposure, with maintenance of muscle energy and heat production based on triacylglycerol utilization and efflux from the hindleg (muscle plus skin and subcutaneous adipose masses) contributing energy to help sustain heat production by the core organs and surrounding brown adipose tissue.

The activities of aspartate and alanine transaminase, serine dehydratase, arginase, glutamate dehydrogenase, adenylate deaminase and glutamine synthetase were determined in the stomach and small intestine of developing rats. Despite the common embryonic origin of the intestine and stomach, their enzymes showed quite different activity levels and patterns of development, depending on their roles. Most enzyme activities were low during late intrauterine life and after birth, attaining adult levels with the change of diet at weaning. No arginase activity was found in the stomach and no changes were detected in adenylate deaminase in the stomach or intestine throughout the period studied. Alanine transaminase, serine dehydratase and, to some extent, glutamine synthetase levels, significantly higher in late intrauterine life, decreased after birth, suggesting that the foetal stomach has a transient ability to handle amino acids.

El mejillón es una especie que, debido a sus funciones reproductivas, consume una gran cantidad de energía, suministrada en su mayor parte por el glucógeno, que el individuo ha ido acumulando en grandes cantidades durante la primera fase de su ciclo biológico; estos niveles pueden ser muy elevados según el momento del ciclo biológico de que se trate, obteniéndose un máximo del 28% de glucógeno referido a peso seco y un mínimo de 1'7%. La presencia de esta enorme cantidad de material gluciálico, que se moviliza de modo general y relativamente rápido, hace que el mejillón disponga de unos sistemas de síntesis y degradación del glucógeno muy eficaces y capaces de ser altamente regulados de un modo estacional, con el fin de utilizar la energía contenida durante la gametogénesis. Hasta el momento de realizar la presente tesis no se habían estudiado de una manera sistemática los mecanismos degradativos y biosintéticos del glucógeno en el mejillón, determinándose en cambio ampliamente los niveles, características físicas y químicas, así como la presencia de otros azúcares, solubles, en su composición. La gran variabilidad en contenido de glucógeno de las diversas clases de moluscos hace que el estudio de tales mecanismos sea complicado. Para poder iniciar nuestro trabajo, hemos seleccionado una serie de muestras de mejillones procedentes de comercialización, pertenecientes a la especie "Mytilus edulis" L., considerando dicha especie en sentido más lato, puesto que la heterogeneidad fenotípica no nos permitía circunscribir el estudio a una cepa uniforme y seleccionada. La variabilidad de las muestras fue comprobada estudiando la correlación entre diversos parámetros biométricos, tales como la longitud máxima de la concha, la anchura máxima, el grosor y el peso escurrido del animal completo. El principal inconveniente que presentaban los animales utilizados era que habían tenido que soportar el relativamente largo viaje desde los centros de producción a los lugares de consumo, a lo que debía añadirse el tiempo de almacenaje previo a la comercialización, que era variable. Durante todo este tiempo el animal mantiene la concha cerrada, conservando dentro el agua intervalvar y sufriendo un período de anoxia superior al que soporta en condiciones naturales durante la marea baja, y que tal vez puede haber afectado de alguna manera a los niveles de enzimas estudiados, aunque supusimos de un modo apriorístico que este efecto sería sólo de tipo cuantitativo y no cualitativo. Otro problema, quizás más importante que el anterior, era que estos animales debían soportar un prolongado periodo de hambre, desde el momento en que eran extraídos del vivero y transportados a la estación depuradora, donde si bien pueden respirar son sometidos a un tiempo variable a un flujo de agua filtrada y desinfectada, por lo que durante este periodo filtran agua que carece de materiales en suspensión y de este modo se vacía el tubo digestivo, de manera que los animales sobreviven a base de sus reservas, especialmente en lo que respecta al hepatopáncreas, donde hemos podido encontrar niveles extraordinariamente variables de glúcidos solubles y de glucógeno según la vivacidad de los animales, que suponemos correlacionada con el tiempo transcurrido entre la captura y el análisis.A la hora de escoger los especímenes objeto de estudio, se desecharon todos aquellos que llegasen defectuosos (concha rota, valvas entreabiertas) y los seleccionados fueron conservados en la nevera, generalmente bajo agua de mar artificial, a unos 4ºC.

Oleoyl-estrone (OE) is a powerful slimming agent that is also present in plasma and adipose tissue, where it is synthesized. It acts through the formation of a derivative W. OE effects (and W levels) are proportional to the dose. OE reduces food intake but maintains energy expenditure (thermogenesis). The energy gap is fulfilled with adipose tissue fat, sparing body protein and maintaining glycemia (and glycogen) with lower insulin and leptin levels. OE (in fact W) acts through specific receptors, different from those of estrogen. OE increases cholesterol catabolism, reducing hypercholesterolemia in obese rats. The main metabolic effect on adipose tissue is a lowering of lipid synthesis, maintaining unchanged the intracellular lipolytic processes; the imbalance favors the progressive loss of fat, which is largely used by the muscle. OE administration induces additive effects with other antiobesity agents, such as β3-adrenergic agonists, forcing a massive loss of lipid. Corticosteroids, markedly limit OE action by altering the liver control of lipogenesis. OE also inhibits the action of 17β-hydroxysteroid dehydrogenase, decreasing the synthesis of β-estradiol and testosterone. Discontinuous treatment allows for maximal efficacy both in rats and humans. OE has the advantage that the loss of fat is maintained and does not require additional dietary limitations.

