bluuebunny/crossref_metadata_2025 huggingface dataset:
Association of SH2-Containing Inositol Phosphatase 2 With the Insulin Resistance of Diabetic db/db Mice
Authors: Hori Hiroyuki , Sasaoka Toshiyasu , Ishihara Hajime , Wada Tsutomu , Murakami Shihou , Ishiki Manabu , Kobayashi Masashi
Published: March 2007

SH-2-containing inositol 5′-phosphatase 2 (SHIP-2) is a physiologically important lipid phosphatase that functions to hydrolyze phosphatidylinositol (PI) 3-kinase product PI(3,4,5)P3 to PI(3,4)P2 in the negative regulation of insulin signaling. We investigated whether SHIP-2 is associated with the insulin resistance of diabetic db/db mice. The amount of SHIP-2 protein was elevated in quadriceps muscle and epididymal fat tissue, but not in the liver, of db/db mice relative to that in control db/+m mice. In accordance with the enhanced expression of SHIP-2, its localization at the membrane preparation was increased in the skeletal muscle and fat tissue of db/db mice. Insulin stimulation of PI 3-kinase activity was modestly decreased in skeletal muscle, fat tissue, and liver of db/db mice compared with that of db/+m mice. In addition to the modest decrease at the level of PI 3-kinase, the activity of Akt and protein kinase C (PKC)-ζ/λ, which are downstream molecules of PI 3-kinase, was more severely reduced in the skeletal muscle and fat tissue, but not in liver of db/db mice. Treatment with the insulin-sensitizing agent rosiglitazone decreased the elevated expression of SHIP-2 in the skeletal muscle and fat tissue of db/db mice. Insulin-induced Akt activation and PKC-ζ/λ phosphorylation were restored to the control level, although insulin-stimulated PI 3-kinase activation was minimally affected in the skeletal muscle and fat tissue of db/db mice. These results indicate that SHIP-2 is a novel molecule associated with insulin resistance in the skeletal muscle and fat tissue, and that insulin-induced activity of the downstream molecules of PI 3-kinase is decreased, at least in part, by the elevated expression of SHIP-2 in diabetic db/db mice.
Synthesis Rate of Muscle Proteins, Muscle Functions, and Amino Acid Kinetics in Type 2 Diabetes
Authors: Halvatsiotis Panagiotis , Short Kevin R. , Bigelow Maureen , Nair K. Sreekumaran
Published: March 2007

Improvement of glycemic status by insulin is associated with profound changes in amino acid metabolism in type 1 diabetes. In contrast, a dissociation of insulin effect on glucose and amino acid metabolism has been reported in type 2 diabetes. Type 2 diabetic patients are reported to have reduced muscle oxidative enzymes and Vo2max. We investigated the effect of 11 days of intensive insulin treatment (T2D+) on whole-body amino acid kinetics, muscle protein synthesis rates, and muscle functions in eight type 2 diabetic subjects after withdrawing all treatments for 2 weeks (T2D−) and compared the results with those of weight-matched lean control subjects using stable isotopes of the amino acids. Whole-body leucine, phenylalanine and tyrosine fluxes, leucine oxidation, and plasma amino acid levels were similar in all groups, although plasma glucose levels were significantly higher in T2D−. Insulin treatment reduced leucine nitrogen flux and transamination rates in subjects with type 2 diabetes. Synthesis rates of muscle mitochondrial, sarcoplasmic, and mixed muscle proteins were not affected by glycemic status or insulin treatment in subjects with type 2 diabetes. Muscle strength was also unaffected by diabetes or glycemic status. In contrast, the diabetic patients showed increased tendency for muscle fatigability. Insulin treatment also failed to stimulate muscle cytochrome C oxidase activity in the diabetic patients, although it modestly elevated citrate synthase. In conclusion, improvement of glycemic status by insulin treatment did not alter whole-body amino acid turnover in type 2 diabetic subjects, but leucine nitrogen flux, transamination rates, and plasma ketoisocaproate level were decreased. Insulin treatments in subjects with type 2 diabetes had no effect on muscle mitochondrial protein synthesis and cytochrome C oxidase, a key enzyme for ATP production.
Selective Antagonism of the NPY Y5 Receptor Does Not Have a Major Effect on Feeding in Rats
Authors: Turnbull Andrew V. , Ellershaw Laraine , Masters Dave J. , Birtles Susan , Boyer Scott , Carroll Debbie , Clarkson Paul , Loxham Sue J.G. , McAulay Pat , Teague Joanne L. , Foote Kevin M. , Pease J. Elizabeth , Block Michael H.
Published: March 2007

