Eduardo Moltó

Central Leptin Regulates Total Ceramide Content and Sterol Regulatory Element Binding Protein-1C Proteolytic Maturation in Rat White Adipose Tissue

Obesity and type 2 diabetes are associated with insulin and leptin resistance, and increased ceramide contents in target tissues. Because the adipose tissue has become a central focus in these diseases, and leptin-induced increases in insulin sensitivity may be related to effects of leptin on lipid metabolism, we investigated herein whether central leptin was able to regulate total ceramide levels and the expression of enzymes involved in ceramide metabolism in rat white adipose tissue (WAT). After 7 d central leptin treatment, the total content of ceramides was analyzed by quantitative shotgun lipidomics mass spectrometry. The effects of leptin on the expression of several enzymes of the sphingolipid metabolism, sterol regulatory element binding protein (SREBP)-1c, and insulin-induced gene 1 (INSIG-1) in this tissue were studied. Total ceramide levels were also determined after surgical WAT denervation. Central leptin infusion significantly decreased both total ceramide content and the long-chain fatty acid ceramide species in WAT. Concomitant with these results, leptin decreased the mRNA levels of enzymes involved in de novo ceramide synthesis (SPT-1, LASS2, LASS4) and ceramide production from sphingomyelin (SMPD-1/2). The mRNA levels of enzymes of ceramide degradation (Asah1/2) and utilization (sphingomyelin synthase, ceramide kinase, glycosyl-ceramide synthase, GM3 synthase) were also down-regulated. Ceramide-lowering effects of central leptin were prevented by local autonomic nervous system denervation of WAT. Finally, central leptin treatment markedly increased INSIG-1 mRNA expression and impaired SREBP-1c activation in epididymal WAT. These observations indicate that in vivo central leptin, acting through the autonomic nervous system, regulates total ceramide levels and SREBP-1c proteolytic maturation in WAT, probably contributing to improve the overall insulin sensitivity.

Isolation and biological characterization of a 6-kDa protein from hepatopancreas of lobster Panulirus argus with insulin-like effects

A protein with insulin-like effects was isolated from the hepatopancreas of the lobster Panulirus argus following a classic method for mammalian insulin purification from the pancreas. After acid-alcoholic extraction and ethanol-ether precipitation followed by molecular filtration chromatography, a protein with an apparent molecular weight of 6 kDa was isolated. This protein is characterized by its ability to interact with anti-insulin antibodies and by mimicking insulin actions as the stimulation of glucose oxidation to CO(2) and lipogenesis in isolated rat adipocytes. In addition, this insulin immunoreactive protein (IIP) was able to stimulate the autophosphorylation of the insulin receptor present in rat adipocyte plasma membranes, in a dose-dependent manner. The immunological and biochemical results obtained are consistent with the hypothesis that protein(s) with insulin-like effects occur in the digestive gland of the lobster P. argus and may be of significance to control metabolic and growth related processes in crustaceans.

The expression of rat resistin isoforms is differentially regulated in visceral adipose tissues: effects of aging and food restriction.

Two variants of the adipose hormone resistin are generated by alternative splicing in Wistar rats. Here we analyzed the expression of these resistin variants in 2 main visceral adipose depots, epididymal and retroperitoneal, as well as the resistin serum concentration during aging and food restriction. Total protein levels of resistin were also analyzed in extracts from both visceral adipose depots. Resistin variants show similar patterns of relative expression in visceral adipose tissues in 3-month-old rats, representing the short variant, s-resistin, which is 15% of the full-length transcript. However, only epididymal, but not retroperitoneal, fat pad shows a decrease in both messenger RNA and protein levels of resistin isoforms with aging. Food restriction decreases adiposity index in 8- and 24-month-old animals to values even lower than those of 3-month-old animals. Food restriction decreases resistin expression in both adipose tissues in 8-month-old but not in 24-month-old rats. Interestingly, concomitant with the improvement of insulin sensitivity asserted by homeostasis model assessment, resistin serum levels decrease only in food-restricted 8-month-old animals. In contrast, food restriction up-regulates s-resistin messenger RNA in epididymal adipose tissue, whereas no significant changes are appreciated in retroperitoneal adipose tissue. These data indicate that both forms of resistin are differentially regulated by fat depot location, aging, and even nutritional status, suggesting that alternative splicing plays a key role in this differential regulation.