Las dietas terapéuticas, utilizadas para el tratamiento de la obesidad representan siempre un compromiso de limitación del aporte energético creando los menos problemas posibles, para que el margen de utilidad sea efectivo se requiere que el aporte calórico sea muy bajo para combatir la caida del gasto energético que lleva aparejada su utilización, pero esto afecta negativamente el metabolismo nitrogenado, por lo que la utifización de estas dietas viene limitado en el tiempo. Su efectividad a largo plazo es también limitada. A pesar de todo siempre es preferible su utifización que las peligrosas dietas cetogénicas o disociadas, o las dietas populares creadas al amparo de campañas publicitarias y cuya efectividad terapéutica es nula. Las dietas sirven para aportar los nutrientes que necesitamos, pero también para dar placer y satisfacción.

1. The blood flow, PO2, pH and PCO2 have been estimated in portal and suprahepatic veins as well as in hepatic artery of fed and overnight starved rats given an oral glucose load. From these data the net intestinal, hepatic and splanchnic balances for oxygen and bicarbonate were calculated. The oxygen consumption of the intact animal has also been measured under comparable conditions. 2. The direct utilization of oxygen balances as energy equivalents when establishing the contribution of energy metabolism of liver and intestine to the overall energy expenses of the rat, has been found to be incorrect, since it incorporates the intrinsic error of interorgan proton transfer through bicarbonate. Liver and intestine produced high net bicarbonate balances in all situations tested, implying the elimination (by means of oxidative pathways, i.e. consuming additional oxygen) of high amounts of H+ generated with bicarbonate. The equivalence in energy output of the oxygen balances was then corrected for bicarbonate production to 11-54% lower values. 3. Intestine and liver consume a high proportion of available oxygen, about one-half in basal (fed or starved) conditions and about one-third after gavage, the intestine consumption being about 15% in all situations tested and the liver decreasing its oxygen consumption with gavage.

[eng] The current theories about endothelial inflammation in the adipose tissue in metabolic syndrome point to hypoxia as one of its main causes. Nevertheless, our group has found that both 3T3L1 adipocytes, under normoxic conditions, and WAT, in vivo, consume large amount of glucose, with high production of lactate and glycerol irrespective of oxygen availability. This thesis is based on the hypothesis that adipocytes act essentially as glycolytic cells impervious to hypoxia, and their metabolism may help reduce the blood glucose levels. We assumed that hypoxia, could affect the cells of the stromal fraction, eliciting an inflammatory response. To this purpose, we have studied ex vivo the glycolytic and anaerobic capacity of adult adipocytes and stromal vascular cells of both sex and obtained from different WAT sites, developing the methodology needed for a quantitative comparative analysis of data obtained from the same cultured cells well. We found that adipocytes, despite being the cells present in WAT in lower numbers, occupied almost the whole volume of the tissue, a consequence of their huge size due to their inert fat vacuole. The overall “live cell" volume represented only about 1.5% of the tissue, thus showing a very high metabolic activity of WAT in relative terms. Adipocytes ex vivo incubated with glucose, also took large amounts of the sugar, irrespective of its concentration, releasing instead 3C metabolites, such as lactate and glycerol to the medium. Lactate was fully derived from glucose and was produced at a steady pace, irrespective the presence of oxygen, to form the ATP needed for cell functions. Glycerol efflux increased over time and its origin shifted from glycolitic to glycolytic-lipolytic: new-formed glycerol was incorporated into TAG, by esterification with acyl-CoA, derived from the same TAG lipolisis. The coexistence of these processes appears as a “futile cycle”, with the probable function further to waste excess energy. Lipogenesis was limited because the size of cells limits the access to oxidative mitochondrial pathways. The release of 3C metabolites seems to be a mechanism to lower glycemia, defend WAT against excess of substrate, and provide 3C fragments as more accessible substrates for other tissues. Mesenteric WAT adipocytes presented the highest metabolic activity, probably to help the hepatic handling of NEFA and reduce the flow of intestinal glucose to the liver. While in almost all WAT sites, excess mitochondrial pyruvate is returned to the cytoplasm to keep forming lactate; in female mesenteric adipocytes it is in part oxidized to acetyl-CoA to fuel lipogenesis. Stromal vascular cells also released lactate, even more than adipocytes per unit of tissue weight, but not glycerol nor NEFA. Red blood cells produced lactate, but its contribution was quantitatively minimal. Thus, stromal cells, acted in consonance with adipocytes, in all sites and sexes examined, wasting glucose in an anaerobic way, producing high amounts of 3C units. Thus, reinforcing the idea that WAT may be an active protagonist both in energy handling and in the body control of glycemia., [spa] Las teorías actuales sobre la inflamación endotelial en el tejido adiposo en el síndrome metabólico apuntan a la hipoxia como una de sus causas. Nuestro grupo ha encontrado que adipocitos 3T3L1, bajo condiciones normóxicas, consumen grandes cantidades de glucosa, con elevada producción de lactato y glicerol. La hipótesis de esta tesis es que los adipocitos actuan como células glucolíticas y su metabolismo ayuda a reducir la glucemia, mientre la hipoxia podría afectar las células estromales. Hemos estudiado ex vivo la capacidad glucolítica de adipocitos adultos y células estromales de ambos sexos y de diferentes ubicaciones del TAB, desarrollando la metodología para un análisis cuantitativo. Hallamos que los adipocitos eran las células menos númerosas del tejido y la parte "viva" representaba solo el 1,5% del total, mostrando así el TAB una actividad metabólica muy elevada. Los adipocitos ex vivo captaban grandes cantidades de glucosa, independientemente de su concentración; liberando metabolitos de 3C. El lactato, liberado a ritmo constante y independientemente de la presencia de oxígeno, procedía totalmente de la glucosa. El eflujo de glicerol aumentó con el tiempo y su origen cambió de glucolítico a glucolítico-lipolítico. El glicerol neoformado se incorporaba a los TAG, reciclado acil-CoA de la misma lipolisis. Este"ciclo fútil", probablemente sirve para desperdiciar e energía, disminuir la glucemia, y proporcionar sustratos energéticos para otros tejidos. Los adipocitos del TAB presentaban la más alta actividad metabólica; probablemente para ayudar al manejo hepático de ácidos grasos y reducir el flujo de glucosa intestinal. Mientras que en todas las localizaciones, el exceso de piruvato mitocondrial regresaba al citoplasma y seguía formando lactato; en los adipocitos de las hembras de mesentérico, parte se oxidaba a acetil-CoA para sostener la lipogénesis. Las células estromales, por unidad de tejido, liberaban más lactato que los adipocitos, pero no glicerol ni ácidos grasos. Los eritrocitos produjeron mínimas cantidades de lactato.Así las células estromales, actúan en consonancia con los adipocitos, en todas las localizaciones estudiadas y en ambos sexos. Estos resultados refuerzan la idea de que el TAB puede ser un protagonista activo en el manejo de la energía y en el control de la glucemia.

