Adipocyte signaling and lipid homeostasis: sequelae of insulin-resistant adipose tissue

For many years adipose tissue was viewed as the site where excess energy was stored, in the form of triglycerides (TGs), and where that energy, when needed elsewhere in the body, was released in the form of fatty acids (FAs). Recently, it has become clear that when the regulation of the storage and release of energy by adipose tissue is impaired, plasma FA levels become elevated and excessive metabolism of FA, including storage of TGs, occurs in nonadipose tissues. Most recently, work by several laboratories has made it clear that in addition to FA, adipose tissue communicates with the rest of the body by synthesizing and releasing a host of secreted molecules, collectively designated as adipokines. Several recent reviews have described how these molecules, along with FA, significantly effect total body glucose metabolism and insulin sensitivity. Relatively little attention has been paid to the effects of adipokines on lipid metabolism. In this review, we will describe, in detail, the effects of molecules secreted by adipose tissue, including FA, leptin, adiponectin, resistin, TNF-alpha, IL-6, and apolipoproteins, on lipid homeostasis in several nonadipose tissues, including liver, skeletal muscle, and pancreatic beta cells.     

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.     

Effects of fenofibrate on lipid metabolism in adipose tissue of rats

The effect of fenofibrate, a peroxisome proliferator-activated receptor alpha agonist, on body weight gain and on reduction of adipose tissue pads has been ascribed to increased fat catabolism in liver mainly through the induction of target enzymes involved in hepatic lipid metabolism. The aim of this study was to investigate whether peroxisome proliferator-activated receptor alpha activation also affects metabolic pathways in adipose tissue of rats treated with fenofibrate (100 mg/kg body weight) for 9 days. Fenofibrate lowered body weight gain and plasma triglyceride, total cholesterol, and high-density lipoprotein cholesterol but had no influence on food intake and on plasma glucose levels. The activity of lipoprotein lipase of treated animals decreased 50% in epididymal, 29% in retroperitoneal, and was not affected in the mesenteric fat pads. In this study, we show a 34% decrease in epididymal adipose tissue de novo lipogenesis by fenofibrate. The results demonstrate that insulin sensitivity of lipolysis is decreased in fenofibrate-treated rats which resulted in 30% higher rate of glycerol release when compared to the control group. These findings suggest that besides its effects on liver, fenofibrate exerts effects on lipid metabolism in adipose tissue which may contribute to decreasing adiposity.     

Dual effect of the adapter growth factor receptor-bound protein 14 (grb14) on insulin action in primary hepatocytes

Tight control of insulin action in liver is a crucial determinant for the regulation of energy homeostasis. Growth factor receptor-bound protein 14 (Grb14) is a molecular adapter, highly expressed in liver, which binds to the activated insulin receptor and inhibits its tyrosine kinase activity. The physiological role of Grb14 in liver metabolism was unexplored. In this study we used RNA interference to investigate the consequences of Grb14 decrease on insulin-regulated intracellular signaling, and on glucose and lipid metabolism in mouse primary cultured hepatocytes. In Grb14-depleted hepatocytes, insulin-induced phosphorylation of Akt, and of its substrates glycogen synthase kinase 3 and fork-head box protein 1, was increased. These effects on insulin signaling are in agreement with the selective inhibitory effect of Grb14 on the receptor kinase. However, the metabolic and genic effects of insulin were differentially regulated after Grb14 down-regulation. Indeed, the insulin-mediated inhibition of hepatic glucose production and gluconeogenic gene expression was slightly increased. Surprisingly, despite the improved Akt pathway, the induction by insulin of sterol regulatory element binding protein-1c maturation was totally blunted. As a result, in the absence of Grb14, glycogen synthesis as well as glycolytic and lipogenic gene expression were not responsive to the stimulatory effect of insulin. This study provides evidence that Grb14 exerts a dual role on the regulation by insulin of hepatic metabolism. It inhibits insulin receptor catalytic activity, and acts also at a more distal step, i.e. sterol regulatory element binding protein-1c maturation, which effect is predominant under short-term inhibition of Grb14 expression.     

