Model Of Diet-induced Insulin Resistance Research Articles

A high-sugar diet produces obesity and insulin resistance in wild-type Drosophila

SUMMARY Insulin-resistant, ‘type 2’ diabetes (T2D) results from a complex interplay between genes and environment. In particular, both caloric excess and obesity are strongly associated with T2D across many genetic backgrounds. To gain insights into how dietary excess affects insulin resistance, we studied the simple model organism Drosophila melanogaster. Larvae reared on a high-sugar diet were hyperglycemic, insulin resistant and accumulated fat – hallmarks of T2D – compared with those reared on control diets. Excess dietary sugars, but not fats or proteins, elicited insulin-resistant phenotypes. Expression of genes involved in lipogenesis, gluconeogenesis and -oxidation was upregulated in high-sugar-fed larvae, as were FOXO targets, consistent with known mechanisms of insulin resistance in humans. These data establish a novel Drosophila model of diet-induced insulin resistance that bears strong similarity to the pathophysiology of T2D in humans.

A high-sugar diet produces obesity and insulin resistance in wild-typeDrosophila

SUMMARYInsulin-resistant, ‘type 2’ diabetes (T2D) results from a complex interplay between genes and environment. In particular, both caloric excess and obesity are strongly associated with T2D across many genetic backgrounds. To gain insights into how dietary excess affects insulin resistance, we studied the simple model organism Drosophila melanogaster. Larvae reared on a high-sugar diet were hyperglycemic, insulin resistant and accumulated fat – hallmarks of T2D – compared with those reared on control diets. Excess dietary sugars, but not fats or proteins, elicited insulin-resistant phenotypes. Expression of genes involved in lipogenesis, gluconeogenesis and β-oxidation was upregulated in high-sugar-fed larvae, as were FOXO targets, consistent with known mechanisms of insulin resistance in humans. These data establish a novel Drosophila model of diet-induced insulin resistance that bears strong similarity to the pathophysiology of T2D in humans.

Intestinal SR-BI is upregulated in insulin-resistant states and is associated with overproduction of intestinal apoB48-containing lipoproteins

Intestinal lipid dysregulation is a common feature of insulin-resistant states. The present study investigated alterations in gene expression of key proteins involved in the active absorption of dietary fat and cholesterol in response to development of insulin resistance. Studies were conducted in two diet-induced animal models of insulin resistance: fructose-fed hamster and high-fat-fed mouse. Changes in the mRNA abundance of lipid transporters, adenosine triphosphate cassette (ABC) G5, ABCG8, FA-CoA ligase fatty acid translocase P4, Niemann-Pick C1-Like1 (NPC1L1), fatty acid transport protein 4 (FATP4), and Scavenger Receptor Class B Type I (SR-BI), were assessed in intestinal fragments (duodenum, jejunum, and ileum) using quantitative real-time PCR. Of all the transporters evaluated, SR-B1 showed the most significant changes in both animal models examined. A marked stimulation of SR-B1 expression was observed in all intestinal segments examined in both insulin-resistant animal models. The link between SR-BI expression and intestinal lipoprotein production was then examined in the Caco-2 cell model. SR-B1 overexpression in Caco-2 cells increased apolipoprotein B (apoB) 100 and apoB48 secretion, whereas RNAi knock down of SR-B1 decreased secretion of both apoB100 and apoB48. We also observed changes in subcellular distribution of SR-B1 in response to exogenous lipid and insulin. Confocal microscopy revealed marked changes in SR-BI subcellular distribution in response to both exogenous lipids (oleate) and insulin. In summary, marked stimulation of intestinal SR-BI occurs in vivo in animal models of diet-induced insulin resistance, and modulation of SR-BI in vitro regulates production of apoB-containing lipoprotein particles. We postulate that apical and/or basolateral SR-BI may play an important role in intestinal chylomicron production and may contribute to chylomicron overproduction normally observed in insulin-resistant states.

