Acute peripheral but not central administration of olanzapine induces hyperglycemia associated with hepatic and extra-hepatic insulin resistance

Atypical antipsychotic drugs such as Olanzapine induce weight gain and metabolic changes associated with the development of type 2 diabetes. The mechanisms underlying the metabolic side-effects of these centrally acting drugs are still unknown to a large extent. We compared the effects of peripheral (intragastric; 3 mg/kg/h) versus central (intracerebroventricular; 30 μg/kg/h) administration of Olanzapine on glucose metabolism using the stable isotope dilution technique (Experiment 1) in combination with low and high hyperinsulinemic-euglycemic clamps (Experiments 2 and 3), in order to evaluate hepatic and extra-hepatic insulin sensitivity, in adult male Wistar rats. Blood glucose, plasma corticosterone and insulin levels were measured alongside endogenous glucose production and glucose disappearance. Livers were harvested to determine glycogen content. Under basal conditions peripheral administration of Olanzapine induced pronounced hyperglycemia without a significant increase in hepatic glucose production (Experiment 1). The clamp experiments revealed a clear insulin resistance both at hepatic (Experiment 2) and extra-hepatic levels (Experiment 3). The induction of insulin resistance in Experiments 2 and 3 was supported by decreased hepatic glycogen stores in Olanzapine-treated rats. Central administration of Olanzapine, however, did not result in any significant changes in blood glucose, plasma insulin or corticosterone concentrations nor in glucose production. In conclusion, acute intragastric administration of Olanzapine leads to hyperglycemia and insulin resistance in male rats. The metabolic side-effects of Olanzapine appear to be mediated primarily via a peripheral mechanism, and not to have a central origin.     

Role of insulin-like growth factor iin maintaining normal glucose homeostasis

Insulin-like growth factor I (IGF-I) has significant structural homology with insulin. IGF-I has been shown to bind to insulin receptors to stimulate glucose transport in fat and muscle, to inhibit hepatic glucose output and to lower blood glucose while simultaneously suppressing insulin secretion. However, the precise role of IGF-I in maintaining normal glucose homeostasis and insulin sensitivity is not well defined. Studies in patients with diabetes have shown that in insulin-deficient states, serum IGF-I concentrations are low and increase with insulin therapy. Similarly, administration of insulin via the portal vein results in optimization of plasma IGF-I concentrations. A patient with an IGF1 gene deletion was shown to have severe insulin resistance that improved with IGF-I therapy. Studies conducted in experimental animals have shown that if IGF-I synthesis by the liver is deleted, the animals become insulin-resistant, and this is improved when IGF-I is administered. Likewise, deletion of the IGF-I receptor in muscle in mice induces severe insulin resistance. Administration of IGF-I to patients with type 2 diabetes mellitus has been shown to result in an improvement in insulin sensitivity and a reduction in the requirement for exogenously administered insulin to maintain glucose homeostasis. A polymorphism in the IGF1 gene that has been shown to reduce serum IGF-I results in an increased prevalence of type 2 diabetes. Taken together, these findings support the conclusion that IGF-I is necessary for normal insulin sensitivity, and impairment of IGF-I synthesis results in a worsening state of insulin resistance.     

Effects of high-fat overfeeding on mitochondrial function, glucose and fat metabolism, and adipokine levels in low-birth-weight subjects

Low birth weight (LBW) is associated with an increased risk of insulin resistance and downregulation of oxidative phosphorylation (OXPHOS) genes when exposed to a metabolic challenge of high-fat overfeeding (HFO). To elaborate further on the differential effects of HFO in LBW subjects, we measured in vivo mitochondrial function, insulin secretion, hepatic glucose production, and plasma levels of key regulatory hormones before and after 5 days of HFO in 20 young LBW and 26 normal-birth-weight (NBW) men. The LBW subjects developed peripheral insulin resistance after HFO due to impaired endogenous glucose storage (9.42 ± 4.19 vs. 5.91 ± 4.42 mg·kg FFM(-1)·min(-1), P = 0.01). Resting muscle phosphorcreatine and total ATP in muscle increased significantly after HFO in LBW subjects only, whereas additional measurements of mitochondrial function remained unaffected. Despite similar plasma FFA levels, LBW subjects displayed increased fat oxidation during insulin infusion compared with normal-birth-weight (NBW) subjects after HFO (0.37 ± 0.35 vs. 0.17 ± 0.33 mg·kg FFM(-1)·min(-1), P = 0.02). In contrast to NBW subjects, the plasma leptin levels of LBW subjects did not increase, and the plasma gastric inhibitory polypeptide (GIP) as well as pancreatic polypeptide (PP) levels increased less in LBW compared with NBW subjects during HFO. In conclusion, HFO unmasks dissociation between insulin resistance and mitochondrial dysfunction in LBW subjects, suggesting that insulin resistance may be a cause, rather than an effect, of impaired muscle OXPHOS gene expression and mitochondrial dysfunction. Reduced increments in response to HFO of fasting plasma leptin, PP, and GIP levels may contribute to insulin resistance, lower satiety, and impaired insulin secretion in LBW subjects.     