[eng] Current lifestyle with high-energy diets and characterized by sedentary is triggering an alarming growth in obesity. Obesity along with metabolic syndrome- related co-morbidities (i.e. insulin resistance, atherosclerosis, sleep apnea, depression, asthma, hypertension and the alteration of blood lipid transport) are the most apparent consequence of the excess energy. Under conditions of excess dietary energy, the body cannot easily dispose of the excess amino-N against the evolutively-adapted schemes that prevent its wastage; thus ammonia and glutamine formation and urea excretion are decreased. High lipid and energy availability limit the utilization of glucose, and high glucose spares the production of ammonium from amino acids, decreasing the synthesis of glutamine and its utilization by the intestine and kidney. In contrast, high protein diets enhance protein synthesis and growth, and the synthesis of non-protein-N-containing compounds. But these outlets are not enough; consequently, less- conventional mechanisms are activated, such as increased synthesis of NO∙ followed by higher nitrite (and nitrate) excretion and changes in the microbiota. In this study we studied how the initial phase of development of metabolic syndrome can affects the function of liver as main site of amino-N metabolism, and to determine whether doubling the protein content in the diet induced significant changes in enzyme of amino acids metabolism along intestine and on liver. The common result obtained by these studies is that, both in case of hyperlipidic or hyperproteic diets, elimination of excess N is necessary but cannot be easily carried out through the metabolic pathways/tissues we evaluated, although possible alternative pathways have been taken into consideration., [cat] L’estil de vida actual amb les dietes d'alt contingut energètic, i caracteritzat pel sedentarisme, està provocant un creixement alarmant de l'obesitat. L'obesitat, juntament amb les comorbiditats relacionades amb la síndrome metabòlica (és a dir, resistència a la insulina, aterosclerosi, apnea del son, depressió, asma, la hipertensió i l'alteració del transport de lípids en la sang) són la conseqüència més evident de l'excés d’energia. En condicions d'excés d'energia de la dieta, el cos no pot eliminar ràpidament l'excés d'amino-N contra els esquemes adaptats evolutivament i que impedeixin el seu deteriorament; així, la formació d'amoníac i de glutamina i l'excreció d'urea disminueixen. Els elevats nivells de lípids i de la disponibilitat d'energia limiten la utilització de la glucosa, i nivells elevats de glucosa estalvia la producció d'amoni a partir dels aminoàcids, disminuint la síntesi de glutamina i la seva utilització per l'intestí i el ronyó. En contrast, les dietes d’elevat contingut en proteïnes incrementen la síntesi de proteïnes i el creixement, i la síntesi de compostos que contenen N i no són proteïnes. Però aquests mecanismes no són suficients i en conseqüència, s'activen mecanismes menys convencionals, com ara augment de la síntesi de NO ∙ seguides per l’augment del nitrit (i nitrat) i la seva excreció, juntament amb canvis en la microbiota. En aquest treball es va estudiar com la fase inicial de desenvolupament de la síndrome metabòlica pot afectar la funció del fetge com lloc principal del metabolisme d'amino-N, i per determinar si la duplicació del contingut de proteïnes en la dieta induïa canvis significatius en els enzims del metabolisme d'aminoàcids al llarg intestí i al fetge. El resultat genèric obtingut per aquests estudis és que, tant en el cas de que la dieta sigui hiperlipídica o hiperproteica, l'eliminació de l'excés de N és necessària, però no es pot dur a terme fàcilment a través de les vies metabòliques / teixits que avaluem, tot i les possibles vies alternatives s'han tingut en consideració.