The hexosamine biosynthetic pathway can mediate myocardial apoptosis in a rat model of diet-induced insulin resistance

Type 2 diabetes is characterized by deranged metabolic pathways that may result in cardiovascular complications. For example, hyperglycaemia promotes flux through the hexosamine biosynthetic pathway (HBP) leading to greater O-GlcNAcylation of target proteins, with pathophysiological outcomes. This study investigated mechanisms whereby increased HBP flux elicits myocardial apoptosis in a rat model of diet-induced hyperglycaemia/insulin resistance. Four-week-old male Wistar rats were fed a high-fat diet (86 days) after which insulin resistance was assessed vs. matched controls. Oxidative stress was evaluated, and apoptotic peptide levels, BAD phosphorylation and overall O-GlcNAcylation assessed by immunoblotting. Protein-specific O-GlcNAcylation and BAD-Bcl-2 dimerization were determined by immunoprecipitation and Western blotting. Rats consuming the high-fat diet exhibited a moderate elevation in body weight, higher fasting insulin and glucose levels, and insulin resistance vs. controls. Overall protein O-GlcNAcylation was increased in hyperglycaemic/insulin-resistant hearts. In parallel, myocardial peptide levels of apoptotic markers (caspase-3, cytochrome-c, BAD) were significantly higher with insulin resistance. To gain mechanistic insight into our findings, we evaluated O-GlcNAcylation of BAD, a pro-apoptotic Bcl-2 homolog. Here we found increased BAD O-GlcNAcylation and decreased BAD phosphorylation (Ser136) in hyperglycaemic/insulin-resistant rat hearts. These data are in agreement with competition by phosphorylation and O-GlcNAcylation for the same or neighbouring site(s) on target proteins. Moreover, we observed increased BAD-Bcl-2 dimerization in hyperglycaemic/insulin-resistant hearts. The main finding of this study is that increased apoptosis in hyperglycaemic/insulin-resistant hearts can also be mediated through HBP-induced BAD O-GlcNAcylation and greater formation of BAD-Bcl-2 dimers (pro-apoptotic).

Neprilysin, obesity and the metabolic syndrome

ObjectiveNeprilysin (NEP), a zinc metallo-endopeptidase, has a role in blood pressure control and lipid metabolism. The present study tested the hypothesis that NEP is associated with insulin resistance and features of the metabolic syndrome (MetS) in a study of 318 healthy human subjects and in murine obesity and investigated NEP production by adipocytes in-vitro.Methods and ResultsIn 318 white European males, plasma NEP was elevated in the MetS and increased progressively with increasing MetS components. Plasma NEP activity correlated with insulin, homeostasis model assessment and body mass index in all subjects (p<0.01). Quantitative RT-PCR and Western blotting showed that in human pre-adipocytes NEP expression is upregulated 25-30 fold during differentiation into adipocytes. Microarray analysis of mRNA from differentiated human adipocytes confirmed high NEP expression comparable to adiponectin and plasminogen activator inhibitor-1. In a murine model of diet-induced insulin resistance, plasma NEP levels were significantly higher in high fat diet (HFD)-fed compared with normal chow diet (NCD)-fed animals (1642±529 and 820±487 pg/μl, respectively; p<0.01). Tissue NEP was increased in mesenteric fat in HFD compared with NCD-fed mice (p<0.05). NEP knock out mice did not display any changes in insulin resistance, glucose tolerance or body and epididymal fat pad weight compared to wild type mice.ConclusionsIn humans, NEP activity correlated with body mass index and measures of insulin resistance with increasing levels in subjects with multiple cardiovascular risk factors. NEP protein production in human adipocytes increased during cell differentiation and plasma and adipose tissue levels of NEP were increased in obese insulin resistant mice. Our results indicate that NEP associates with cardio-metabolic risk in the presence of insulin resistance and increases in obesity.

The Effects of Acarbose Treatment on Intimal Hyperplasia in a Rat Carotid Endarterectomy Model of Diet-Induced Insulin Resistance

Increased carotid restenosis due to revascularization therapy is associated with insulin resistance. We hypothesize that glucose control using acarbose may attenuate intimal hyperplasia in rat carotid endarterectomy model of diet-induced insulin resistance. Rats were fed low-fat complex carbohydrate (control) or high-fat sucrose (insulin resistance) for 4 months. Three days preoperatively, some high-fat-sucrose rats were on acarbose, remainder of the rats received placebo. Rat carotids were assessed with duplex pre-and postoperatively. Acarbose and placebo continued for 2 weeks. Glucose, insulin, blood flow velocities and intimal hyperplasia were determined. High-fat sucrose plus acarbose attenuated intimal hyperplasia. Post-drug high-fat sucrose glucose decreased. Blood flow velocities postoperatively elevated above baseline. High-fat sucrose increased blood flow velocities postoperatively, which was attenuated with acarbose. Glucose control by acarbose in rat carotid endarterectomy model of diet-induced insulin resistance resulted in attenuation of intimal hyperplasia.