Neuropeptide Y (NPY) is thought to play a key role in stimulating feeding, thus making NPY receptors attractive appetite suppressant drug targets for treating obesity. Because the orexigenic effects of NPY have been ascribed to actions at the NPY Y5 receptor, we have determined the role of this receptor in feeding in rats, using a small molecule antagonist of this receptor. NPY5RA-972 is a selective and potent (<10 nmol/l) NPY Y5 receptor antagonist. This compound is central nervous system (CNS) penetrant, and an oral dose of 10 mg/kg NPY5RA-972 to rats produced concentrations in cerebrospinal fluid that Xgreatly exceeded the in vitro IC50 (inhibitory concentration 50%). Indeed, at doses to rats as low as 1 mg/kg, NPY5RA-972 inhibited feeding induced by intracerebroventricular (ICV) administration of a selective NPY Y5 agonist ([cPP1–7,NPY19–23,Ala31,Aib32,Gln34]-hPP). However, in the dose range 1–10 mg/kg, NPY5RA-972 had no significant effect on food intake in Wistar rats induced to feed by either ICV NPY or 24 h fasting or in free-feeding Wistar or obese Zucker rats. Chronic administration of NPY5RA-972 (10 mg/kg twice daily) had no effect on food intake or body weight in either free-feeding Wistar rats or dietary obese rats. These data indicate that NPY5RA-972 is a potent, selective, orally active, and CNS-penetrant antagonist of the NPY Y5 receptor that prevents feeding driven by activation of this receptor. The data obtained with this antagonist indicate that the NPY Y5 receptor is not a major regulator of feeding in the rat.
Improved Insulin Sensitivity and Resistance to Weight Gain in Mice Null for the Ahsg Gene
Authors: Mathews Suresh T. , Singh Gurmant P. , Ranalletta Mollie , Cintron Vivian J. , Qiang Xiaoling , Goustin Anton Scott , Jen Kai-Lin Catherine , Charron Maureen J. , Jahnen-Dechent Willi , Grunberger George
Published: March 2007

Fetuin inhibits insulin-induced insulin receptor (IR) autophosphorylation and tyrosine kinase activity in vitro, in intact cells, and in vivo. The fetuin gene (AHSG) is located on human chromosome 3q27, recently identified as a susceptibility locus for type 2 diabetes and the metabolic syndrome. Here, we explore insulin signaling, glucose homeostasis, and the effect of a high-fat diet on weight gain, body fat composition, and glucose disposal in mice carrying two null alleles for the gene encoding fetuin, Ahsg (B6, 129-Ahsgtm1Mbl). Fetuin knockout (KO) mice demonstrate increased basal and insulin-stimulated phosphorylation of IR and the downstream signaling molecules mitogen-activated protein kinase (MAPK) and Akt in liver and skeletal muscle. Glucose and insulin tolerance tests in fetuin KO mice indicate significantly enhanced glucose clearance and insulin sensitivity. Fetuin KO mice subjected to euglycemic-hyperinsulinemic clamp show augmented sensitivity to insulin, evidenced by increased glucose infusion rate (P = 0.077) and significantly increased skeletal muscle glycogen content (P < 0.05). When fed a high-fat diet, fetuin KO mice are resistant to weight gain, demonstrate significantly decreased body fat, and remain insulin sensitive. These data suggest that fetuin may play a significant role in regulating postprandial glucose disposal, insulin sensitivity, weight gain, and fat accumulation and may be a novel therapeutic target in the treatment of type 2 diabetes, obesity, and other insulin-resistant conditions.
PDX-1 Induces Differentiation of Intestinal Epithelioid IEC-6 Into Insulin-Producing Cells
Authors: Yoshida Shigeru , Kajimoto Yoshitaka , Yasuda Tetsuyuki , Watada Hirotaka , Fujitani Yoshio , Kosaka Hiroshi , Gotow Takahiro , Miyatsuka Takeshi , Umayahara Yutaka , Yamasaki Yoshimitsu , Hori Masatsugu
Published: March 2007

A homeodomain containing transcription factor PDX-1 can induce β-cell–specific gene expressions in some non–β-cells and may therefore be useful for future diabetes gene/cell therapy. Among the potential target organs or tissues for transcription factor–mediated induction of β-cell–like differentiation are the intestinal epithelial cells. They have certain merits over other tissues and organs in terms of accessibility for gene delivery and of similarity in developmental background to the pancreatic primordium. In this study, we used an intestinal epithelium–derived cell line, IEC-6 cells, and investigated the possible effects of PDX-1 expression in those cells. By exogenous expression of the PDX-1 gene, IEC-6 cells started expressing multiple β-cell–specific genes such as amylin, glucokinase, and Nkx6.1, which were not found in the original IEC-6 cells. Insulin gene expression, which was missing initially even in the PDX-1–transfected IEC-6 cells, became detectable when the cells were transplanted under the renal capsule of a rat. When the PDX-1+ IEC-6 cells were kept in vitro, treatment with betacellulin could also confer insulin gene expression to them. Although insulin secretory granules became visible by electron microscopy, they were secreted regardless of glucose concentration. The in vivo or in vitro inductions of the insulin gene expression were not observed in the PDX-1– IEC-6 cells. Thus, our present observations demonstrate the potency of intestinal epithelial cells as a tool for diabetes gene/cell therapy and provide further support for the potency of PDX-1 in driving β-cell–like differentiation in non–β-cells.
Foxa2 Controls Pdx1 Gene Expression in Pancreatic β-Cells In Vivo
Authors: Lee Catherine S. , Sund Newman J. , Vatamaniuk Marko Z. , Matschinsky Franz M. , Stoffers Doris A. , Kaestner Klaus H.
Published: March 2007

