The effect of PPARgamma ligands on the adipose tissue in insulin resistance

Insulin resistance is frequently accompanied by obesity and both obesity and type 2 diabetes are associated with a mild chronic inflammation. Elevated levels of various cytokines, such as TNF-alpha and IL-6, are typically found in the adipose tissue in these conditions. It has been suggested that many cytokines produced in the adipose tissue are derived from infiltrated inflammatory cells. However, the adipose tissue itself has proven to be an important endocrine organ, secreting several hormones and cytokines, usually referred to as adipokines. Peroxisome proliferator-activated receptor (PPAR)gamma is essential for adipocyte proliferation and differentiation. In recent years, PPARgamma and its ligands, the thiazolidinediones (TZD), have achieved great attention due to their insulin sensitizing and anti-inflammatory properties. Treatment with TZDs result in improved insulin signaling and adipocyte differentiation, increased adipose tissue influx of free fatty acids and inhibition of cytokine expression and action. As a result, PPARgamma plays a central role in maintaining a functional and differentiated adipose tissue.     

Monomeric tartrate resistant acid phosphatase induces insulin sensitive obesity

Background

Obesity is associated with macrophage infiltration of adipose tissue, which may link adipose inflammation to insulin resistance. However, the impact of inflammatory cells in the pathophysiology of obesity remains unclear. Tartrate resistant acid phosphatase (TRAP) is an enzyme expressed by subsets of macrophages and osteoclasts that exists either as an enzymatically inactive monomer or as an active, proteolytically processed dimer.

Principal Findings

Using mice over expressing TRAP, we show that over-expression of monomeric, but not the dimeric form in adipose tissue leads to early onset spontaneous hyperplastic obesity i.e. many small fat cells. In vitro, recombinant monomeric, but not proteolytically processed TRAP induced proliferation and differentiation of mouse and human adipocyte precursor cells. In humans, monomeric TRAP was highly expressed in the adipose tissue of obese individuals. In both the mouse model and in the obese humans the source of TRAP in adipose tissue was macrophages. In addition, the obese TRAP over expressing mice exhibited signs of a low-grade inflammatory reaction in adipose tissue without evidence of abnormal adipocyte lipolysis, lipogenesis or insulin sensitivity.

Conclusion

Monomeric TRAP, most likely secreted from adipose tissue macrophages, induces hyperplastic obesity with normal adipocyte lipid metabolism and insulin sensitivity.     

Protease-activated receptor 2-dependent phosphorylation of the tissue factor cytoplasmic domain

Tissue factor (TF) is the physiological activator of the coagulation cascade that plays pathophysiological roles in metastasis, angiogenesis, and inflammation. Downstream in coagulation, thrombin is the central protease that signals through G protein-coupled, protease-activated receptors (PARs). However, the TF-VIIa-Xa complex upstream in coagulation also activates PAR1 and 2. Here, we address the question of whether signaling of the TF initiation complex is a relevant pathway that leads to TF cytoplasmic domain phosphorylation. In heterologous expression systems and primary endothelial cells, we demonstrate that the ternary TF-VIIa-Xa complex induces TF phosphorylation specifically by activating PAR2 but not through PAR1 signaling. In addition, TF cytoplasmic domain phosphorylation is induced only by TF-dependent signaling but not by other coagulation factors in endothelial cells. Phosphorylation of the Pro-directed kinase target site Ser258 is dependent on prior phosphorylation of Ser253 by protein kinase C (PKC) alpha. TF phosphorylation is somewhat delayed and coincides with sustained PKCalpha activation downstream of PAR2 but not PAR1 signaling. Phosphatidylcholine-dependent phospholipase C is the major pathway that leads to prolonged PKCalpha recruitment downstream of PAR2. Thus, PAR2 signaling specifically phosphorylates TF in a receptor cross-talk that distinguishes upstream from downstream coagulation protease signaling.     

Insulin resistance in adipose tissue: direct and indirect effects of tumor necrosis factor-alpha

Insulin resistance is a fundamental defect that precedes the development of the full insulin resistance syndrome as well as beta cell failure and type 2 diabetes. Tumor necrosis factor-alpha (TNF-alpha), a paracrine/autocrine factor highly expressed in adipose tissues of obese animals and human subjects, is implicated in the induction of insulin resistance seen in obesity and type 2 diabetes. Here, we review several molecular aspects of adipose tissue physiology, and highlight the direct effects of TNF-alpha on the functions of adipose tissue including induction of lipolysis, inhibition of insulin signaling, and alterations in expression of adipocyte important genes through activation of NF-kappaB, as well as their pertinence to insulin sensitivity of adipocytes. We also review the ability of TNF-alpha to inhibit synthesis of several adipocyte-specific proteins including Acrp30 (adiponectin) and enhance release of free fatty acids (FFAs) from adipose tissue, and discuss how these factors may act as systemic mediators of TNF-alpha and affect whole body energy homeostasis and overall insulin sensitivity. On the basis of these mechanisms, we examine the therapeutic potential of blocking specific autocrine/paracrine signaling pathways in adipocytes, particularly those involving NF-kappaB, in the treatment of type 2 diabetes.     