Long-Term Corticosterone Exposure Decreases Insulin Sensitivity and Induces Depressive-Like Behaviour in the C57BL/6NCrl Mouse

Chronic stress or long-term administration of glucocorticoids disrupts the hypothalamus-pituitary-adrenal system leading to continuous high levels of glucocorticoids and insulin resistance (IR). This pre-diabetic state can eventually develop into type 2 diabetes mellitus and has been associated with a higher risk to develop depressive disorders. The mechanisms underlying the link between chronic stress, IR and depression remains unclear. The present study aimed to establish a stress-depression model in mice to further study the effects of stress-induced changes upon insulin sensitivity and behavioural consequences. A pilot study was conducted to establish the optimal administration route and a pragmatic measurement of IR. Subsequently, 6-month-old C57BL/6NCrl mice were exposed to long-term oral corticosterone treatment via the drinking water. To evaluate insulin sensitivity changes, blood glucose and plasma insulin levels were measured at different time-points throughout treatment and mice were behaviourally assessed in the elevated zero maze (EZM), forced swimming test (FST) and open field test to reveal behavioural changes. Long-term corticosterone treatment increased body weight and decreased insulin sensitivity. The latter was revealed by a higher IR index and increased insulin in the plasma, whereas blood glucose levels remained unchanged. Corticosterone treatment induced longer immobility times in the FST, reflecting depressive-like behaviour. No effects were observed upon anxiety as measured in the EZM. The effect of the higher body weight of the CORT treated animals at time of testing did not influence behaviour in the EZM or FST, as no differences were found in general locomotor activity. Long-term corticosterone treatment via the drinking water reduces insulin sensitivity and induces depressive-like behaviour in the C57BL/6 mouse. This mouse model could thus be used to further explore the underlying mechanisms of chronic stress-induced T2DM and its association with increased prevalence of major depressive disorder on the short-term and other behavioural adaptations on the longer term.     

Attenuation of inflammation and cellular stress‐related pathways maintains insulin sensitivity in obese type I interleukin‐1 receptor knockout mice on a high‐fat diet

The development of insulin resistance in the obese is associated with chronic, low‐grade inflammation. We aimed to identify novel links between obesity, insulin resistance and the inflammatory response by comparing C57BL/6 with type I interleukin‐1 receptor knockout (IL‐1RI^?/?) mice, which are protected against diet‐induced insulin resistance. Mice were fed a high‐fat diet for 16 wk. Insulin sensitivity was measured and proteomic analysis was performed on adipose, hepatic and skeletal muscle tissues. Despite an equal weight gain, IL‐1RI^?/? mice had lower plasma glucose, insulin and triacylglycerol concentrations, compared with controls, following dietary treatment. The higher insulin sensitivity in IL‐1RI^?/? mice was associated with down‐regulation of antioxidant proteins and proteasomes in adipose tissue and hepatic soluble epoxide hydrolase, consistent with a compromised inflammatory response as well as increased glycolysis and decreased fatty acid β‐oxidation in their muscle. Their lower hepatic triacylglycerol concentrations may reflect decreased flux of free fatty acids to the liver, decreased hepatic fatty acid‐binding protein expression and decreased lipogenesis. Correlation analysis revealed down‐regulation of classical biomarkers of ER stress in their adipose tissue, suggesting that disruption of the IL‐1RI‐mediated inflammatory response may attenuate cellular stress, which was associated with significant protection from diet‐induced insulin resistance, independent of obesity.     