Differentiation of early foregut endoderm into pancreatic endocrine and exocrine cells depends on a cascade of gene activation events controlled by various transcription factors. Prior in vitro analysis has suggested that the forkhead/winged helix transcription factor Foxa2 (formerly HNF-3β) is a major upstream regulator of Pdx1, a homeobox gene essential for pancreatic development. Pdx1 is also essential for the maintenance of glucose homeostasis, as its human orthologue, IPF-1, is mutated in a subset of patients with early-onset type 2 diabetes (MODY4). To analyze the Foxa2/Pdx1 regulatory cascade during pancreatic β-cell differentiation, we used conditional gene ablation of Foxa2 in mice. We demonstrated that the deletion of Foxa2 in β-cell−specific knockout mice results in downregulation of Pdx1 mRNA and subsequent reduction of PDX-1 protein levels in islets. These data represent the first in vivo demonstration that Foxa2 acts upstream of Pdx1 in the differentiated β-cell.
Increased Glucose Sensitivity of Stimulus-Secretion Coupling in Islets FromPsammomys obesusAfter Diet Induction of Diabetes
Authors: Pertusa Jose A.G. , Nesher Rafael , Kaiser Nurit , Cerasi Erol , Henquin Jean-Claude , Jonas Jean-Christophe
Published: March 2007

When fed a high-energy (HE) diet, diabetes-prone (DP) Psammomys obesus develop type 2 diabetes with altered glucose-stimulated insulin secretion (GSIS). β-Cell stimulus-secretion coupling was investigated in islets isolated from DP P. obesus fed a low-energy (LE) diet (DP-LE) and after 5 days on a HE diet (DP-HE). DP-LE islets cultured overnight in 5 mmol/l glucose displayed glucose dose-dependent increases in NAD(P)H, mitochondrial membrane potential, ATP/(ATP + ADP) ratio, cytosolic calcium concentration ([Ca2+]c), and insulin secretion. In comparison, DP-HE islets cultured overnight in 10 mmol/l glucose were 80% degranulated and displayed an increased sensitivity to glucose at the level of glucose metabolism, [Ca2+]c, and insulin secretion. These changes in DP-HE islets were only marginally reversed after culture in 5 mmol/l glucose and were not reproduced in DP-LE islets cultured overnight in 10 mmol/l glucose, except for the 75% degranulation. Diabetes-resistant P. obesus remain normoglycemic on HE diet. Their β-cell stimulus-secretion coupling was similar to that of DP-LE islets, irrespective of the type of diet. Thus, islets from diabetic P. obesus display an increased sensitivity to glucose at the level of glucose metabolism and a profound β-cell degranulation, both of which may affect their in vivo GSIS.
Cerivastatin Improves Insulin Sensitivity and Insulin Secretion in Early-State Obese Type 2 Diabetes
Authors: Paniagua J.A. , López-Miranda J. , Escribano A. , Berral F.J. , Marín C. , Bravo D. , Paz-Rojas E. , Gómez P. , Barcos M. , Moreno J.A. , Pérez-Jiménez F.
Published: March 2007