Proteomic profiling of intestinal prechylomicron transport vesicle (PCTV)-associated proteins in an animal model of insulin resistance (94 char)

Intestinal overproduction of apolipoprotein B (apoB)-48-containing chylomicrons is increasingly recognized as an underlying factor in metabolic dyslipidemia commonly observed in insulin-resistant states. Enhanced chylomicron assembly and secretion has been documented in animal models of insulin resistance, but the underlying mechanistic factors are unknown. Chylomicron assembly occurs through a series of complex vesicular interactions involving prechylomicron transport vesicles (PCTVs), which transport lipids from the endoplasmic reticulum (ER) to the Golgi. We report proteomic profiles of PCTVs isolated from the enteric ER in the small intestine of the fructose-fed hamster, an established model of diet-induced insulin resistance. Using 2D gel electrophoresis and tandem mass spectrometry, PCTVs were characterized and proteomic profiles of PCTV-associated proteins from insulin-resistant and control enterocytes were developed, with the intention of identifying proteins involved in insulin signaling attenuation and lipoprotein overproduction. A number of PCTV-associated proteins were found to be differentially expressed including microsomal triglyceride transfer protein (MTP), apoB-48, Sar1 and VAMP7. We postulate that altered expression of Sar1 and MTP may contribute to increased chylomicron assembly in the fructose-fed hamster. These findings have increased our understanding of the intracellular assembly and transport of nascent chylomicrons and potential cellular factors responsible for lipoprotein overproduction in insulin-resistant states.

Partial A1Adenosine Receptor Agonist Regulates Cardiac Substrate Utilization in Insulin-Resistant Rats in Vivo

Reducing the availability and uptake of fatty acids is a plausible pharmaceutical target to ameliorate glucose intolerance and insulin resistance. CVT-3619 [2-{6-[((1R,2R)-2-hydroxycyclopentyl) amino]purin-9-yl(4S,5S,2R,3R)-5-[(2-fluorophenylthio)methyl]oxolane-3,4-diol] is a partial A(1) adenosine receptor agonist with antilipolytic properties. Aims of the present study were to examine the acute effects of CVT-3619 on whole-body and cardiac glucose and fatty acid kinetics in vivo in normal and diet-induced insulin-resistant rats. Male Sprague-Dawley rats were fed either a chow (CH) or high-fat (HF) diet for 4 weeks. Catheters were then chronically implanted in the carotid artery and jugular vein for sampling and infusions, respectively. After 5 days of recovery, fasted animals (10 h) received either saline or CVT-3619 (0.4 mg/kg bolus + 1 mg/kg/h). Indices of glucose and fatty acid utilization were obtained by the administration of 2-deoxy[(14)C]glucose and [9,10-(3)H]-(R)-2-bromopalmitate. HF feeding resulted in elevated, fasting insulin and free fatty acid (FFA) levels compared with CH. CVT-3619 caused a 64 and 86% reduction of FFA and insulin in HF (p < 0.05) but less (N.S.) in CH diet-fed animals. In HF diet-fed rats, CVT-3619 increased whole-body glucose clearance with no change in fatty acid kinetics. Likewise, analysis of cardiac tissue metabolism showed that CVT-3619 caused an increased glucose but not fatty acid clearance in HF-fed animals. Results show that the acute administration of CVT-3619 lowers circulating fatty acid levels, leading to improved whole-body and cardiac glucose clearance in a model of diet-induced insulin resistance. As such, CVT-3619 may be a treatment option for the restoration of substrate balance in the insulin-resistant heart.

Green tea increases insulin sensitivity and decreases brain oxidative stress in fructose‐fed rats

Hyperglycemia and insulin resistance are leading causes of early brain alterations. Our objective was to investigate the in vivo effects of green tea extract on insulin sensitivity, insulin signaling, and brain oxidative stress using an experimental rodent model of diet‐induced insulin resistance, the fructose‐fed rat. Wistar rats, 10 per group, received a fructose rich diet (FD) or FD plus 1g green tea extract/kg diet. After consuming tea extracts, plasma insulin decreased from 412 ± 100 to 13 ± 28 pmol/L. There were also associated decreases in glucose and triglycerides. Real‐time PCR measurements showed that increases in insulin sensitivity were associated with an up‐regulation of GLUT4, insulin receptor, IRS‐2, glycogen synthetase kinase, p‐110 phosphoinositol kinase (PI3‐K), and protein kinase B in the liver. Addition of green tea extract to the diet resulted in significant decreases in lipid oxidation and DNA oxidative damage in brain tissue while sulfhydryl groups and glutathione remained unchanged. Ferric‐reducing/antioxidant power (FRAP) in brain was also enhanced. This study shows that green tea extract protected against brain oxidative damage and suggests possible neuroprotective effects against insulin resistance‐induced, early oxidative brain alterations. (Funded by USDA/ARS and Unilever, France).

