Search

Peroxisome proliferator-activated receptor-γ (PPAR-γ) plays a critical role in regulating insulin sensitivity and glucose homeostasis. In this study, we identified highly efficient small interfering RNA (siRNA) sequences and used lentiviral short hairpin RNA and electroporation of siRNAs to deplete PPAR-γ from 3T3-L1 adipocytes to elucidate its role in adipogenesis and insulin signaling. We show that PPAR-γ knockdown prevented adipocyte differentiation but was not required for maintenance of the adipocyte differentiation state after the cells had undergone adipogenesis. We further demonstrate that PPAR-γ suppression reduced insulin-stimulated glucose uptake without affecting the early insulin signaling steps in the adipocytes. Using dual siRNA strategies, we show that this effect of PPAR-γ deletion was mediated by both GLUT4 and GLUT1. Interestingly, PPAR-γ-depleted cells displayed enhanced inflammatory responses to TNF-α stimulation, consistent with a chronic anti-inflammatory effect of endogenous PPAR-γ. In summary, 1) PPAR-γ is essential for the process of adipocyte differentiation but is less necessary for maintenance of the differentiated state, 2) PPAR-γ supports normal insulin-stimulated glucose transport, and 3) endogenous PPAR-γ may play a role in suppression of the inflammatory pathway in 3T3-L1 cells. [ABSTRACT FROM AUTHOR]

The link between intra-abdominal adiposity and type II diabetes has been known for decades, and adipose tissue macrophage (ATM)-associated inflammation has recently been linked to insulin resistance. However, the mechanisms associated with ATM recruitment remain ill defined. Herein, we describe in vitro chemotaxis studies, in which adipocyte conditioned medium was used to stimulate macrophage migration. We demonstrate that tumor necrosis factor α and free fatty acids, key inflammatory stimuli involved in obesity-associated autocrine/paracrine inflammatory signaling, stimulate adipocyte expression and secretion of macrophage chemoattractants. Pharmacological studies showed that peroxisome proliferator-activated receptor γ agonists and glucocorticoids potently inhibit adipocyteinduced recruitment of macrophages. This latter effect was mediated by the glucocorticoid receptor, which led to decreased chemokine secretion and expression. In vivo results were quite comparable; treatment of high fat diet-fed mice with dexamethasone prevented ATM accumulation in epididymal fat. This decrease in ATM was most pronounced for the promflammatory F4/80+, CD11b+, CD11c+ M-1-like ATM subset. Overall, our results elucidate a beneficial function of peroxisome proliferator-activated receptor γ activation and glucocorticoid receptor/glucocorticoids in adipose tissue and indicate that pharmacologic prevention of ATM accumulation could be beneficial. [ABSTRACT FROM AUTHOR]

PPAR gamma is required for fat cell development and is the molecular target of antidiabetic thiazolidinediones (TZDs), which exert insulin-sensitizing effects in adipose tissue, skeletal muscle, and liver. Unexpectedly, we found that inactivation of PPAR gamma in macrophages results in the development of significant glucose intolerance plus skeletal muscle and hepatic insulin resistance in lean mice fed a normal diet. This phenotype was associated with increased expression of inflammatory markers and impaired insulin signaling in adipose tissue, muscle, and liver. PPAR gamma-deficient macrophages secreted elevated levels of factors that impair insulin responsiveness in muscle cells in a manner that was enhanced by exposure to FFAs. Consistent with this, the relative degree of insulin resistance became more severe in mice lacking macrophage PPAR gamma following high-fat feeding, and these mice were only partially responsive to TZD treatment. These findings reveal an essential role of PPAR gamma in macrophages for the maintenance of whole-body insulin action and in mediating the antidiabetic actions of TZDs. [ABSTRACT FROM AUTHOR]

Impaired estrogen action is associated with the metabolic syndrome in humans. We sought to determine whether impaired estrogen action in female C57Bl6 mice, produced by whole body Esr1 ablation, could recapitulate aspects of this syndrome, including inflammation, insulin resistance, and obesity. Indeed, we found that global knockout (KO) of the estrogen receptor (ER)alpha leads to reduced oxygen uptake and caloric expenditure compared with wild-type (WT) mice. In addition, fasting insulin, leptin, and PAI-1 levels were markedly elevated, whereas adiponectin levels were reduced in normal chow-fed KO. Furthermore, ERalpha-KO mice exhibited impaired glucose tolerance and marked skeletal muscle insulin resistance that was accompanied by the accumulation of bioactive lipid intermediates, inflammation, and diminished PPARalpha, PPARdelta, and UCP2 transcript levels. Although the relative glucose intolerance and insulin resistance phenotype in KO mice became more severe with high-fat feeding, WT mice were refractory to these dietary-induced effects, and this protection coincided with a marked increase in circulating adiponectin and heat shock protein 72 levels in muscle, liver, and fat. These data indicate that ERalpha is critical for the maintenance of whole body insulin action and protection against tissue inflammation during both normal chow and high-fat feeding.