In a double-blind, placebo-controlled, randomized crossover study, 15 stable mild hyperglycemic patients without treatment and with features of metabolic syndrome were treated with cerivastatin (0.4 mg/day) or placebo for 3 months. The insulin sensitivity index during the euglycemic-hyperinsulinemic clamp (EHC; 5.4 mmol/l; 80 mU · m−2 · min−1) was increased by cerivastatin treatment (66.39 ± 3.9 nmol · lean body mass [LBM]−1 · min−1 · pmol−1 · l−1) as compared with placebo (58.37 ± 3.69 nmol · LBM−1 · min−1 · pmol−1 · l− 1; P < 0.01) by 13.7%. Glucose oxidation during EHC was significantly higher with statin treatment (16.1 ± 1.37 μmol · LBM−1 · min−1) as compared with placebo (14.58 ± 1.48 μmol · LBM−1 · min−1; P < 0.05). During hyperinsulinemia (∼800 pmol/l) in EHC steady-state, lipid oxidation was significantly decreased and respiratory quotient was significantly increased with statin treatment (0.33 ± 0.05 mg · LBM−1 · min− 1, 0.94 ± 0.01) as compared with placebo (0.48 ± 0.06 mg · LBM−1 · min−1, 0.91 ± 0.01; P < 0.01 and P < 0.05, respectively). During statin treatment, the first-phase insulin response increased from 2.07 ± 0.28 to 2.82 ± 0.38 pmol · l−1 · pmol−1 (P < 0.05). The second phase of insulin responses examined by C-peptide and insulin levels averaged during the hyperglycemic clamp (20 mmol/l) was unchanged. In conclusion, this study demonstrates that 0.4 mg cerivastatin therapy improves first-phase insulin secretion and increases insulin-mediated glucose uptake and respiratory quotient in the early state of obese type 2 diabetes.
Effects of Mesangium Glycation on Matrix Metalloproteinase Activities
Authors: McLennan S.V. , Martell S.K.Y. , Yue D.K.
Published: March 2007

High glucose concentrations can decrease degradation of mesangium by reducing the activities of matrix metalloproteinases (MMPs). The aim of this study was to investigate the effects of glycation of mesangium matrix on MMP-2, the principal MMP secreted by mesangial cells to degrade type IV collagen. Also examined were membrane type 1 MMP (MT1-MMP), tissue inhibitors of MMPs (TIMP)-1 and -2, and transforming growth factor-β (TGF-β), which together regulate MMP-2 activities in an interacting manner. Human fetal mesangial cells were grown on mesangium matrix glycated by incubation in 500 mmol/l ribose, with or without aminoguanidine. The activities and gene expression of the abovementioned enzymes/inhibitors were measured by degradation of radiolabeled mesangium matrix, RT-PCR, and zymography. Glycation of mesangium matrix resulted in a threefold increase in advance glycation end products and reduced by 45% the matrix-degrading activity of MMPs secreted by mesangial cells. Analogous to the direct effects of high glucose concentrations, glycation of matrix increased the gene expression of MMP-2 and TIMP-1 (control 100 ± 16.9 vs. glycated 197.3 ± 30.6% and control 100 ± 5.3 vs. glycated 152.1 ± 20.1%, respectively; P < 0.05) and decreased MT1-MMP (control 100 ± 1.17 vs. glycated 54.1 ± 15.2%; P < 0.05). However, unlike high glucose concentrations, glycation was not associated with decreased activation of MMP-2. Similarly, glycation but not high glucose increased expression of TIMP-2 (control 100 ± 5.9 vs. glycated 168.2 ± 31.4%; P < 0.05), and the effects of glycation on degradation can be abolished by anti-TIMP-2 antibody. Glycation of matrix decreased TGF-β mRNA by 38.2% and total and active TGF-β by 35.5 and 21.5%, respectively, opposite the effects of high glucose concentrations. Our results indicate that glycation of matrix affects the balance between MMP-2 and its activator and inhibitors, but this phenomenon is not due to TGF-β. The process of glycation may impart to the mesangium matrix a memory effect that contributes to the long-term toxicity of hyperglycemia.
Glucose Increases Endothelial-Dependent Superoxide Formation in Coronary Arteries by NAD(P)H Oxidase Activation
Authors: Christ Michael , Bauersachs Johann , Liebetrau Claudia , Heck Marina , Günther Andreas , Wehling Martin
Published: March 2007

Increased vascular superoxide anion (O2−) formation is essentially involved in the pathophysiology of atherosclerosis. Chronic hyperglycemia induces endothelial dysfunction, probably due to increased formation of reactive oxygen intermediates. However, little is known about the localization, modulators, and molecular mechanisms of vascular O2− formation during hyperglycemia. In porcine coronary segments, high glucose significantly increased O2− formation (1,703.5 ± 394.9 vs. 834.1 ± 91.7 units/mg for control, n = 64, P < 0.05; measured by lucigenin-enhanced chemiluminescence). This effect was completely blocked after removal of the endothelium. Coincubation with 10 μmol/l atorvastatin, a lipophilic inhibitor of 3-hydroxy-3-methylglutaryl-coenzyme A reductase, attenuated basal and glucose-induced O2− formation (328.1 ± 46.5 and 332.8 ± 50.3 units/mg, P < 0.05 vs. without atorvastatin). Incubation with mevalonic acid reversed this effect. High glucose increased mRNA expression of the oxidase subunit p22phox, which was blocked by 10 μmol/l atorvastatin, whereas expression of gp91phox was unchanged. In conclusion, glucose-induced increase of vascular O2− formation is endothelium dependent and is probably mediated by increased p22phox subunit expression. Beneficial effects of statins in diabetic patients may be explained in part by attenuation of vascular O2− formation independent of lipid lowering.
Regulation of Dietary Fatty Acid Entrapment in Subcutaneous Adipose Tissue and Skeletal Muscle
Authors: Evans Kevin , Burdge Graham C. , Wootton Stephen A. , Clark Mo L. , Frayn Keith N.
Published: March 2007