Enhanced prereceptor glucocorticoid metabolism and lipogenesis impair insulin signaling in the liver of fructose-fed rats

Overconsumption of fructose, as a highly lipogenic sugar, may profoundly affect hepatic metabolism and has been associated with many components of the metabolic syndrome, particularly with insulin resistance and Type 2 diabetes. In this study, we proposed that high fructose diet may enhance lipogenesis and decrease insulin sensitivity in the liver through dysregulation of glucocorticoid signaling. Therefore, we examined the effects of long-term consumption of 10% fructose solution on triglyceridemia, liver histology and intracellular corticosterone level, as well as on 11β-hydroxysteroid dehydrogenase type 1 (11βHSD1) and hexose-6-phosphate dehydrogenase (H6PDH) mRNA and protein levels in the rat liver. Glucocorticoid action was assessed by glucocorticoid receptor (GR) expression and intracellular redistribution. We also analyzed the expression of enzymes involved in gluconeogenesis and lipogenesis, phosphoenolpyruvate carboxykinase (PEPCK) and lipin-1. The results have shown that fructose-rich diet led to increase in 11βHSD1 and H6PDH protein levels, while hepatic corticosterone concentration remained unchanged. Concomitantly, GR was increasingly accumulated in the cytoplasm, whereas its nuclear level was unchanged and accompanied by diminished PEPCK mRNA level. Elevation of lipin-1 in the liver microsomes suggested that fructose diet led to an increase in lipogenesis and consequently to hypertriglyceridemia. The observed increase of insulin receptor supstrate-1 phosphorylation on Ser³⁰⁷ represents a hallmark of impaired insulin signaling in the liver of fructose-fed rat and probably is a consequence of the alterations in 11βHSD1 and lipin-1 levels. Overall, our findings suggest that fructose-rich diet may perturb hepatic prereceptor glucocorticoid metabolism and lipogenesis, resulting in hypertriglyceridemia and attenuated hepatic insulin sensitivity.     

Dissociation of inositol-requiring enzyme (IRE1α)-mediated c-Jun N-terminal kinase activation from hepatic insulin resistance in conditional X-box-binding protein-1 (XBP1) knock-out mice

Hepatic insulin resistance has been attributed to both increased endoplasmic reticulum (ER) stress and accumulation of intracellular lipids, specifically diacylglycerol (DAG). The ER stress response protein, X-box-binding protein-1 (XBP1), was recently shown to regulate hepatic lipogenesis, suggesting that hepatic insulin resistance in models of ER stress may result from defective lipid storage, as opposed to ER-specific stress signals. Studies were designed to dissociate liver lipid accumulation and activation of ER stress signaling pathways, which would allow us to delineate the individual contributions of ER stress and hepatic lipid content to the pathogenesis of hepatic insulin resistance. Conditional XBP1 knock-out (XBP1Δ) and control mice were fed fructose chow for 1 week. Determinants of whole-body energy balance, weight, and composition were determined. Hepatic lipids including triglyceride, DAGs, and ceramide were measured, alongside markers of ER stress. Whole-body and tissue-specific insulin sensitivity were determined by hyperinsulinemic-euglycemic clamp studies. Hepatic ER stress signaling was increased in fructose chow-fed XBP1Δ mice as reflected by increased phosphorylated eIF2α, HSPA5 mRNA, and a 2-fold increase in hepatic JNK activity. Despite JNK activation, XBP1Δ displayed increased hepatic insulin sensitivity during hyperinsulinemic-euglycemic clamp studies, which was associated with increased insulin-stimulated IRS2 tyrosine phosphorylation, reduced hepatic DAG content, and reduced PKCε activity. These studies demonstrate that ER stress and IRE1α-mediated JNK activation can be disassociated from hepatic insulin resistance and support the hypothesis that hepatic insulin resistance in models of ER stress may be secondary to ER stress modulation of hepatic lipogenesis.     

MRNA and enzyme activity of hepatic 11beta-hydroxysteroid dehydrogenase type 1 are elevated in C57BL/KsJ-db/db mice.