Aunque se ha caracterizado extensamente la eficacia de la administración de la oleoil estrona sobre la pérdida de grasa corporal, todavía no se ha conseguido averiguar su mecanismo de acción. Esta tesis doctoral se ha centrado en el estudio de la acción de la oleoil estrona a nivel periférico, utilizando ratas Wistar macho con sobrepeso (previamente alimentadas con una dieta de cafetería), y en un modelo de restricción energética forzada equivalente (pair fed), con el fin de discernir los efectos específicos de la oleoil estrona sobre vías metabólicas, partición de la energía y regulación hormonal. Los resultados obtenidos indican que la administración de oleoil estrona junto con una restricción energética adicional no potencia su efecto sobre la pérdida de grasa corporal, aunque se induce la movilización de proteína corporal. Por otro lado, tanto la disminución de las reservas de grasa producidas por el tratamiento con oleoil estrona como el perfil de expresión génica relacionado con el metabolismo lipídico en el tejido adiposo, parecen ser consecuencia, en buena parte, de la disminución de la ingesta. El hígado es, probablemente, el tejido que marca las diferencias en el metabolismo energético tras el tratamiento con oleoil estrona. La acción de la oleoil estrona en el hígado da lugar a cambios profundos y casi siempre en sentido contrario al observado en las ratas pair fed. Concretamente, la administración de oleoil estrona, a pesar de la diminución de la ingesta, disminuye la expresión hepática de los genes implicados en la gluconeogénesis, mantiene la de los relacionados con la glucolisis y la lipogénesis e incluso incrementa la expresión de los implicados en la síntesis de triacilgliceroles, de acuerdo con el incremento del factor de transcripción SREBP-1c, lo que diferencia claramente la acción de la oleoil estrona de la simple restricción energética. Estos resultados explican en parte el mantenimiento de la glucemia y el aumento del consumo de triacilgliceroles por parte de los órganos periféricos. Además, el tratamiento con oleoil estrona incrementa en el hígado la expresión hepática de genes y niveles de proteínas que participan en el transporte reverso de colesterol y en su oxidación a ácidos biliares. Estos efectos están de acuerdo con la disminución del colesterol circulante que se observa tras el tratamiento con oleoil estrona. El efecto adelgazante de la administración de la oleoil estrona se ve limitado por el efecto contraregulador de los glucocorticoides. Los resultados obtenidos indican que las ratas tratadas con oleoil estrona aumentan tanto la síntesis de glucocorticoides en las glándulas adrenales como su eliminación en el hígado, lo que modera el incremento de glucocorticoides circulantes, a diferencia del grupo pair fed, sometido a una restricción energética forzada. La hidrólisis de la oleoil estrona libera estrona, un estrógeno débil pero fácilmente activable a estradiol. La sobrecarga de estrona producida tras la administración de la oleoil estrona inhibe su activación de estradiol, lo que limita los efectos estrogénicos, pero a la vez se inhibe la síntesis y la activación de testosterona, con lo que se produce una marcada disminución de la testosterona circulante. En definitiva, el tratamiento con oleoil estrona disminuye la grasa corporal como consecuencia del incremento de la sensación de saciedad, pero a diferencia de una simple restricción energética mantiene la glucemia y la síntesis de lípidos hepáticos, a la vez que limita el incremento de glucocorticoides circulantes, lo que da lugar a la moderación del nivel de estrés de los animales., This thesis has been focused on the study of the action of oleoyl estrone using overweight male Wistar rats, and a model of restricted energy intake (pair fed), in order to discern the specific effects on metabolic pathways of oleoyl estrone. The decrease in fat reserves produced by treatment with oleoyl estrone as suggests by the expression profile of genes associated with lipid metabolism in adipose tissue seems to be due, largely, to the decrease in food intake. The oleoyl estrone treatment results in profound changes in liver metabolism. In spite of decreased food intake, administration of oleoyl estrone decreases the hepatic expression of genes involved in gluconeogenesis, maintaining those of glycolysis and lipogenesis; there are, even, an increase in those of triacylglycerol synthesis. This is in agreement with SREBP-1c increase, a condition which clearly differentiates the action of oleoyl estrone from pair fed energy restriction. These results explain at least partly the maintenance of blood glucose and the increased consumption of triacylglycerols by peripheral organs. Treatment with oleoyl estrone also increases the hepatic expression of genes involved in reverse cholesterol transport and its oxidation to bile acids. These effects are consistent with the marked decrease of circulating cholesterol observed under treatment with oleoyl estrone. Rats treated with oleoyl estrone increase both glucocorticoid synthesis in the adrenal glands and their oxidative inactivation in the liver, thereby moderating the increase in circulating glucocorticoids, unlike the pair fed group, subjected to a non voluntary restriction in energy intake. The estrone overload produced after the administration of the oleoyl estrone inhibits its own activation to estradiol, thereby limiting the overall estrogenic effects. However, this process also inhibits the synthesis and activation of testosterone, resulting in a decrease its circulating levels. In conclusion, treatment with oleoyl estrone decreases body fat due to a higher level of satiety, but unlike pair fed energy restriction, blood glucose and hepatic lipid synthesis are maintained, and the increase of circulating glucocorticoids is limited, lowering the level of stress endured by the rats.