Using stable isotopic labeling of dietary fatty acids in conjunction with arteriovenous difference measurements, we have assessed the regulation of lipoprotein lipase-derived fatty acid entrapment in subcutaneous adipose tissue and forearm muscle in healthy subjects in the postprandial state. Eight volunteers fasted overnight and were then given a mixed meal containing [ 1-13C]palmitic acid and [1-13C]oleic acid. At baseline and for 6 h after the meal, blood samples were obtained from an arterialized hand vein and veins draining subcutaneous abdominal adipose tissue and forearm muscle, and arteriovenous differences were calculated. Entrapment of labeled fatty acids released by circulating triacylglycerol hydrolysis was close to 100% at 60 min, decreasing to 10–30% by 360 min. Entrapment of labeled fatty acids in forearm muscle was >100% and did not change with time. This study shows that entrapment of dietary fatty acids in adipose tissue in the postprandial period is a highly regulated process (varying with time) and that this can be studied in humans using stable isotope- labeled fatty acids in combination with measurement of appropriate arteriovenous differences. Also, fatty acid trapping in skeletal muscle is fundamentally different from that in adipose tissue, in that all the fatty acids released by lipoprotein lipase in skeletal muscle are taken up by the tissue.
Troglitazone Treatment Increases Protein Kinase B Phosphorylation in Skeletal Muscle of Normoglycemic Subjects at Risk for the Development of Type 2 Diabetes
Authors: Meyer Marco M. , Levin Klaus , Grimmsmann Thomas , Perwitz Nina , Eirich Alexandra , Beck-Nielsen Henning , Klein Harald H.
Published: March 2007

We investigated whether the effect of troglitazone on glucose disposal is associated with altered insulin signaling. Nondiabetic first-degree relatives of type 2 diabetic patients (age 30 ± 2 years, BMI 30 ± 1 kg/m2; n = 20) were randomized in a double-blind manner to 3 months of troglitazone (200 mg/day) or placebo treatment. Before and after treatment, 3-h euglycemic-hyperinsulinemic glucose clamps (40 mU · m−2 · min−1) were performed, and muscle biopsies were obtained immediately before and after the clamps. In the biopsies, insulin receptor kinase (IRK) activity, insulin receptor substrate (IRS)-1-associated phosphatidylinositol 3-kinase (PI3K) activity, Ser473 and Thr308 phosphorylation of protein kinase B (PKB), and protein expression of IRS-1, IRS-2, phosphoinositol-dependent kinase-1 (PDK-1), PKB, and GLUT-4 were determined. After troglitazone treatment, insulin-stimulated glucose disposal was increased compared with pretreatment and placebo (279 ± 37 vs. 211 ± 26 and 200 ± 25 mg · m−2 · min−1; both P < 0.05). IRK and PI3K activities were not altered by troglitazone, but PKB Ser473 phosphorylation was enhanced compared with pretreatment and placebo at the clamp insulin level (138 ± 36 vs. 77 ± 16 and 55 ± 13 internal standard units; both P < 0.05) and with pretreatment at the basal level (31 ± 9 vs. 14 ± 4 internal standard units; P < 0.05). PKB Thr308 phosphorylation also tended to be higher, but this was not statistically significant. Troglitazone did not alter insulin receptor number or IRS-1, IRS-2, PKB, PDK-1, or GLUT-4 protein expression. We conclude that increased PKB phosphorylation may contribute to the insulin-sensitizing effects of thiazolidinediones in human skeletal muscle.
Need for GLUT4 Activation to Reach Maximum Effect of Insulin-Mediated Glucose Uptake in Brown Adipocytes Isolated From GLUT4myc-Expressing Mice
Authors: Konrad Daniel , Bilan Philip J. , Nawaz Zafar , Sweeney Gary , Niu Wenyan , Liu Zhi , Antonescu Costin N. , Rudich Assaf , Klip Amira
Published: March 2007