Regulation of Insulin-Stimulated Glucose Uptake in Rat White Adipose Tissue upon Chronic Central Leptin Infusion: Effects on Adiposity

Leptin enhances the glucose utilization in most insulin target tissues and paradoxically decreases it in white adipose tissue (WAT), but knowledge of the mechanisms underlying the inhibitory effect of central leptin on the insulin-dependent glucose uptake in WAT is limited. After 7 d intracerebroventricular leptin treatment (0.2 μg/d) of rats, the overall insulin sensitivity and the responsiveness of WAT after acute in vivo insulin administration were analyzed. We also performed unilateral WAT denervation to clarify the role of the autonomic nervous system in leptin effects on the insulin-stimulated [(3)H]-2-deoxyglucose transport in WAT. Central leptin improved the overall insulin sensitivity but decreased the in vivo insulin action in WAT, including insulin receptor autophosphorylation, insulin receptor substrate-1 tyrosine-phosphorylation, and Akt activation. In this tissue, insulin receptor substrate-1 and glucose transporter 4 mRNA and protein levels were down-regulated after central leptin treatment. Additionally, a remarkable up-regulation of resistin, together with an augmented expression of suppressor of cytokine signaling 3 in WAT, was also observed in leptin-treated rats. As a result, the insulin-stimulated glucose transporter 4 insertion at the plasma membrane and the glucose uptake in WAT were impaired in leptin-treated rats. Finally, denervation of WAT abolished the inhibitory effect of central leptin on glucose transport and decreased suppressor of cytokine signaling 3 and resistin levels in this tissue, suggesting that resistin, in an autocrine/paracrine manner, might be a mediator of central leptin antagonism of insulin action in WAT. We conclude that central leptin, inhibiting the insulin-stimulated glucose uptake in WAT, may regulate glucose availability for triacylglyceride formation and accumulation in this tissue, thereby contributing to the control of adiposity.

Food Restriction is Required to Preserve the Antisteatotic Effects of Central Leptin in the Liver of Middle-Aged Rats: Central Leptin and Fatty Liver in Middle-Aged Rats

Aging is a significant risk factor for the development of obesity and hepatic steatosis associated with insulin and leptin resistance. Food restriction (FR) is commonly used for reducing body weight (BW), adiposity, and liver steatosis. Thus, this study aimed to determine whether FR in middle‐aged rats can recover the central leptin antisteatotic effects observed in the liver in young animals.

S-resistin, a non secretable resistin isoform, impairs the insulin signalling pathway in 3T3-L1 adipocytes

S-resistin is a non-secretable resistin spliced variant, which is expressed mainly in the white adipose tissue from Wistar rats. Previous results confirmed that 3T3-L1 cells expressing s-resistin (3T3-L1-s-res) showed an inflammatory response and exhibited a decrease in glucose transport, both basal and insulin-stimulated. Here we present evidences demonstrating for the first time that s-resistin, like resistin, blocks insulin signalling pathway by inhibiting insulin receptor, insulin receptor substrate 1, protein kinase B/Akt and the mammalian target of rapamycin phosphorylation, and increasing the suppressor of cytokine signalling 3 levels being the later probably due to augmented of leptin expression. Thus, our data suggest that s-resistin could act by a still unknown intracrine pathway as an intracellular sensor, regulating the adipocyte insulin sensitivity.

Changes in Visceral Adipose Tissue Plasma Membrane Lipid Composition in Old Rats Are Associated With Adipocyte Hypertrophy With Aging

Increased adiposity, through adipocyte hypertrophy, and/or hyperplasia, characterizes aging and obesity. Both are leptin-resistant states, associated with disturbed lipid metabolism, reduced insulin sensitivity and inflammation. Nevertheless, fat tissue dysfunction appears earlier in obesity than in normal aging. In contrast, lipodystrophy is accompanied by diabetes, and improving the fat cell capacity to expand rescues the diabetic phenotype. Fat tissue dysfunction is extensively studied in the diet-induced obesity, but remains relatively neglected in the aging-associated obesity. In the Wistar rat, as occurs in humans, early or middle aging is accompanied by an increase in adiposity. Using this experimental model, we describe the molecular mechanisms contributing to the white adipose tissue (WAT) hypertrophy. WAT from middle-old age rats is characterized by decreased basal lipogenesis and lipolysis, increased esterification, as demonstrated by the higher TAG and cholesterol content in visceral WAT, and the maintenance of total ceramide levels within normal values. In addition, we describe alterations in the adipose tissue plasma membrane lipid composition, as increased total ether-phosphatidylcholine, sphingomyelin, and free cholesterol levels that favor an enlarged fat cell size with aging. All these metabolic changes may be regarded as a survival advantage that prevents the aged rats from becoming overtly diabetic.