The novel endocrine disruptor tolylfluanid impairs insulin signaling in primary rodent and human adipocytes through a reduction in insulin receptor substrate-1 levels

Emerging data suggest that environmental endocrine disrupting chemicals may contribute to the pathophysiology of obesity and diabetes. In a prior work, the phenylsulfamide fungicide tolylfluanid (TF) was shown to augment adipocyte differentiation, yet its effects on mature adipocyte metabolism remain unknown. Because of the central role of adipose tissue in global energy regulation, the present study tested the hypothesis that TF modulates insulin action in primary rodent and human adipocytes. Alterations in insulin signaling in primary mammalian adipocytes were determined by the phosphorylation of Akt, a critical insulin signaling intermediate. Treatment of primary murine adipose tissue in vitro with 100nM TF for 48h markedly attenuated acute insulin-stimulated Akt phosphorylation in a strain- and species-independent fashion. Perigonadal, perirenal, and mesenteric fat were all sensitive to TF-induced insulin resistance. A similar TF-induced reduction in insulin-stimulated Akt phosphorylation was observed in primary human subcutaneous adipose tissue. TF treatment led to a potent and specific reduction in insulin receptor substrate-1 (IRS-1) mRNA and protein levels, a key upstream mediator of insulin's diverse metabolic effects. In contrast, insulin receptor-β, phosphatidylinositol 3-kinase, and Akt expression were unchanged, indicating a specific abrogation of insulin signaling. Additionally, TF-treated adipocytes exhibited altered endocrine function with a reduction in both basal and insulin-stimulated leptin secretion. These studies demonstrate that TF induces cellular insulin resistance in primary murine and human adipocytes through a reduction of IRS-1 expression and protein stability, raising concern about the potential for this fungicide to disrupt metabolism and thereby contribute to the pathogenesis of diabetes.     

Adipocyte/macrophage fatty acid binding proteins control integrated metabolic responses in obesity and diabetes

Fatty acid binding proteins (FABPs) are cytosolic fatty acid chaperones whose biological role and mechanisms of action are not well understood. Here, we developed mice with targeted mutations in two related adipocyte FABPs, aP2 and mal1, to resolve their role in systemic lipid, glucose, and energy metabolism. Mice lacking aP2 and mal1 exhibited a striking phenotype with strong protection from diet-induced obesity, insulin resistance, type 2 diabetes, and fatty liver disease. These mice have altered cellular and systemic lipid transport and composition, leading to enhanced insulin receptor signaling, enhanced muscle AMP-activated kinase (AMP-K) activity, and dramatically reduced liver stearoyl-CoA desaturase-1 (SCD-1) activity underlying their phenotype. Taken together with the previously reported strong protection against atherosclerosis, these results demonstrate that adipocyte/macrophage FABPs have a robust impact on multiple components of metabolic syndrome, integrating metabolic and inflammatory responses in mice and constituting a powerful target for the treatment of these diseases.     

Identification of Pigment Epithelium-Derived Factor as an Adipocyte-Derived Inflammatory Factor

Obesity is a major risk factor for insulin resistance, type 2 diabetes mellitus and cardiovascular disease. The pathophysiology of obesity is associated with chronic low-grade inflammation. Adipose tissue in obesity is significantly infiltrated by macrophages that secrete cytokines. The mechanisms of interaction between macrophages and adipocytes, leading to macrophage activation and increased cytokine release, remain to be elucidated. We reasoned that an adipocyte-derived factor might stimulate activation of macrophages. We have identified pigment epithelium-derived factor (PEDF) as a mediator of inflammation that is secreted by adipocytes and mediates macrophage activation. Recombinant PEDF activates macrophages to release tumor necrosis factor (TNF) and interleukin-1 (IL-1). The PEDF receptor adipose triglyceride lipase (ATGL) is required for PEDF-mediated macrophage activation. Selective inhibition of ATGL on macrophages attenuates PEDF-induced TNF production, and PEDF enhances the phosphorylation of p38 and extracellular signal-regulated kinase 1/2 mitogen-activated protein kinases. PEDF administration to rats results in increased serum TNF levels, and insulin resistance. Together, these findings suggest that PEDF secreted by adipocytes contributes to the onset and maintenance of chronic inflammation in obesity, and may be a therapeutic target in ameliorating insulin resistance.     