La finalitat d'aquest treball ha estat continuar caracteritzant els efectes aprimadors de l'oleoïl-estrona (OE), aprofundint en el coneixement del seu mecanisme d'acció, i de com els seus efectes sobre la ingesta, la composició corporal i la millora de la sensibilitat a la insulina, poden ser modulats al ser combinada amb altres fàrmacs, comprovant especialment la seguretat d'aquestes combinacions.El nostre grup de treball, basant-se en el model homeostàtic postulat per Kennedy per explicar la complexitat dels mecanismes de regulació del pes corporal, va descobrir l'oleoïl-estrona, una molècula que indueix una pèrdua de pes de manera dosi depenent deguda a la disminució de la ingestió d'aliment mantenint la despesa energètica i facilitant així la mobilització de greixos. L'OE va ser proposada com a possible senyal ponderòstat i com a possible base d'una teràpia farmacològica per a l'obesitat.En un primer objectiu es pretenia demostrar si els efectes de la OE sobre el balanç energètic eren mitjançats a través dels receptors d'estrògens, pel que es va realitzar un estudi en el que s'administrava un ampli rang de dosis d'estrona lliure comprovant la diferència d'efectes metabòlics existents entre aquesta i l'OE. Continuant amb la mateixa línia, es va realitzar una administració conjunta de l'OE amb el tamoxifè, un antagonista dels receptors d'estrògens àmpliament utilitzat, el que ens permetria confirmar que el tamoxifè no evita els efectes aprimadors de l'OE, suggerint que el bloqueig d'aquests receptors no impedeix l'acció de l'OE, i per tant, que aquesta no actua a través de les vies estrogèniques.En un segon objectiu es va combinar l'OE amb altres agents aprimadors (sibutramina, dexfenfluramina, fentermina, un agonista dels receptors beta-3-adrenèrgics (CL316,243), i rimonabant), fet que ens va permetre caracteritzar una possible teràpia farmacològica combinada efectiva i segura per al tractament de l'obesitat i aproximar-nos de manera indirecta al coneixement del mecanisme d'acció de l'OE.En un tercer objectiu, mitjançant la combinació de l'OE amb la rosiglitazona es va estudiar els mecanismes a través dels quals aquests dos compostos milloren la sensibilitat a la insulina. Malgrat que la rosiglitazona i l'OE presenten efectes metabòlics pràcticament oposats, la seva combinació es presenta con una bona alternativa per al tractament de la diabetis mellitus de tipus 2 en els pacients que presenten obesitat.Finalment, per tal d'aprofundir en els efectes moduladors de l'OE sobre la ingesta, es van analitzar els efectes d'aquesta sobre l'expressió dels principals pèptids gastrointestinals que regulen la ingestió d'aliments a curt termini. Es va comprovar com l'administració d'OE provoca una ràpida i marcada inhibició de l'expressió del pèptid orexigènic ghrelina, amb efectes pràcticament nuls sobre l'expressió dels pèptids anorexigènics CCK, PYY i GLP-1., SUMMARU:The aim of this work was to continue the characterization of the the slimming effects of oleoyl-estrone (OE) studying its mechanism of action and whether its effects on food intake, energy balance and insulin sensitivity could be modulated by its combination with other drugs. Possible synergisms and risks of the combination of OE with known drugs were studied. Our research group discovered OE, a molecule synthesized in white adipose tissue that elicits a marked dose-dependent loss of body weight. OE treatment reduces food intake while maintaining energy expenditure thus creating an energy gap filled by the expense of body fat stores. OE was proposed as a possible ponderostat signal and a possible agent for obesity treatment. Initially, we focused our study in determining if OE's effects were mediated by the estrogen receptor. First, a wide range of estrone doses were tested in order to evaluate differences between estrone and OE on the energetic balance. Later, we studied the combined effects of OE and tamoxifen, a estrogen receptor antagonist, in order to prove that OE does not act through estrogenic pathways. Tamoxifen does not prevent the mobilization of body lipids elicited by OE.Secondly, OE was combined with other drugs regularly used to treat obesity (sibutramine, dexfenfluramine, phentermine, a specific b3-adrenergic receptor agonist (CL316,243) and rimonabant), in order to characterize an effective and safe pharmacological therapy. This set of experiments also provided insights regarding the mechanism of action of OE.Afterwards, we combined OE and rosiglitazone in order to study how these two agents improve insulin sensitivity. Despite their diverging effects on body mass fat, the combination of OE and rosiglitazone may be positive for type 2 diabetics that are obese or show a marked predisposition to the accumulation of fat.Finally, in order to elucidate the effects of OE on food intake we analyzed the expression of several gut peptides that affect short-term feeding behavior. Results showed that the ghrelin expression was markedly reduced by OE administration. However its effects on the anorexigenic peptides studied (CCK, GLP-1 and PYY) are practically non-existent.

Aquest treball té com a objectiu principal l'estudi dels mecanismes de control del pes corporal, objectiu que es concreta en l'estudi de les causes bioquímiques i fisiològiques de l'obesitat. Per tal de dur a terme aquest treball es van utilitzar animals de la soca Wistar i de la soca Zucker; aquesta última soca es caracteritza per presentar obesitat genètica i es van utilitzar grups d'animals prims i obesos (rata Zucker fa/fn). Igualment, per tal d'estudiar un model d'obesitat induida per la dieta es va utilitzar una dieta de "cafeteria" i per estudiar la ingesta es va mantenir un grup d'animals en gàbies metàl.liques dels trenta als seixanta dies d'edat. Per altra banda un dels termes que més ens va interessar de quantificar són les pèrdues d'energia en forma de calor, que van ser mesurades gràcies al desenvolupament d'un aparell de calorimetria directe, així com un de calorimetria indirecte d'anàlisi de gasos; aquestes mesures de calorimetria es realitzaven al final del període estudiat (59-60 dies d'edat).Es van realitzar mesures de les pèrdues de calor mitjancant calorimetria directa en condicions basals, després de l'administració intragàstrica de sacarosa i havent administrat sacarosa, però després de dotze hores de privació d'aliments. Es va poder comprovar que les rates obeses presenten unes pèrdues de calor (expressades en W+pes(4)) més baixes que les rates primes (Zucker o Wistar) i que l'administració de sacarosa promou un augment de la despesa, si més no en les rates Wistar. Pel que fa a la privació d'aliments, va mostrar un efecte molt marcat de disminució de les pèrdues de calor en els dos grups de rates primes, mentre que aquest efecte era pràcticament nul per les rates obeses, indicant que la producció de calor en les rates obeses ja es troba a uns nivells prou baixos, de manera que ni la manca d'aliments pot fer que disminuexi més. Aquest va ser un estudi de l'efecte de l'administració aguda de sacarosa. Per veure si l'administració crònica tenia altres efectes es van tractar els animals amb dosis diàries de sacarosa durant deu dies, observant-se que no es donava un efecte significatiu al llarg del temps i confirmant-se una menor producció