To evaluate the importance of 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1) in insulin resistant diabetic C57BL/KsJ-db/db mice, we measured the activity and mRNA level of 11beta-HSD1 in the liver of db/db mice and their heterozygote litter mates, db/+m mice. The blood glucose, plasma insulin, and corticosterone levels of db/db mice were significantly higher than those of db/+m mice. Despite hyperinsulinemia, the activity level of this enzyme was significantly higher in db/db mice, and the mRNA level of hepatic 11beta-HSD1 was also significantly higher in db/db mice. Since hepatic 11beta-HSD1 in vivo mainly functions as 11-keto-reductase and does not work as 11beta-oxidase, these results suggest that the rate of hepatic conversion of 11-dehydrocorticosterone to corticosterone is increased in db/db mice, resulting in higher glucocorticoid activity in the liver. The increased hepatic corticosterone concentration due to the elevation of 11beta-HSD1 and high plasma corticosterone concentration may antagonize the action of insulin and cause insulin resistance. These findings have a potentially important implication for relationships between increased hepatic 11beta-HSD1 and insulin resistance in db/db mice. The present paper is the first to demonstrate the increased activities and mRNA level of hepatic 11beta-HSD1 in db/db mice.     

Prematurity and insulin sensitivity

Premature infants of low and extremely low birth weight represent a challenge for neonatal intensive care units and paediatricians. These neonates may be at increased risk of insulin resistance and diabetes perinatally and during childhood. During the first week of postnatal life, infants born prematurely are at risk of abnormalities in glucose homeostasis. Additionally, there are major differences in their glucose/insulin homeostasis compared with infants born at term. Preterm infants are at risk of hypoglycaemia, due to decreases in deposits of glycogen and fat that occur during the third trimester, and also to transient hyperinsulinaemia. Hyperglycaemia may also be observed in preterm infants during the perinatal period. These infants are unable to suppress glucose production within a large range of glucose and insulin concentrations, insulin secretory response is inappropriate, insulin processing is immature and there is an increased ratio of the glucose transporters Glut-1/Glut-2 in fetal tissues, which limits sensitivity and hepatocyte reaction to increments in glucose/insulin concentration during hyperglycaemia. In addition, increased concentrations of tumour necrosis factor alpha present in intrauterine growth retardation (IUGR) and induce insulin resistance. It has been proposed that the reduced insulin sensitivity may result from adaptation to an adverse in utero environment during a critical period of development. We have investigated postnatal insulin resistance in 60 children born with very low birth weight and either small for gestational age or at an appropriate size for gestational age. This study showed that IUGR, rather than low birth weight itself, was associated with increased fasting insulin levels. As poor fetal growth may be associated with the development of obesity, type 2 diabetes and the metabolic syndrome in later life, it is important that we continue to increase our understanding of the effects of IUGR on postnatal growth and metabolism.     

Retinol-binding protein 4 in young men with low versus normal birth weight: physiological response to short-term overfeeding

Retinol-binding protein 4 (RBP4) is a plasma protein which is elevated in obesity and type 2 diabetes. We aimed to investigate whether RBP4 represents a mechanism underlying the associations between low birth weight (LBW), high-fat diet, and insulin resistance. Forty-six young, lean men with low (n = 20) or normal (n = 26) birth weight underwent a 5-day high-fat high-calorie (HFHC) dietary intervention. In vivo glucose metabolism was assessed by euglycemic-hyperinsulinemic clamp, glucose tracer and intravenous glucose tolerance test techniques. Body composition was measured by a dual-energy x-ray absorptiometry scan, and plasma RBP4 by an enzyme-linked immunosorbent assay. RBP4 was not associated with birth weight, but with BMI (β = 0.9 μg/ml (0.08; 1.8) (95% confidence interval), P = 0.03) and plasma levels of low-density lipoprotein cholesterol (β = 5.3 μg/ml (1.9; 8.7), P = 0.03) and triglycerides (β = 15.4 μg/ml (9.5; 21.3), P < 0.001). Under baseline diet conditions, RBP4 was associated with decreased disposition index (D(i)) (β = -2.4% (-4.5%; -0.2%), P = 0.04) and increased basal hepatic glucose production rate (HGP) (β = 0.02 mg kg(-1) min(-1) (0.002; 0.04), P = 0.03), but not associated with peripheral glucose disposal rate or hepatic insulin resistance index. RBP4 levels were not influenced by overfeeding or related to peripheral and hepatic insulin resistance provoked by the dietary intervention. In conclusion, plasma RBP4 in young men associates with components of the metabolic syndrome, but is not determined by birth weight and seems not to be involved in short-term high-fat diet-induced insulin resistance.     