Regulation of adenosine 5',monophosphate-activated protein kinase and lipogenesis by androgens contributes to visceral obesity in an estrogen-deficient state

Menopause is associated with an accumulation of visceral fat. An emerging concept suggests that relatively elevated levels of circulating androgens, compared with estrogens in postmenopausal women, underlie this shift in body fat distribution. In this study we administered dihydrotestosterone (DHT) to ovariectomized mice to examine the effect of relative androgen excess on adipose tissue distribution and function in estrogen-deficient mice. Compared with controls, DHT-treated mice exhibited increased body weight and visceral fat mass associated with triglyceride accumulation. Phosphorylation of AMP-activated protein kinase (AMPK) and acetyl CoA carboxylase was significantly decreased by DHT in visceral fat. In 3T3-L1 cells, DHT decreased phosphorylation of AMPK in a dose-dependent manner. In addition, DHT increased the expression of lipogenic genes (fatty acid synthase, sterol regulatory element binding protein-2, and lipoprotein lipase) in visceral fat. These data provide the first in vivo evidence that an increased androgen to estrogen ratio can promote visceral fat accumulation by inhibiting AMPK activation and stimulating lipogenesis.     

Effects of nutritional state, insulin, and glucagon on lipid mobilization in rainbow trout, Oncorhynchus mykiss.

The effects of nutritional state, insulin, and glucagon on lipid mobilization were determined in rainbow trout, Oncorhynchus mykiss. In nutritional state experiments, fish were either fed continuously (except 24 to 36 hr prior to experimentation) with commercial trout chow or fasted for 4 weeks. Lipase activity in liver tissue isolated from fasted fish and cultured for 5 hr was greater than that in tissue isolated from fed fish and cultured. The presence of glucose (5.55 mM) in the incubation medium accentuates lipolytic activity in both liver and adipose tissue. Hormone response was assessed both in vivo and in vitro. Salmon insulin was injected into anesthetized fish (fed continuously except 24 hr prior to injections) in 10 microliters of saline/g body weight; final hormone dose was 100 ng/g body weight. Tissue and plasma were sampled 1 and 3 hr after injection. Insulin resulted in depressed plasma FA concentration and reduced hepatic triacylglycerol lipase activity. In vitro effects of hormones were evaluated by incubating liver and adipose tissue pieces in Hanks-MEM. Glucagon (bovine/porcine) directly stimulated lipid breakdown in both liver and adipose tissue. These actions were manifested by enhanced FA and glycerol released into the culture medium and by elevated triacylglycerol lipase activity. Insulin (bovine) generally appeared antilipolytic as this agent inhibited glucagon-stimulated lipase activity and glucagon-stimulated FA release. Furthermore, insulin (in the presence of glucose) reduced net lipolysis, as indicated by glycerol release, compared to control cultures. These results indicate that nutritional state and glucose are important modulators of lipid mobilization and that glucagon and insulin act directly on lipid storage sites to coordinate lipolysis in rainbow trout.     

Leptin mRNA表达的组织分布及在大鼠急性肠道损伤中的变化

目的:探讨leptin在急性炎症反应中的作用.方法:采集正常大鼠下丘脑、肺、肝、脾、胃、十二指肠、肾、附睾脂肪垫、睾丸等重要脏器标本,以RT-PCR法检测leptinmRNA表达的组织分布;并建立大鼠盲肠结扎穿孔模型,设立假手术组(A)和脂肪乳组(B)、单纯损伤组(C)、雌二醇组(D)、胰岛素组(E)等实验组,采用RT-PCR检测脂肪、肝及肺内leptinmRNA表达的变化.结果:正常大鼠的上述9种重要脏器内均有leptinmRNA表达,肾脏内含量最高而睾丸内含量最低.大鼠盲肠结扎穿孔12h后,与A组leptinmRNA表达水平相比,其在B组脂肪内表达显著增高而在肝、肺内表达显著降低,在C组肝内表达无显著差异而在脂肪、肺内表达显著降低,在D组肺内表达显著增高而在脂肪、肝内表达显著降低,在E组肺内表达无显著差异而在脂肪、肝内表达显著降低.脂肪乳对leptinmRNA表达的影响具有中枢分泌组织(脂肪)内诱导而外周脏器内抑制的双向模式.结论:LeptinmRNA表达水平在干预急性肠道损伤后能量代谢和神经-内分泌功能时发生敏感变化,提示leptin可能作为一种核心保护因子促进内环境的稳定.     