There is a need to understand whether the amount of GLUT4 at the cell surface determines the extent of glucose uptake in response to insulin. Thus, we created a heterozygous mouse expressing modest levels of myc-tagged GLUT4 (GLUT4myc) in insulin-sensitive tissues under the control of the human GLUT4 promoter. Insulin stimulated 2-deoxyglucose uptake 6.5-fold in isolated brown adipocytes. GLUT1 did not contribute to the insulin response. The stimulation by insulin was completely blocked by wortmannin and partly (55 ± 2%) by the p38 mitogen-activated protein kinase (MAPK) inhibitor SB203580. Insulin increased surface exposure of GLUT4myc twofold (determined by fluorescent or enzyme-linked myc immunodetection in intact adipocytes). Such increase was completely blocked by wortmannin but insensitive to SB203580. Insulin increased the kinase activity of the p38 MAPK β-isoform 1.9-fold without affecting p38-α. In summary, the GLUT4myc mouse is a promising model for measuring GLUT4 translocation in intact primary cells. It affords direct comparison between GLUT4 translocation and glucose uptake in similar cell preparations, allowing one to study the regulation of GLUT4 activity. Using this animal model, we found that stimulation of glucose uptake into brown adipocytes involves both GLUT4 translocation and activation.
Androgens Decrease Plasma Adiponectin, an Insulin-Sensitizing Adipocyte-Derived Protein
Authors: Nishizawa Hitoshi , Shimomura Iichiro , Kishida Ken , Maeda Norikazu , Kuriyama Hiroshi , Nagaretani Hiroyuki , Matsuda Morihiro , Kondo Hidehiko , Furuyama Naoki , Kihara Shinji , Nakamura Tadashi , Tochino Yoshihiro , Funahashi Tohru , Matsuzawa Yuji
Published: March 2007

Adiponectin, an adipose-specific secretory protein, exhibits antidiabetic and antiatherogenic properties. In the present study, we examined the effects of sex hormones on the regulation of adiponectin production. Plasma adiponectin concentrations were significantly lower in 442 men (age, 52.6 ± 11.9 years [mean ± SD]) than in 137 women (53.2 ± 12.0 years) but not different between pre- and postmenopausal women. In mice, ovariectomy did not alter plasma adiponectin levels. In contrast, high levels of plasma adiponectin were found in castrated mice. Testosterone treatment reduced plasma adiponectin concentration in both sham-operated and castrated mice. In 3T3-L1 adipocytes, testosterone reduced adiponectin secretion into the culture media, using pulse-chase study. Castration-induced increase in plasma adiponectin was associated with a significant improvement of insulin sensitivity. Our results indicate that androgens decrease plasma adiponectin and that androgen-induced hypoadiponectinemia may be related to the high risks of insulin resistance and atherosclerosis in men.
Differential Interactions of Nateglinide and Repaglinide on the Human β-Cell Sulphonylurea Receptor 1
Authors: Hansen Ann Maria K. , Christensen Inge T. , Hansen John Bondo , Carr Richard D. , Ashcroft Frances M. , Wahl Philip
Published: March 2007

Repaglinide and nateglinide represent a new class of insulin secretagogues, structurally unrelated to sulphonylureas, that were developed for the treatment of type 2 diabetes. The inhibitory effect of these drugs was investigated on recombinant wild-type and mutant Kir6.2/SUR1 channels expressed in HEK293 cells. Nateglinide and repaglinide dose-dependently inhibited whole-cell Kir6.2/SUR1 currents with half-maximal inhibitory concentration (IC50) values of 800 and 21 nmol/l, respectively. Mutation of serine 1237 in SUR1 to tyrosine (S1237Y) abolished tolbutamide and nateglinide block, suggesting that these drugs share a common point of interaction on the SUR1 subunit of the ATP-sensitive K+ channel. In contrast, repaglinide inhibition was unaffected by the S1237Y mutation (IC50 = 23 nmol/l). Radioligand binding studies revealed a single high-affinity binding site for [3H]repaglinide on membranes prepared from HEK293 cells expressing wild-type (equilibrium dissociation constant [KD] = 0.40 nmol/l) or mutant (KD = 0.31 nmol/l) Kir6.2/SUR1 channels. Nateglinide and tolbutamide displaced [3H]repaglinide binding to wild-type channels with IC50 values of 0.7 and 26 μmol/l, respectively, but produced <10% displacement of [3H]repaglinide bound to mutant channels. This is consistent with the idea that binding of nateglinide and tolbutamide, but not repaglinide, is abolished by the SUR1[S1237Y] mutation and that the binding site for repaglinide is not identical to that of nateglinde/tolbutamide. These results are discussed in terms of a conformational analysis of the drug molecules.
Augmentation of the Neutrophil Respiratory Burst Through the Action of Advanced Glycation End Products
Authors: Wong Richard K.M. , Pettit Andrew I. , Davies Joan E. , Ng Leong L.
Published: March 2007