Acute Glucagon Induces Postprandial Peripheral Insulin Resistance

Glucagon levels are often moderately elevated in diabetes. It is known that glucagon leads to a decrease in hepatic glutathione (GSH) synthesis that in turn is associated with decreased postprandial insulin sensitivity. Given that cAMP pathway controls GSH levels we tested whether insulin sensitivity decreases after intraportal (ipv) administration of a cAMP analog (DBcAMP), and investigated whether glucagon promotes insulin resistance through decreasing hepatic GSH levels.Insulin sensitivity was determined in fed male Sprague-Dawley rats using a modified euglycemic hyperinsulinemic clamp in the postprandial state upon ipv administration of DBcAMP as well as glucagon infusion. Glucagon effects on insulin sensitivity was assessed in the presence or absence of postprandial insulin sensitivity inhibition by administration of L-NMMA. Hepatic GSH and NO content and plasma levels of NO were measured after acute ipv glucagon infusion. Insulin sensitivity was assessed in the fed state and after ipv glucagon infusion in the presence of GSH-E. We founf that DBcAMP and glucagon produce a decrease of insulin sensitivity, in a dose-dependent manner. Glucagon-induced decrease of postprandial insulin sensitivity correlated with decreased hepatic GSH content and was restored by administration of GSH-E. Furthermore, inhibition of postprandial decrease of insulin sensitivity L-NMMA was not overcome by glucagon, but glucagon did not affect hepatic and plasma levels of NO. These results show that glucagon decreases postprandial insulin sensitivity through reducing hepatic GSH levels, an effect that is mimicked by increasing cAMP hepatic levels and requires physiological NO levels. These observations support the hypothesis that glucagon acts via adenylate cyclase to decrease hepatic GSH levels and induce insulin resistance. We suggest that the glucagon-cAMP-GSH axis is a potential therapeutic target to address insulin resistance in pathological conditions.     

Improvement of insulin sensitivity by antagonism of the renin-angiotensin system

The reduced capacity of insulin to stimulate glucose transport into skeletal muscle, termed insulin resistance, is a primary defect leading to the development of prediabetes and overt type 2 diabetes. Although the etiology of this skeletal muscle insulin resistance is multifactorial, there is accumulating evidence that one contributor is overactivity of the renin-angiotensin system (RAS). Angiotensin II (ANG II) produced from this system can act on ANG II type 1 receptors both in the vascular endothelium and in myocytes, with an enhancement of the intracellular production of reactive oxygen species (ROS). Evidence from animal model and cultured skeletal muscle cell line studies indicates ANG II can induce insulin resistance. Chronic ANG II infusion into an insulin-sensitive rat produces a markedly insulin-resistant state that is associated with a negative impact of ROS on the skeletal muscle glucose transport system. ANG II treatment of L6 myocytes causes impaired insulin receptor substrate (IRS)-1-dependent insulin signaling that is accompanied by augmentation of NADPH oxidase-mediated ROS production. Further critical evidence has been obtained from the TG(mREN2)27 rat, a model of RAS overactivity and insulin resistance. The TG(mREN2)27 rat displays whole body and skeletal muscle insulin resistance that is associated with local oxidative stress and a significant reduction in the functionality of the insulin receptor (IR)/IRS-1-dependent insulin signaling. Treatment with a selective ANG II type 1 receptor antagonist leads to improvements in whole body insulin sensitivity, enhanced insulin-stimulated glucose transport in muscle, and reduced local oxidative stress. In addition, exercise training of TG(mREN2)27 rats enhances whole body and skeletal muscle insulin action. However, these metabolic improvements elicited by antagonism of ANG II action or exercise training are independent of upregulation of IR/IRS-1-dependent signaling. Collectively, these findings support targeting the RAS in the design of interventions to improve metabolic and cardiovascular function in conditions of insulin resistance associated with prediabetes and type 2 diabetes.     