Effect of fasting on PPARgamma and AMPK activity in adipocytes

We investigated the effects of fasting on gene expression and intracellular signals regulating energy metabolism in adipose tissue. Following fasting for 15 h or 39 h, epididymal fat pads were isolated from Wistar rats. PPARγ mRNA levels decreased in the adipose tissues isolated from rats fasted for 39 h, whereas adipocyte lipid-binding protein (aP2) and lipoprotein lipase (LPL) mRNA levels increased. Overnight fasting increased the AMP/ATP ratio and AMP-activated protein kinase (AMPK) in adipose tissue, but not in muscle or liver tissue. In addition, the effect of 5-aminoimidazole-4-carboxyamide-ribonucleoside (AICAR) on PPARγ expression in primary cultured adipocytes was investigated. AICAR reduced PPARγ mRNA levels but increased aP2 and LPL mRNA levels. Thus, fasting-induced AMPK activation may affect on the regulation of gene expression in adipocytes.     

Expression of neuromedin B in adipose tissue and its regulation by changes in energy balance

Bombesin is one of the most powerful substances showing anorexic effects in the hypothalamus (Moody TW & Merali Z 2004 Bombesin-like peptides and associated receptors within the brain: distribution and behavioral implications. Peptides 25 511-520). In mammals, neuromedin B (NMB) is one member of a family of bombesin-like peptides, which have been shown to reduce food intake when administered systemically. Using Taqman real-time PCR with specific primers, we report the expression of NMB mRNA in both human and rodent adipose tissue. Expression of NMB in rodent epididymal adipose tissue was higher than in other tissues studied. Expression of NMB in adipose tissue appears to be regulated by changes in energy balance and leptin. It is decreased fourfold in the epididymal fat depot of ob/ob mice when compared with the same fat depot in lean mice. It is further decreased with the intra-peritoneally (i.p.) administration of leptin in both lean and obese ob/ob mice. This may relate to its function in food intake regulation or to changes in energy expenditure. We demonstrate that NMB expression in rodent adipose tissue is decreased in cold exposed animals. However, when we investigated the effects of NMB on resting metabolic rate by i.p. injection, there was no effect on oxygen consumption, RQ or physical activity when compared with saline-treated controls. In conclusion, NMB is expressed in both human and rodent adipose tissue and appears to be regulated by changes in energy balance. Given its anorexic effects centrally, it may form part of a new adipose tissue--hypothalamic axis regulating food intake.     

Dehydroepiandrosterone down-regulates the expression of peroxisome proliferator-activated receptor gamma in adipocytes

Dehydroepiandrosterone (DHEA) is expected to have a weight-reducing effect. In this study, we evaluated the effect of DHEA on genetically obese Otsuka Long Evans Fatty rats (OLETF) compared with Long-Evans Tokushima rats (LETO) as control. Feeding with 0.4% DHEA-containing food for 2 wk reduced the weight of sc, epididymal, and perirenal adipose tissue in association with decreased plasma leptin levels in OLETF. Adipose tissue from OLETF showed increased expression of peroxisome proliferator-activated receptor gamma (PPARgamma) protein, which was prevented by DHEA treatment. Further, we examined the effect of DHEA on PPARgamma in primary cultured adipocytes and monolayer adipocytes differentiated from rat preadipocytes. PPARgamma protein level was decreased in a time- and concentration-dependent manner, and DHEA significantly reduced mRNA levels of PPARgamma, adipocyte lipid-binding protein, and sterol regulatory element-binding protein, but not CCAAT/enhancer binding protein alpha. DHEA-sulfate also reduced the PPARgamma protein, but dexamethasone, testosterone, or androstenedione did not alter its expression. In addition, treatment with DHEA for 5 d reduced the triglyceride content in monolayer adipocytes. These results suggest that DHEA down-regulates adiposity through the reduction of PPARgamma in adipocytes.     