de calor en les rates genèticament obeses. Amb aquest mateix plantejament es va estudiar l'augment de pes i de deposició de lípids en aquests animals, comparant-lo amb animals control i animals sotmesos a dieta de cafeteria: es va observar que en cap cas l'administració de sacarosa promovia increments de pes més alts que els dels altres tractaments i l'efecte sobre la deposició de nutrients era variable segons la soca. El mateix podem dir pel que fa a la seva potencialitat termogènica, perquè en aquest cas no ha provocat una despesa energetica més gran. En canvi, l'administració de la dicta de cafeteria, contrariament a l'obesitat genètica, sí va donar una producció de calor més gran. L'energia ingerida i emmagatzemada pels animals mantinguts en gabies metabòliques va ser més alta en els animals alimentats amb dieta de cafeteria i en els animals obesos, mostrant-se un augment de l'eficiència de deposició en ambdós tipus d'obesitat.Per altra banda, l'anàlisi dels ritmes de producció de calor en rates Wister va posar de manifest l'existència de ritmes circadians, de ritmes de vuit, dues i mitja hora. L'estudi paral.lel dels ritmes de variació de la temperatura nuclear, mesurada mitjançant la implantació crònica de termoparells a l'aorta, va indicar que aquests ritmes eren comparables als anteriors, però que també es donaven ritmes de quatre i cinc hores, indetectables en les mesures de pèrdues de calor i que aquests paràmetres estaven correlacionats, però no eren totalment coincidents, existint un desplaçament d'una mitja hora.Pel que fa a la utilització de les tècniques de calorimetria indirecta, l'anàlisi de gasos ens va permetre mesurar l'aigua evapotranspirada, i a partir de les dades, també de calorimetria indirecta, dels substrats oxidats, l'aigua metabòlica produïda. Això ens va permetre completar l'estudi del balanç d'aigua en les tres soques estudiades. Les rates de la soca Wistar van ingerir més aigua, com a tal, i van presentar unes pèrdues per orina més grans que les rates de la soca Zucker, de manera que en les primeres donava lloc a una orina molt més diluida. En canvi, les perdues d'aigua amb les femtes van ser més grans en les rates Zucker obeses, de manera que la relació de grams de sòlids en femta a grams de sòlids ingerids era més alta en aquests animals, suggerint una pèrdua d'eficiència digestiva relacionada amb aquest tipus d'obesitat.El fet d'utilitzar la calorimetria indirecta per l'estudi de l'obesitat ens va fer plantejar si les equacions que s'utilitzen habitualment eren directament aplicables als nostres estudis. Per tal d'esbrinar això, a partir de les dades de composició en àcids grassos dels lípids, vam poder deduïr l'estructura del triacilglicerol mig corresponents als lípids ingerits, acumulats, oxidats i els que es troben en el cos dels animals, i de la mateixa manera pels aminoàcids constituents de les proteines. A partir d'aquests dos grups de dades vam deduïr les equacions que s'apliquen a les mesures de calorimetria indirecta, les quals presenten la mateixa estructura bàsica que les descrites per altres autors (Ferrannini i Brouwer), amb la salvedat que en les que nosaltres proposem no s'utilitza el nitrogen urinari com a mesura de la proteina oxidada perquè, segons dades del nostre propi grup de recerca, existeix una part significativa del nitrogen provinent de l'oxidació proteica que no es troba com a pèrdues de nitrogen en l'orina. Aquestes equacions van ser validades comparant els seus resultats amb els obtinguts amb les equacions d'altres autors i amb l'estudi del balanç de substrats. La seva aplicació a les dades de consum d'oxígen i producció de diòxid de carboni al llarg del temps ens va permetre conèixer el patró d'utilització de substrats al llarg del dia. Vam obserusr que mentre que les rates Wister presenten un màxim d'oxidació de glúcids amb síntesi de lípids dins el període fosc, les rates de la soca Zucker utilitzen glúcids per a sintetitzar lípids la major part del dia, consumint part d'aquests durant la nit. Això ens suggereix uns hàbits més diürns per les rates Zucker en comparació als nocturns, tipics de la rata Wister. També es va estudiar el balanç d'àcids grassos en els diferents grups, que va indicar que la composició en àcids grassos dels lípids corporals de la rata segueix un patró molt uniforme, independent de la soca, l'estat d'obesitat i fins i tot del tipus de dieta, amb una predominancia clara de 1'àcid palmític i l'oleic, per tant els efectes d'ambdós models d'obesitat són més de tipus quantitatiu que qualitatiu. E1 patró d'ingesta en àcids grassos era molt semblant per ambdues dietes, però les rates primes mantingudes en dieta control sintetitzaven una part dels àcids grassos, mentre que les mantingudes en dieta de cafeteria n'oxidaven. En canvi les rates obeses seguien sintetitzant alguns àcids grassos malgrat una ingesta ja elevada de greixos.Això també es va observar mitjançant la utilització de la calorimetria indirecta en animals sotmesos a diferents temperatures, de manera que les rates obeses i les rates Wistar mantingudes a 10 ºC mostren síntesi de lípids, un procés que té una eficiència del 75%, perdent-se la resta en forma de calor, de manera que mentre que les rates primes mobilitzen les seves reserves per fer front al fred a temperatures extremes, les rates obeses i les Wistar en fred moderat utilitzaven la síntesi de lípids com a mecanisme de generar calor. Però aquest procés funcionaria a totes les temperatures en els animals obesos, de manera que mantenen una ingesta elevada a canvi de tenir cert control sobre la seva producció de calor, fet que podria explicar perquè són hiperfàgics i perquè esdevenen obesos., This work deals with the biochemical and physiological causes of obesity within a line of research studying body weight regulation. A series of methods have been developed to measure heat production and output, by means of direct and indirect (gas analysis) calorimetry. These techniques have been applied to the study of animal models of genetic obesity (Zucker fa/fa rat), and diet-induced obesity (use of a cafeteria diet), as well as their additive interaction. We have found that energy deposition efficiency was higher in both obesity models compared with controls; their combined effect was not completely additive, since the cafeteria diet had a less marked effect on obese rats than in lean controls. Sucrose administration had different effects on heat production depending on the rat strain, with undetectable effects as to body weight change. Wistar rats showed distinct heat production rhythms, which were partially correlated (but displaced in time) with core temperature rhythms. Fatty acid composition patterns of rat lipids were very uniform desipite of diet and genetic obesity situations. In the rat maintained at different ambient temperatures, thermic homeostasis took preeminence over energy homeostasis. Genetically obese rats actively synthesize lipids from carbohydrate in order to produce residual heat and maintain their thermic equilibrium. Non-obese rats also showed this trend when exposed to 10 ºC, but their normal operation of thermogenesis from lipids precluded their accumulation of fat; the fa/fa rats deficient thermogenesis could not eliminate this fat and they store it becoming obese.