An accelerated accumulation of advanced glycation end products (AGEs) occurs in diabetes secondary to the increased glycemic burden. In this study, we investigated the contribution of AGEs to intravascular oxidant stress by examining their action on the neutrophil burst of reactive oxygen species (ROS); this may be a significant donor to the overall vascular redox status and to vasculopathy. AGEs exerted a dose-dependent enhancement on the neutrophil respiratory burst in response to a secondary mechanical stimulus (up to 265 ± 42%, P = 0.022) or chemical stimulation with formyl-methylleucylphenylalanine 100 nmol/l (up to 218 ± 19%, P < 0.001), although they possessed no ability to augment the neutrophil respiratory burst alone. This phenomenon was both immediate and reversible and depended on the simultaneous presence of AGEs with the additional stimulus. It appeared to work through an upregulation of the neutrophil NADPH oxidase, the enzyme responsible for ROS generation, as seen by a diphenyleneiodonium-dependent suppression of basal and augmented ROS output. Moreover, this action of AGEs was found to be complementary to that of neutrophil priming agents, also known to upregulate neutrophil ROS production, implying the presence of distinct intracellular transduction pathways mediating the effect of these two classes of agents.
Liver Pyruvate Kinase Polymorphisms Are Associated With Type 2 Diabetes in Northern European Caucasians
Authors: Wang Hua , Chu Winston , Das Swapan K. , Ren Qianfang , Hasstedt Sandra J. , Elbein Steven C.
Published: March 2007

Pyruvate kinase is a key glycolytic enzyme. Isoforms that are expressed in the red cell, liver, pancreatic β-cells, small intestine, and proximal renal tubule are encoded by the 12 exons of the PKLR gene, which maps to chromosome 1q23. We hypothesized that common variants of the PKLR gene could account for the linkage of diabetes to this region. We screened the promoter regions, exons and surrounding introns, and the 3′ untranslated region for mutations. We identified five single-nucleotide polymorphisms (SNPs), and only one (V506I, exon 11) altered the coding sequence. We tested the five SNPs, a poly-T insertion-deletion polymorphism, and an ATT triplet repeat in 131 unrelated diabetic patients and 118 nondiabetic control subjects. The V506I variant was rare and not associated with type 2 diabetes. The four SNPs and the insertion-deletion polymorphism were associated with diabetes, with a 10% difference between individuals with diabetes and nondiabetic individuals (P = 0.001–0.011, relative risk for minor allele 1.85). The same trend was found for the ATT repeat (P = 0.029). Common variants in the PKLR are associated with increased risk of type 2 diabetes, but because of strong linkage disequilibrium between variants, the actual susceptibility allele may be in a different gene.
A Comprehensive, Statistically Powered Analysis of GAD2 in Type 1 Diabetes
Authors: Johnson Gillian C.L. , Payne Felicity , Nutland Sarah , Stevens Helen , Tuomilehto-Wolf Eva , Tuomilehto Jaakko , Todd John A.
Published: March 2007

GAD2 maps to chromosome 10p11.23 and encodes the 65-kDa isoform of GAD65, a major autoantigen in type 1 diabetes. The genetic variation that influences expression of preproinsulin mRNA, encoding another major autoantigen in type 1 diabetes, has already been shown to be genetically associated with disease. Previous reports that have assessed the association of GAD2 with type 1 diabetes have not used a dense map of markers surrounding the gene and have relied on very small clinical sample sizes. Consequently, no definite conclusions can be drawn from their negative results. We have therefore systematically searched all exons, the 3′ untranslated region (UTR), the 5′ UTR, and the 5′ upstream region of GAD2, for polymorphisms in 32 white European individuals. We have genotyped these polymorphisms in a maximum of 472 U.K. type 1 diabetic affected sib pair families exhibiting linkage to type 1 diabetes on chromosome 10p and have tested both single variants and haplotypes in the GAD2 region for association with disease. We subsequently followed up our results by genotyping a subset of these single-nucleotide polymorphisms in a maximum of 873 Finnish families with at least one affected child. Our results suggest that GAD2 does not play a major role in type 1 diabetes in these two European populations.
Polymorphic Susceptibility to the Molecular Causes of Neural Tube Defects During Diabetic Embryopathy
Authors: Pani Lydie , Horal Melissa , Loeken Mary R.
Published: March 2007

Previously, we demonstrated that neural tube defects (NTDs) are significantly increased in a mouse model of diabetic pregnancy. In addition, expression of Pax-3, a gene encoding a transcription factor required for neural tube development, is significantly decreased. This suggests that diabetic embryopathy results from impaired expression of genes regulating essential morphogenetic processes. Here, we report that in one mouse strain, C57Bl/6J, embryos are resistant to the effects of maternal diabetes on NTDs and Pax-3 expression, in contrast to a susceptible strain, FVB, in which maternal diabetes significantly increases NTDs (P = 0.02) and inhibits Pax-3 expression (P = 0.01). Resistance to NTDs caused by diabetic pregnancy is a dominant trait, as demonstrated by heterozygous embryos of diabetic or nondiabetic mothers of either strain. There was no significant difference between strains in expression of genes that regulate free radical scavenging pathways, suggesting that susceptibility to oxidative stress does not account for the genetic differences. Understanding the genetic bases for differential susceptibility to altered gene expression and NTDs in diabetic mice may be important in delineating the mechanisms by which maternal hyperglycemia interferes with embryo gene expression. Moreover, if susceptibility to diabetic embryopathy is variable in humans as well as in mice, it may be possible to screen individuals at increased risk for this complication of diabetes.
Are Oxidative Stress−Activated Signaling Pathways Mediators of Insulin Resistance and β-Cell Dysfunction?
Authors: Evans Joseph L. , Goldfine Ira D. , Maddux Betty A. , Grodsky Gerold M.
Published: March 2007