Hepatic Insulin Resistance in Obese Non-Diabetic Subjects and in Type 2 Diabetic Patients

Objective: Obese non-diabetic patients are characterized by an extra-hepatic insulin resistance. Whether obese patients also have decreased hepatic insulin sensitivity remains controversial. Research Methods and Procedures: To estimate their hepatic insulin sensitivity, we measured the rate of exogenous insulin infusion required to maintain mildly elevated glycemia in obese patients with type 2 diabetes, obese non-diabetic patients, and lean control subjects during constant infusions of somatostatin and physiological low-glucagon replacement infusions. To account for differences in insulin concentrations among the three groups of subjects, an additional protocol was also performed in healthy lean subjects with higher insulin infusion rates and exogenous dextrose infusion. Results: The insulin infusion rate required to maintain glycemia at 8.5 mM was increased 4-fold in obese patients with type 2 diabetes and 1.5-fold in obese non-diabetic patients. The net endogenous glucose production (measured with 6,6-²H₂-glucose) and total glucose output (measured with 2-²H₁-glucose) were ∼30% lower in the patients than in the lean subjects. Net endogenous glucose production and total glucose output were both markedly increased in both groups of obese patients compared with lean control subjects during hyperinsulinemia. Discussion: Our data indicate that both obese non-diabetic and obese type 2 diabetic patients have a blunted suppressive action of insulin on glucose production, indicating hepatic and renal insulin resistance.     

Crucial role of a long-chain fatty acid elongase, Elovl6, in obesity-induced insulin resistance

Insulin resistance is often associated with obesity and can precipitate type 2 diabetes. To date, most known approaches that improve insulin resistance must be preceded by the amelioration of obesity and hepatosteatosis. Here, we show that this provision is not mandatory; insulin resistance and hyperglycemia are improved by the modification of hepatic fatty acid composition, even in the presence of persistent obesity and hepatosteatosis. Mice deficient for Elovl6, the gene encoding the elongase that catalyzes the conversion of palmitate to stearate, were generated and shown to become obese and develop hepatosteatosis when fed a high-fat diet or mated to leptin-deficient ob/ob mice. However, they showed marked protection from hyperinsulinemia, hyperglycemia and hyperleptinemia. Amelioration of insulin resistance was associated with restoration of hepatic insulin receptor substrate-2 and suppression of hepatic protein kinase C epsilon activity resulting in restoration of Akt phosphorylation. Collectively, these data show that hepatic fatty acid composition is a new determinant for insulin sensitivity that acts independently of cellular energy balance and stress. Inhibition of this elongase could be a new therapeutic approach for ameliorating insulin resistance, diabetes and cardiovascular risks, even in the presence of a continuing state of obesity.     

C-C Chemokine Receptor 2 Inhibitor Ameliorates Hepatic Steatosis by Improving ER Stress and Inflammation in a Type 2 Diabetic Mouse Model

Hepatic steatosis is the accumulation of excess fat in the liver. Recently, hepatic steatosis has become more important because it occurs in the patients with obesity, type 2 diabetes, and hyperlipidemia and is associated with endoplasmic reticulum (ER) stress and insulin resistance. C-C chemokine receptor 2 (CCR2) inhibitor has been reported to improve inflammation and glucose intolerance in diabetes, but its mechanisms remained unknown in hepatic steatosis. We examined whether CCR2 inhibitor improves ER stress-induced hepatic steatosis in type 2 diabetic mice. In this study, db/db and db/m (n = 9) mice were fed CCR2 inhibitor (2 mg/kg/day) for 9 weeks. In diabetic mice, CCR2 inhibitor decreased plasma and hepatic triglycerides levels and improved insulin sensitivity. Moreover, CCR2 inhibitor treatment decreased ER stress markers (e.g., BiP, ATF4, CHOP, and XBP-1) and inflammatory cytokines (e.g., TNFα, IL-6, and MCP-1) while increasing markers of mitochondrial biogenesis (e.g., PGC-1α, Tfam, and COX1) in the liver. We suggest that CCR2 inhibitor may ameliorate hepatic steatosis by reducing ER stress and inflammation in type 2 diabetes mellitus.     