Dissection of the Insulin-Sensitizing Effect of Liver X Receptor Ligands

The liver X receptors (LXRalpha and beta) are nuclear receptors that coordinate carbohydrate and lipid metabolism. Treatment of insulin-resistant mice with synthetic LXR ligands enhances glucose tolerance, inducing changes in gene expression expected to decrease hepatic gluconeogenesis (via indirect suppression of gluconeogenic enzymes) and increase peripheral glucose disposal (via direct up-regulation of glut4 in fat). To evaluate the relative contribution of each of these effects on whole-body insulin sensitivity, we performed hyperinsulinemic-euglycemic clamps in high-fat-fed insulin-resistant rats treated with an LXR agonist or a peroxisome proliferator-activated receptor gamma ligand. Both groups showed significant improvement in insulin action. Interestingly, rats treated with LXR ligand had lower body weight and smaller fat cells than controls. Insulin-stimulated suppression of the rate of glucose appearance (Ra) was pronounced in LXR-treated rats, but treatment failed to enhance peripheral glucose uptake (R'g), despite increased expression of glut4 in epididymal fat. To ascertain whether LXR ligands suppress hepatic gluconeogenesis directly, mice lacking LXRalpha (the primary isotype in liver) were treated with LXR ligand, and gluconeogenic gene expression was assessed. LXR activation decreased expression of gluconeogenic genes in wild-type and LXRbeta null mice, but failed to do so in animals lacking LXRalpha. Our observations indicate that despite inducing suggestive gene expression changes in adipose tissue in this model of diet-induced insulin resistance, the antidiabetic effect of LXR ligands is primarily due to effects in the liver that appear to require LXRalpha. These findings have important implications for clinical development of LXR agonists as insulin sensitizers.

Chronic Treatment With Sildenafil Improves Energy Balance and Insulin Action in High Fat–Fed Conscious Mice

Stimulation of nitric oxide-cGMP signaling results in vascular relaxation and increased muscle glucose uptake. We show that chronically inhibiting cGMP hydrolysis with the phosphodiesterase-5 inhibitor sildenafil improves energy balance and enhances in vivo insulin action in a mouse model of diet-induced insulin resistance. High-fat-fed mice treated with sildenafil plus L-arginine or sildenafil alone for 12 weeks had reduced weight and fat mass due to increased energy expenditure. However, uncoupling protein-1 levels were not increased in sildenafil-treated mice. Chronic treatment with sildenafil plus L-arginine or sildenafil alone increased arterial cGMP levels but did not adversely affect blood pressure or cardiac morphology. Sildenafil treatment, with or without l-arginine, resulted in lower fasting insulin and glucose levels and enhanced rates of glucose infusion, disappearance, and muscle glucose uptake during a hyperinsulinemic (4 mU x kg(-1) x min(-1))-euglycemic clamp in conscious mice. These effects occurred without an increase in activation of muscle insulin signaling. An acute treatment of high fat-fed mice with sildenafil plus l-arginine did not improve insulin action. These results show that phosphodiesterase-5 is a potential target for therapies aimed at preventing diet-induced energy imbalance and insulin resistance.

Nutrient Sensing, Leptin and Insulin Action

The glucose-fatty acid cycle as proposed four decades ago by Randle suggests that insulin resistance develops in consequence of alterations of the metabolic pressure of lipids. The more recently published ‘hexosamine pathway theory’ and the ‘malonyl-CoA hypothesis’ depict insulin resistance as a consequence of an imbalance between utilization of lipids and carbohydrates. The latter is finely tuned by entry of fatty acids into the mitochondria and/or by entry of glucose to the hexosamine pathway. A significant body of evidence has also been accumulated which points to the complex effects of leptin, an adipocyte-derived signal of lipid stores, on the storage and metabolism of fats and carbohydrates. These are mediated either directly, through actions on specific tissues, or indirectly, via CNS, endocrine and neural mechanisms. The available literature also provides good evidence that leptin orchestrates the metabolic changes in a number of organs and tissues, and alters nutrient fluxes to favor energy expenditure over energy storage. In this article, the proposed lipopenic effects of leptin as studied in various animal models of diet-induced insulin resistance, and possible regulations of leptin production and action by marine fish oil feeding are reviewed.