El tejido adiposo marrón (TAM) está considerado como el principal efector de la termogénesis no temblorosa inducida por el frío y de la termogénesis facultativa inducida por la dieta en la rata y otras especies de mamíferos.Debido a su importancia en estos procesos; al metabolismo del TAM ha sido muy estudiado, si bien los aspectos relacionados con el metabolismo de los aminoácidos han recibido muy poca atención. El objetivo del presente estudio ha sido determinar las características generales del metabolismo de los aminoácidos en el TAM interescapular (TAMI) de la rata en distintas situaciones fisiológicas, habiendo sido dividido en tres bloques experimentales:1) Estudio de las actividades enzimáticas relacionadas con el metabolismo de los aminoácidos en el TAMI.2) Estudio de la utilización de la alanina por el TAMI «in vitro».3) Estudio de la captación/liberación de aminoácidos y glucosa por el TAMI «in vivo».En la primera parte se han estudiado las actividades de ocho de los principales enzimas en el metabolismo de los aminoácidos. No se actividades detectables de los enzimas serina deshidratasa arginasa. Los otros enzimas estudiados han sido la transaminasa (GPT), aspartato transaminasa (CiO"I'), transaminasa de aminoácidos de cadena ramificada (BCT), glutamina sintetasa (GST), glutamato deshidrogenasa (GDH) y adenilato desaminasa (AMD); para todos ellos las actividades en el TAMI son mayores que las observadas en el tejido adiposo blanco, y comparables a las del hígado y músculo esquelético. El ayuno induce pocos cambios en dichas actividades, mientras que la exposición aguda al frío determina un incremento en la BCT; por su parte, la exposición crónica al frío induce un notable incremento en la BCT y también en la GDH, mientras que disminuye la actividad AMD. Por otra parte, el acervo de aminoácidos libres del TAMI muestra variaciones en función de la situación fisiológica del animal. De este modo, el ayuno y la exposición aguda al frío inducen una disminución en la concentración de los aminoácidos tisulares; en los animales expuestos al frío de forma crónica la concentración total de aminoácidos no se modifica, aunque es en este grupo donde las variaciones individuales con respecto a los controles son más numerosas. Al relacionar estas concentraciones con el aminograma sanguíneo destacan las variaciones observadas en el grupo de aminoácidos ramificados cambios que parecen estar de acuerdo con una utilización activa de los mismos en situaciones de exposición aguda o crónica al frío. En cualquier caso, se verifica que dichos cambios no son debidos a un recambio proteico especialmente rápido en el TAMI.En la segunda parte se ha procedido a estudiar la utilización de la alanina por el TAMI «in vitro» en un modelo de incubaciones de fragmentos de tejido. Primeramente se ha determinado la validez de la técnica, para lo que se ha comprobado que la relación ATP/ADP se mantiene constante para incubaciones de hasta una hora de duración, al igual que la linealidad respecto al tiempo en la incorporación de los carbonos del aminoácido a las distintas fracciones tisulares estudiadas, siendo los destinos más importantes de la alanina su oxidación hasta CO2 y su incorporación a los ácidos grasos del tejido.En un primer experimento se ha determinado el efecto de la glucosa sobre la utilización de la alanina, a distintas concentraciones de ambos substratos, comprobándose un efecto estimulador de la glucosa sobre la incorporación de los carbonos de la alanina al tejido, aunque no se modifica la oxidación del aminoácido. En el experimento complementario, en el que se ha estudiado el efecto de la alanina sobre la utilización de la glucosa, se verifica que para concentraciones superiores a 3 mM de glucosa, la alanina disminuye la oxidación de este substrato aunque sin afectar su oxidación.Se ha comprobado también el efecto de la insulina, la noradrenalina y el glucagón sobre la utilización de la alanina por el TAMI, observándose que la primera hormona tiene un claro efecto estimulador sobre la utilización de la alanina, mientras que las otras dos disminuyen dicha utilización, si bien debe señalarse que en el caso del glucagón este efecto tan sólo se verifica a concentraciones por encima del rango fisiológico. Cabe señalar, asimismo, que todos estos efectos tan sólo se verifican en el caso de que la glucosa esté presente en el medio de incubación, mientras que en ausencia de la misma son menores tanto la oxidación del aminoácido como sobre todo su incorporación al tejido.Una vez verificados todos estos aspectos, se ha estudiado la utilización de la alanina «in vitro» por el TAMI procedente de animales sometidos a distintas situaciones fisiológicas, tanto en condiciones basales corno en presencia de insulina y/o noradrenalina. El TAMI procedente de los animales sometidos a frío agudo muestra la mayor utilización de alanina, mientras que el de los sometidos a ayuno o a una exposición crónica al frío presenta una menor utilización que el tejido de los animales del grupo control.En el tercer apartado experimental se ha determinado la captación o liberación de los distintos aminoácidos y de la glucosa por parte del TAMI en distintas situaciones