Suppression of diacylglycerol acyltransferase-2 (DGAT2), but not DGAT1, with antisense oligonucleotides reverses diet-induced hepatic steatosis and insulin resistance

Nonalcoholic fatty liver disease (NAFLD) is a major contributing factor to hepatic insulin resistance in type 2 diabetes. Diacylglycerol acyltransferase (Dgat), of which there are two isoforms (Dgat1 and Dgat2), catalyzes the final step in triglyceride synthesis. We evaluated the metabolic impact of pharmacological reduction of DGAT1 and -2 expression in liver and fat using antisense oligonucleotides (ASOs) in rats with diet-induced NAFLD. Dgat1 and Dgat2 ASO treatment selectively reduced DGAT1 and DGAT2 mRNA levels in liver and fat, but only Dgat2 ASO treatment significantly reduced hepatic lipids (diacylglycerol and triglyceride but not long chain acyl CoAs) and improved hepatic insulin sensitivity. Because Dgat catalyzes triglyceride synthesis from diacylglycerol, and because we have hypothesized that diacylglycerol accumulation triggers fat-induced hepatic insulin resistance through protein kinase C epsilon activation, we next sought to understand the paradoxical reduction in diacylglycerol in Dgat2 ASO-treated rats. Within 3 days of starting Dgat2 ASO therapy in high fat-fed rats, plasma fatty acids increased, whereas hepatic lysophosphatidic acid and diacylglycerol levels were similar to those of control rats. These changes were associated with reduced expression of lipogenic genes (SREBP1c, ACC1, SCD1, and mtGPAT) and increased expression of oxidative/thermogenic genes (CPT1 and UCP2). Taken together, these data suggest that knocking down Dgat2 protects against fat-induced hepatic insulin resistance by paradoxically lowering hepatic diacylglycerol content and protein kinase C epsilon activation through decreased SREBP1c-mediated lipogenesis and increased hepatic fatty acid oxidation.     

Influence of the hepatic eukaryotic initiation factor 2alpha (eIF2alpha) endoplasmic reticulum (ER) stress response pathway on insulin-mediated ER stress and hepatic and peripheral glucose metabolism

Recent studies have implicated endoplasmic reticulum (ER) stress in insulin resistance associated with caloric excess. In mice placed on a 3-day high fat diet, we find augmented eIF2α signaling, together with hepatic lipid accumulation and insulin resistance. To clarify the role of the liver ER stress-dependent phospho-eIF2α (eIF2α-P) pathway in response to acute caloric excess on liver and muscle glucose and lipid metabolism, we studied transgenic mice in which the hepatic ER stress-dependent eIF2α-P pathway was inhibited by overexpressing a constitutively active C-terminal fragment of GADD34/PPP1R15a, a regulatory subunit of phosphatase that terminates ER stress signaling by phospho-eIF2α. Inhibition of the eIF2α-P signaling in liver led to a decrease in hepatic glucose production in the basal and clamped state, which could be attributed to reduced gluconeogenic gene expression, resulting in reduced basal plasma glucose concentrations. Surprisingly, hepatic eIF2α inhibition also impaired insulin-stimulated muscle and adipose tissue insulin sensitivity. This latter effect could be attributed at least in part by an increase in circulating IGFBP-3 levels in the transgenic animals. In addition, infusion of insulin during a hyperinsulinemic-euglycemic clamp induced conspicuous ER stress in the 3-day high fat diet-fed mice, which was aggravated through continuous dephosphorylation of eIF2α. Together, these data imply that the hepatic ER stress eIF2α signaling pathway affects hepatic glucose production without altering hepatic insulin sensitivity. Moreover, hepatic ER stress-dependent eIF2α-P signaling is implicated in an unanticipated cross-talk between the liver and peripheral organs to influence insulin sensitivity, probably via IGFBP-3. Finally, eIF2α is crucial for proper resolution of insulin-induced ER stress.     