Peroxisome proliferator-activated receptor-gamma (PPAR-gamma) plays a critical role in regulating insulin sensitivity and glucose homeostasis. In this study, we identified highly efficient small interfering RNA (siRNA) sequences and used lentiviral short hairpin RNA and electroporation of siRNAs to deplete PPAR-gamma from 3T3-L1 adipocytes to elucidate its role in adipogenesis and insulin signaling. We show that PPAR-gamma knockdown prevented adipocyte differentiation but was not required for maintenance of the adipocyte differentiation state after the cells had undergone adipogenesis. We further demonstrate that PPAR-gamma suppression reduced insulin-stimulated glucose uptake without affecting the early insulin signaling steps in the adipocytes. Using dual siRNA strategies, we show that this effect of PPAR-gamma deletion was mediated by both GLUT4 and GLUT1. Interestingly, PPAR-gamma-depleted cells displayed enhanced inflammatory responses to TNF-alpha stimulation, consistent with a chronic anti-inflammatory effect of endogenous PPAR-gamma. In summary, 1) PPAR-gamma is essential for the process of adipocyte differentiation but is less necessary for maintenance of the differentiated state, 2) PPAR-gamma supports normal insulin-stimulated glucose transport, and 3) endogenous PPAR-gamma may play a role in suppression of the inflammatory pathway in 3T3-L1 cells.

We show that Topiramate (TPM) treatment normalizes whole body insulin sensitivity in high-fat diet (HFD)-fed male Wistar rats. Thus drug treatment markedly lowered glucose and insulin levels during glucose tolerance tests and caused increased insulin sensitization in adipose and muscle tissues as assessed by euglycemic clamp studies. The insulin-stimulated glucose disposal rate increased twofold (indicating enhanced muscle insulin sensitivity), and suppression of circulating FFAs increased by 200 to 300%, consistent with increased adipose tissue insulin sensitivity. There were no effects of TPM on hepatic insulin sensitivity in these TPM-treated HFD-fed rats. In addition, TPM administration resulted in a three- to fourfold increase in circulating levels of total and high-molecular-weight (HMW) adiponectin (Acrp30). Western blot analysis revealed normal AMPK (Thr(172)) phosphorylation in liver with a twofold increased phospho-AMPK in skeletal muscle in TPM-treated rats. In conclusion, 1) TPM treatment prevents overall insulin resistance in HFD male Wistar rats; 2) drug treatment improved insulin sensitivity in skeletal muscle and adipose tissue associated with enhanced AMPK phosphorylation; and 3) the tissue "specific" effects are associated with increased serum levels of adiponectin, particularly the HMW component.

Lipid infusion and high fat feeding are established causes of systemic and adipose tissue insulin resistance. In this study, we treated 3T3-L1 adipocytes with a mixture of free fatty acids (FFAs) to investigate the molecular mechanisms underlying fat-induced insulin resistance. FFA treatment impaired insulin receptor-mediated signal transduction and decreased insulin-stimulated GLUT4 translocation and glucose transport. FFAs activated the stress/inflammatory kinases c-Jun N-terminal kinase (JNK) and IKKbeta, and the suppressor of cytokine signaling protein 3, increased secretion of the inflammatory cytokine tumor necrosis factor (TNF)-alpha, and decreased secretion of adiponectin into the medium. RNA interference-mediated down-regulation of JNK blocked JNK activation and prevented most of the FFA-induced defects in insulin action. Blockade of TNF-alpha signaling with neutralizing antibodies to TNF-alpha or its receptors or with a dominant negative TNF-alpha peptide had a partial effect to inhibit FFA-induced cellular insulin resistance. We found that JNK activation by FFAs was not inhibited by blocking TNF-alpha signaling, whereas the FFA-induced increase in TNF-alpha secretion was inhibited by RNA interference-mediated JNK knockdown. Together, these results indicate that 1) JNK can be activated by FFAs through TNF-alpha-independent mechanisms, 2) activated JNK is a major contributor to FFA-induced cellular insulin resistance, and 3) TNF-alpha is an autocrine/paracrine downstream effector of activated JNK that can also mediate insulin resistance.