Macrophage α1 AMP-activated Protein Kinase (α1AMPK) Antagonizes Fatty Acid-induced Inflammation through SIRT1

In this study, we aim to determine cellular mechanisms linking nutrient metabolism to the regulation of inflammation and insulin resistance. The nutrient sensors AMP-activated protein kinase (AMPK) and SIRT1 show striking similarities in nutrient sensing and regulation of metabolic pathways. We find that the expression, activity, and signaling of the major isoform α1AMPK in adipose tissue and macrophages are substantially down-regulated by inflammatory stimuli and in nutrient-rich conditions, such as exposure to lipopolysaccharide (LPS), free fatty acids (FFAs), and diet-induced obesity. Activating AMPK signaling in macrophages by 5-aminoimidazole-4-carboxamide-1-β4-ribofuranoside or constitutively active α1AMPK (CA-α1) significantly inhibits; although inhibiting α1AMPK by short hairpin RNA knock-down or dominant-negative α1AMPK (DN-α1) increases LPS- and FFA-induced tumor necrosis factor α expression. Chromatin immunoprecipitation and luciferase reporter assays show that activation of AMPK by CA-α1 in macrophages significantly inhibits LPS- or FFA-induced NF-κB signaling. More importantly, in a macrophage-adipocyte co-culture system, we find that inactivation of macrophage AMPK signaling inhibits adipocyte insulin signaling and glucose uptake. Activation of AMPK by CA-α1 increases the SIRT1 activator NAD⁺ content and SIRT1 expression in macrophages. Furthermore, α1AMPK activation mimics the effect of SIRT1 on deacetylating NF-κB, and the full capacity of AMPK to deacetylate NF-κB and inhibit its signaling requires SIRT1. In conclusion, AMPK negatively regulates lipid-induced inflammation, which acts through SIRT1, thereby contributing to the protection against obesity, inflammation, and insulin resistance. Our study defines a novel role for AMPK in bridging the signaling between nutrient metabolism and inflammation.     

Novel adipocyte lines from brown fat: a model system for the study of differentiation, energy metabolism, and insulin action

Adipose tissue has emerged as an important endocrine regulator of glucose metabolism and energy homeostasis. By virtue of the mitochondrial protein uncoupling protein-1 (UCP-1), brown fat additionally plays a unique role in thermoregulation. Interest has focused on this tissue not only as a target for pharmacotherapy of obesity and insulin resistance but also as an endocrine tissue with leptin secretion and high insulin sensitivity. Most studies of adipocytes have been limited either to primary cell culture or to a small number of established cell lines. Recently, we have generated immortalized brown adipocyte cell lines from single newborn mice of different knockout mouse models. These cell lines retain the main characteristics of primary cells including UCP-1 expression. They display sensitive and diverse metabolic responses to insulin and adrenergic stimulation and have proven to be useful in the characterization of UCP regulation and the role of key insulin signaling elements for insulin action. Here, we outline common approaches to the generation of adipose tissue cell lines. Furthermore, we propose that the novel technique of generating brown adipocyte lines from a single newborn mouse will be instrumental in gaining further insight into the role of a broad range of signaling molecules in adipose tissue biology and in the pathogenesis of insulin resistance.     

Mechanism of insulin resistance in A-ZIP/F-1 fatless mice

Insulin resistance is a major factor in the pathogenesis of type 2 diabetes and may be related to alterations in fat metabolism. Fatless mice have been created using dominant-negative protein (A-ZIP/F-1) targeted gene expression in the adipocyte and shown to develop diabetes. To understand the mechanism responsible for the insulin resistance in these mice, we conducted hyperinsulinemic-euglycemic clamps in awake fatless and wild type littermates before the development of diabetes and examined insulin action and signaling in muscle and liver. We found the fatless mice to be severely insulin-resistant, which could be attributed to defects in insulin action in muscle and liver. Both of these abnormalities were associated with defects in insulin activation of insulin receptor substrate-1 and -2-associated phosphatidylinositol 3-kinase activity and a 2-fold increase in muscle and liver triglyceride content. We also show that upon transplantation of fat tissue into these mice, triglyceride content in muscle and liver returned to normal as does insulin signaling and action. In conclusion, these results suggest that the development of insulin resistance in type 2 diabetes may be due to alterations in the partitioning of fat between the adipocyte and muscle/liver leading to accumulation of triglyceride in the latter tissues with subsequent impairment of insulin signaling and action.     