二甲双胍对人肝细胞Huh7脂质蓄积的影响及作用机制

目的研究二甲双胍对肝细胞脂质蓄积的影响及作用机制。方法体外培养人源Huh7肝细胞,细胞随机分为空白对照组及5 mmol/L二甲双胍、10 mmol/L二甲双胍、15 mmol/L二甲双胍处理组,确定最佳浓度后,使用LDL荷脂,然后分为空白对照组、LDL处理组和LDL+15 mmol/L二甲双胍处理组。实验终点,采用细胞增殖及毒性检测试剂盒检测细胞存活率,油红O染色和BODIPY脂质探针检测细胞内脂质含量,Dil-LDL检测细胞脂质摄取能力,Western blot分析固醇调节元件结合蛋白2(SREBP-2)、SREBP-1c、低密度脂蛋白受体(LDLR)和前蛋白转化酶枯草溶菌素9(PCSK9)蛋白的表达情况。结果二甲双胍可抑制LDL诱导的肝细胞脂质蓄积,降低SREBP-2、SREBP-1c、LDLR和PCSK9的蛋白表达。结论二甲双胍可能通过减少转录因子SREBP-2和SREBP-1c,下调LDLR表达,从而抑制细胞脂质蓄积。     

Age-specific effects of short- and long-term caloric restriction on the expression of adiponectin and adiponectin receptors: influence of intensity of food restriction

Hormonal signals from adipose tissue regulate energy homeostasis and may be involved in anti-aging effects of caloric restriction (CR). The adipokine adiponectin is abundantly expressed in adipose tissue and directly sensitizes the body to insulin. The purpose of the current study was to investigate age-dependent effects of different levels of CR (16%, 2 months or 40%, 6 months) on adiponectin and on its receptors AdipoR1 and AdipoR2 in the left ventricle (LV). In young and senescent rats, 2 months of moderate CR reduced serum leptin. The same diet was sufficient to enhance serum adiponectin, adiponectin expression (visceral fat) and left ventricular AdipoR1 expression in young but not in senescent rats. The higher degree of CR, however, resulted in a mild induction of adiponectin expression in adipose tissue and release into plasma together with increased LV AdipoR1 also in old rats, while these effects were more pronounced in young rats. These changes in adiponectin activation were associated with reduced LV triglyceride content, suggesting an adiponectin-mediated reduced ectopic lipid deposition in nonadipose tissues. Thus, aging is associated with a loss of adiponectin inducibility by moderate CR. This reduction can only partially be overcome by increasing the degree and duration of CR.     

Atrial natriuretic peptide inhibits the production of adipokines and cytokines linked to inflammation and insulin resistance in human subcutaneous adipose tissue

Aims/hypothesis Increased adipose tissue secretion of adipokines and cytokines has been implicated in the chronic low-grade inflammation state and insulin resistance associated with obesity. We tested here whether the cardiovascular and metabolic hormone atrial natriuretic peptide (ANP) was able to modulate adipose tissue secretion of several adipokines (derived from adipocytes) and cytokines (derived from adipose tissue macrophages). Subjects and methods We used protein array to measure the secretion of adipokines and cytokines after a 24-h culture of human subcutaneous adipose tissue pieces treated or not with a physiological concentration of ANP. The effect of ANP on protein secretion was also directly studied on isolated adipocytes and macrophages. Gene expression was measured by real-time RT-quantitative PCR. Results ANP decreased the secretion of the pro-inflammatory cytokines IL-6 and TNF-α, of several chemokines, and of the adipokines leptin and retinol-binding protein-4 (RBP-4). The secretion of the anti-inflammatory molecules IL-10 and adiponectin remained unaffected. The cytokines were mainly expressed in macrophages that expressed all components of the ANP-dependent signalling pathway. The adipokines, leptin, adiponectin and RBP-4 were specifically expressed in mature adipocytes. ANP directly inhibited the secretion of IL-6 and monocyte chemoattractant protein-1 by macrophages. The inhibitory effects of ANP on leptin and growth-related oncogene-α secretions were not seen under selective hormone-sensitive lipase inhibition. Conclusions/interpretation We suggest that ANP, either by direct action on adipocytes and macrophages or through activation of adipocyte hormone-sensitive lipase, inhibits the secretion of factors involved in inflammation and insulin resistance.