fisiológicas. Se ha utilizado a tal efecto la técnica de la determinación de las diferencias arteriovenosas para cada uno de dichos substratos metabólicos, a partir de sus concentraciones en sangre arterial (aorta abdominal) y sangre venosa (vena de Sulzer). La similitud en la concentración de aminoácidos en la sangre arterial de los animales de los diferentes grupos experimentales indica que las diferencias observables deben ser atribuidas a un factor distinto de su asequibilidad. Por otra parte, se han determinado los flujos sanguíneos a través del TAMI utilizando la técnica de las microesferas radiactivas, marcadas con (46)Sc; el flujo se presenta disminuido en los animales ayunados pero sumamente elevado en los animales expuestos de forma aguda o crónica al frío, de acuerdo con la mayor actividad termogénica descrita en estas situaciones.En los animales alimentados tan sólo se observa una liberación neta de prolina y taurina, así como una tendencia global hacia una pequeña liberación de aminoácidos. Con el ayuno el TAMI pasa a exportar alanina, glutamina, glicina y citrulina, así como el conjunto de aminoácidos no esenciales. Sin embargo el frío agudo induce una clara tendencia a la captación de aminoácidos, especialmente de los esenciales como la isoleucina, si bien el tejido continúa exportando prolina. Con la exposición crónica al frío hay una liberación muy importante de alanina, arginina y taurina, mientras que sigue captando isoleucina y también glutamina. En conjunto los resultados parecen indicar que este tejido capta activamente aminoácidos únicamente ante una estimulación aguda por el frío, pero estos compuestos no deben ser importantes substratos para el TAMI en otras situaciones. Sin embargo, los aminoácidos ramificados son utilizados tanto en una situación de estimulación aguda como durante la aclimatación al frío, por lo que podrían considerarse como un importante substrato energético de este tejido.Este estudio «in vivo» se ha hecho extensivo para el caso de la glucosa, considerada como un importante substrato metabólico del TAMI. Su utilización por este tejido es especialmente notable tanto en los animales expuestos de forma aguda al frío como sobre todo en los aclimatados al frío, confirmándose de esta manera su importante papeltermogénico.Los resultados obtenidos en los distintos bloques experimentales realizados en la presente tesis doctoral sugieren que el TAMI utiliza algunos aminoácidos como la alanina y los ramificados como substratos en situaciones de emergencia, como puede ser una exposición aguda al frío, si bien en general no parecen contribuir de forma significativa, en otras situaciones a la capacidad termogénica de este tejido., Interscapular brown adipose tissue (IBAT) presented high activities of some enzymes related with aminoacid metabolism such as alanine, aspartate and branched chain amino acid transaminases, glutamate dehydrogenase, glutamine synthetase and adenylate deaininase, but neither arginase nor serine dehydratase were detectable. These enzymatic activities were in a range between those of liver and skeletal muscle. Chronic cold-exposure induced some changes both in the tissue enzymatic activities and amino acid pool composition, but starvation or acute cold-exposure did not induce deep effects.IBAT took up actively alanine and glucose, in "in vitro" incubations of tissue slices. Insulin enhanced the utilization of alanine, but noradrenaline and glucagon decreased it. These hormones had effect only when glucose was present in the incubating media. The IBAT of rats subjected to acute cold-exposure presented an active utilization of alanine, but the capacity of the starved or chronically cold-exposed rat tissue was decreased with respect to the control animals."In vivo" utilization of amino acids and glucose was estimated through the measurement of arterio-venous differences having into account the blood flow (measured by means of radiactive microspheres). The results indicated that IBAT took up some amino acids, as glutamine, and released glycine and proline, maintaining its amino acid nitrogen balance. Branched chain amino acids seem to play an important role as energetic substrates of IBAT under conditions of tissue stimulation, as cold-exposure; this is also true for glucose uptake.The results obtained in the present work show that IBAT can utilize some amino acids as alanine or branched chain amino acids for energetic purposes, especially under situations of acute cold-exposure.

[spa] Se han estudiado los niveles de aminoácidos individuales (lactato, glucosa, glicerol, etc.) en plasma y actividad de GLU-deshidrogenasa, ASP-transaminasa, ALA-transaminasa, TYR-transaminasa, GLN-sintetasa, SER-deshidratasa, AUP-desaminasa y arginasa en hígado, musculo estriado, riñón, intestino, estómago, tejido adiposo, encéfalo y piel de ratas durante la lactancia y después de destetar las crías. El metabolismo nitrogenado tiende al ahorro de los aminoácidos durante la lactancia con el fin de que el organismo materno los pueda incorporar al metabolismo de las proteínas de la leche. El ayuno de las ratas lactantes provoca una demanda más intensa sobre las reservas de la madre que sobre los controles vírgenes.