Malfunctioning of adipocytes in obesity is linked to quantitative surfaceome changes

Increased triglyceride accumulation in adipocytes caused by a misbalance between energy intake and energy consumption, results in increased adipocyte size, excess adipose tissue, increased body weight and ultimately, obesity. It is well established that enlarged adipocytes exhibit malfunctions that contribute to whole body insulin resistance, a key factor for the development of type 2 diabetes. However, the underlying molecular cause for dysfunctional adipocyte behavior and signaling is poorly understood. Since the adipocyte cell surface proteome, or surfaceome, represents the cellular signaling gateway to the microenvironment, we studied the contribution of this subproteome to adipocyte malfunctions in obesity. By using the chemoproteomic Cell Surface Capture (CSC) technology, we established surfaceome maps of primary adipocytes derived from different mouse models for metabolic disorders. Relative quantitative comparison between these surfaceome maps revealed a set of cell surface glycoproteins with modulated location-specific abundance levels. RNAi mediated targeting of a subset of the detected obesity modulated cell surface glycoproteins in an in vitro model system provided functional evidence for their role in adiponectin secretion and the lipolytic activity of adipocytes. Thus, we conclude that the identified cell surface glycoproteins which exhibit obesity induced abundance changes and impact adipocyte function at the same time contribute to adipocyte malfunction in obesity. The regulation of their concerted activities could improve adipocyte function in obesity.     

GIP-overexpressing mice demonstrate reduced diet-induced obesity and steatosis, and improved glucose homeostasis

Glucose-dependent insulinotropic polypeptide (GIP) is a gastrointestinal hormone that potentiates glucose-stimulated insulin secretion during a meal. Since GIP has also been shown to exert β-cell prosurvival and adipocyte lipogenic effects in rodents, both GIP receptor agonists and antagonists have been considered as potential therapeutics in type 2 diabetes (T2DM). In the present study, we tested the hypothesis that chronically elevating GIP levels in a transgenic (Tg) mouse model would increase adipose tissue expansion and exert beneficial effects on glucose homeostasis. In contrast, although GIP Tg mice demonstrated enhanced β-cell function, resulting in improved glucose tolerance and insulin sensitivity, they exhibited reduced diet-induced obesity. Adipose tissue macrophage infiltration and hepatic steatosis were both greatly reduced, and a number of genes involved in lipid metabolism/inflammatory signaling pathways were found to be down-regulated. Reduced adiposity in GIP Tg mice was associated with decreased energy intake, involving overexpression of hypothalamic GIP. Together, these studies suggest that, in the context of over-nutrition, transgenic GIP overexpression has the potential to improve hepatic and adipocyte function as well as glucose homeostasis.     

Adipocyte NCoR knockout decreases PPARγ phosphorylation and enhances PPARγ activity and insulin sensitivity

Insulin resistance, tissue inflammation, and adipose tissue dysfunction are features of obesity and Type 2 diabetes. We generated adipocyte-specific Nuclear Receptor Corepressor (NCoR) knockout (AKO) mice to investigate the function of NCoR in adipocyte biology, glucose and insulin homeostasis. Despite increased obesity, glucose tolerance was improved in AKO mice, and clamp studies demonstrated enhanced insulin sensitivity in liver, muscle, and fat. Adipose tissue macrophage infiltration and inflammation were also decreased. PPARγ response genes were upregulated in adipose tissue from AKO mice and CDK5-mediated PPARγ ser-273 phosphorylation was reduced, creating a constitutively active PPARγ state. This identifies NCoR as an adaptor protein that enhances the ability of CDK5 to associate with and phosphorylate PPARγ. The dominant function of adipocyte NCoR is to transrepress PPARγ and promote PPARγ ser-273 phosphorylation, such that NCoR deletion leads to adipogenesis, reduced inflammation, and enhanced systemic insulin sensitivity, phenocopying the TZD-treated state.     

Lipid-Overloaded Enlarged Adipocytes Provoke Insulin Resistance Independent of Inflammation

In obesity, adipocyte hypertrophy and proinflammatory responses are closely associated with the development of insulin resistance in adipose tissue. However, it is largely unknown whether adipocyte hypertrophy per se might be sufficient to provoke insulin resistance in obese adipose tissue. Here, we demonstrate that lipid-overloaded hypertrophic adipocytes are insulin resistant independent of adipocyte inflammation. Treatment with saturated or monounsaturated fatty acids resulted in adipocyte hypertrophy, but proinflammatory responses were observed only in adipocytes treated with saturated fatty acids. Regardless of adipocyte inflammation, hypertrophic adipocytes with large and unilocular lipid droplets exhibited impaired insulin-dependent glucose uptake, associated with defects in GLUT4 trafficking to the plasma membrane. Moreover, Toll-like receptor 4 mutant mice (C3H/HeJ) with high-fat-diet-induced obesity were not protected against insulin resistance, although they were resistant to adipose tissue inflammation. Together, our in vitro and in vivo data suggest that adipocyte hypertrophy alone may be crucial in causing insulin resistance in obesity.     