Angiotensin II-induced skeletal muscle insulin resistance mediated by NF-kappaB activation via NADPH oxidase

Reduced insulin sensitivity is a key factor in the pathogenesis of type 2 diabetes and hypertension. Skeletal muscle insulin resistance is particularly important for its major role in insulin-mediated glucose disposal. Angiotensin II (ANG II) is integral in regulating blood pressure and plays a role in the pathogenesis of hypertension. In addition, we have documented that ANG II-induced skeletal muscle insulin resistance is associated with generation of reactive oxygen species (ROS). However, the linkage between ROS and insulin resistance in skeletal muscle remains unclear. To explore potential mechanisms, we employed the transgenic TG(mRen2)27 (Ren-2) hypertensive rat, which harbors the mouse renin transgene and exhibits elevated tissue ANG II levels, and skeletal muscle cell culture. Compared with Sprague-Dawley normotensive control rats, Ren-2 skeletal muscle exhibited significantly increased oxidative stress, NF-kappaB activation, and TNF-alpha expression, which were attenuated by in vivo treatment with an angiotensin type 1 receptor blocker (valsartan) or SOD/catalase mimetic (tempol). Moreover, ANG II treatment of L6 myotubes induced NF-kappaB activation and TNF-alpha production and decreased insulin-stimulated Akt activation and GLUT-4 glucose transporter translocation to plasma membranes. These effects were markedly diminished by treatment of myotubes with valsartan, the antioxidant N-acetylcysteine, NADPH oxidase-inhibiting peptide (gp91 ds-tat), or NF-kappaB inhibitor (MG-132). Similarly, NF-kappaB p65 small interfering RNA reduced NF-kappaB p65 subunit expression and nuclear translocation and TNF-alpha production but improved insulin-stimulated phosphorylation (Ser(473)) of Akt and translocation of GLUT-4. These findings suggest that NF-kappaB plays an important role in ANG II/ROS-induced skeletal muscle insulin resistance.     

Metabolic Adaptations to Dexamethasone‐Induced Insulin Resistance in Healthy Volunteers

Objective : Insulin resistance is observed in individuals with normal glucose tolerance. This indicates that increased insulin secretion can compensate for insulin resistance and that additional defects are involved in impaired glucose tolerance or type 2 diabetes. The objective of this study was to evaluate a procedure aimed at assessing the compensatory mechanisms to insulin resistance. Research Methods and Procedures : Eight healthy nonobese female patients were studied on two occasions, before and after administration of 2 mg/d dexamethasone for 2 days during a two‐step hyperglycemic clamp. Insulin secretion was assessed from plasma insulin concentrations. Insulin sensitivity was assessed from the ratio of whole‐body glucose use (6, 6 ²H₂ glucose) to plasma insulin concentrations. This procedure is known to induce a reversible impairment of glucose tolerance and insulin resistance. Results : In all subjects, dexamethasone induced a decrease in insulin sensitivity and a proportionate increase in first‐phase insulin secretion and in insulin concentrations at both steps of glycemia. The resulting hyperinsulinemia allowed the restoration of normal whole‐body glucose uptake and the suppression of plasma free fatty acids and triglycerides. In contrast, the suppression of endogenous glucose production was impaired after dexamethasone (p < 0.01). Discussion : Increased insulin secretion fully compensates dexamethasone‐induced insulin resistance in skeletal muscle and adipose tissue but not in the liver. This suggests that failure to overcome hepatic insulin resistance can impair glucose tolerance. The compensatory insulin secretion in response to insulin resistance can be assessed by means of a hyperglycemic clamp after a dexamethasone challenge.     

Diabetes in Old Male Offspring of Rat Dams Fed a Reduced Protein Diet

Restricted fetal growth is associated with increased risk for the future development of Type 2 diabetes in humans. The study aim was to assess the glucose tolerance of old (seventeen months) male rats, which were growth restricted in early life due to maternal protein restriction during gestation and lactation. Rat mothers were fed diets containing either 20% or 8% protein and all offspring weaned onto a standard rat diet. In old-age fasting plasma glucose concentrations were significantly higher in the low protein offspring: 8.4 (1.3)mmol/l v. 5.3 (1.3)mmol/l (p = 0.005), Areas under the curves were increased by 67% for glucose (p = 0.01) and 81% for insulin (p = 0.01) in these rats in intravenous glucose tolerance tests, suggesting (a degree of) insulin resistance. These results show that early growth retardation due to maternal protein restriction leads to the development of diabetes in old male rat offspring. The diabetes is predominantly associated with insulin resistance.