The farnesoid X receptor modulates adiposity and peripheral insulin sensitivity in mice

The farnesoid X receptor (FXR) is a bile acid (BA)-activated nuclear receptor that plays a major role in the regulation of BA and lipid metabolism. Recently, several studies have suggested a potential role of FXR in the control of hepatic carbohydrate metabolism, but its contribution to the maintenance of peripheral glucose homeostasis remains to be established. FXR-deficient mice display decreased adipose tissue mass, lower serum leptin concentrations, and elevated plasma free fatty acid levels. Glucose and insulin tolerance tests revealed that FXR deficiency is associated with impaired glucose tolerance and insulin resistance. Moreover, whole-body glucose disposal during a hyperinsulinemic euglycemic clamp is decreased in FXR-deficient mice. In parallel, FXR deficiency alters distal insulin signaling, as reflected by decreased insulin-dependent Akt phosphorylation in both white adipose tissue and skeletal muscle. Whereas FXR is not expressed in skeletal muscle, it was detected at a low level in white adipose tissue in vivo and induced during adipocyte differentiation in vitro. Moreover, mouse embryonic fibroblasts derived from FXR-deficient mice displayed impaired adipocyte differentiation, identifying a direct role for FXR in adipocyte function. Treatment of differentiated 3T3-L1 adipocytes with the FXR-specific synthetic agonist GW4064 enhanced insulin signaling and insulin-stimulated glucose uptake. Finally, treatment with GW4064 improved insulin resistance in genetically obese ob/ob mice in vivo. Although the underlying molecular mechanisms remain to be unraveled, these results clearly identify a novel role of FXR in the regulation of peripheral insulin sensitivity and adipocyte function. This unexpected function of FXR opens new perspectives for the treatment of type 2 diabetes.     

内质网应激响应在脂肪组织中的代谢调节作用

在真核细胞中,内质网是蛋白合成、加工及质量监控的关键细胞器,也是Ca2+储存及脂质合成的重要场所.细胞通过未折叠蛋白响应(unfolded protein response,UPR)感应外界不同刺激引发的内质网应激,在维持细胞功能稳态中发挥至关重要的作用.在哺乳动物中,三个位于内质网的跨膜蛋白——肌醇依赖酶la(ino...     

Essential role of protein tyrosine phosphatase 1B in obesity-induced inflammation and peripheral insulin resistance during aging

Protein tyrosine phosphatase 1B (PTP1B) is a negative regulator of insulin signaling and a therapeutic target for type 2 diabetes (T2DM). In this study, we have evaluated the role of PTP1B in the development of aging-associated obesity, inflammation, and peripheral insulin resistance by assessing metabolic parameters at 3 and 16 months in PTP1B(-/-) mice maintained on mixed genetic background (C57Bl/6J × 129Sv/J). Whereas fat mass and adipocyte size were increased in wild-type control mice at 16 months, these parameters did not change with aging in PTP1B(-/-) mice. Increased levels of pro-inflammatory cytokines, crown-like structures, and hypoxia-inducible factor (HIF)-1α were observed only in adipose tissue from 16-month-old wild-type mice. Similarly, islet hyperplasia and hyperinsulinemia were observed in wild-type mice with aging-associated obesity, but not in PTP1B(-/-) animals. Leanness in 16-month-old PTP1B(-/-) mice was associated with increased energy expenditure. Whole-body insulin sensitivity decreased in 16-month-old control mice; however, studies with the hyperinsulinemic-euglycemic clamp revealed that PTP1B deficiency prevented this obesity-related decreased peripheral insulin sensitivity. At a molecular level, PTP1B expression and enzymatic activity were up-regulated in liver and muscle of 16-month-old wild-type mice as were the activation of stress kinases and the expression of p53. Conversely, insulin receptor-mediated Akt/Foxo1 signaling was attenuated in these aged control mice. Collectively, these data implicate PTP1B in the development of inflammation and insulin resistance associated with obesity during aging and suggest that inhibition of this phosphatase by therapeutic strategies might protect against age-dependent T2DM.