In both type 1 and type 2 diabetes, diabetic complications in target organs arise from chronic elevations of glucose. The pathogenic effect of high glucose, possibly in concert with fatty acids, is mediated to a significant extent via increased production of reactive oxygen species (ROS) and reactive nitrogen species (RNS) and subsequent oxidative stress. ROS and RNS directly oxidize and damage DNA, proteins, and lipids. In addition to their ability to directly inflict damage on macromolecules, ROS and RNS indirectly induce damage to tissues by activating a number of cellular stress-sensitive pathways. These pathways include nuclear factor-κB, p38 mitogen-activated protein kinase, NH2-terminal Jun kinases/stress-activated protein kinases, hexosamines, and others. In addition, there is evidence that in type 2 diabetes, the activation of these same pathways by elevations in glucose and free fatty acid (FFA) levels leads to both insulin resistance and impaired insulin secretion. Therefore, we propose here that the hyperglycemia-induced, and possibly FFA-induced, activation of stress pathways plays a key role in the development of not only the late complications in type 1 and type 2 diabetes, but also the insulin resistance and impaired insulin secretion seen in type 2 diabetes.
Measurements of Insulin Secretory Capacity and Glucose Tolerance to Predict Pancreatic β-Cell Mass In Vivo in the Nicotinamide/Streptozotocin Göttingen Minipig, a Model of Moderate Insulin Deficiency and Diabetes
Authors: Larsen Marianne O. , Rolin Bidda , Wilken Michael , Carr Richard D. , Gotfredsen Carsten F.
Published: March 2007

Knowledge about β-cell mass and/or function could be of importance for the early diagnosis and treatment of diabetes. However, measurement of β-cell function as an estimate of β-cell mass is currently the only method possible in humans. The present study was performed to investigate different functional tests as predictors of β-cell mass in the Göttingen minipig. β-cell mass was reduced in the Göttingen minipig with a combination of nicotinamide (100 [n = 6], 67 [n = 25], 20 [n = 2], or 0 mg/kg [n = 4]) and streptozotocin (125 mg/kg). Six normal pigs were included. An oral glucose tolerance test (OGTT) (n = 43) and insulin secretion test (n = 30) were performed and pancreata obtained for stereological determination of β-cell mass. During OGTT, fasting glucose (r2 = 0.1744, P < 0.01), area under the curve for glucose (r2 = 0.2706, P < 0.001), maximum insulin secretion (r2 = 0.2160, P < 0.01), and maximum C-peptide secretion (r2 = 0.1992, P < 0.01) correlated with β-cell mass. During the insulin secretion test, acute insulin response to 0.3 g/kg (r2 = 0.6155, P < 0.0001) and 0.6 g/kg glucose (r2 = 0.7321, P < 0.0001) and arginine (67 mg/kg) (r2 = 0.7732, P < 0.0001) and maximum insulin secretion (r2 = 0.8192, P < 0.0001) correlated with β-cell mass. This study supports the use of functional tests to evaluate β-cell mass in vivo and has established a validated basis for developing a mathematical method for estimation of β-cell mass in vivo in the Göttingen minipig.
Intramyocellular Lipid and Insulin Resistance
Authors: Kuhlmann Johanna , Neumann-Haefelin Claudia , Belz Ulrich , Kalisch Jürgen , Juretschke Hans-Paul , Stein Marion , Kleinschmidt Elke , Kramer Werner , Herling Andreas W.
Published: March 2007

Insulin resistance plays an important role in the pathogenesis of human type 2 diabetes. In humans, a negative correlation between insulin sensitivity and intramyocellular lipid (IMCL) content has been shown; thus, IMCL becomes a marker for insulin resistance. Recently, magnetic resonance spectroscopy (MRS) has been established as a dependable method for selective detection and quantification of IMCL in humans. To validate the interrelation between insulin sensitivity and IMCL in an animal model of type 2 diabetes, we established volume selective 1H-MRS at 7 Tesla to noninvasively assess IMCL in the rat. In male obese Zucker Diabetic Fatty rats and their lean littermates, IMCL levels were determined repeatedly over 4 months, and insulin sensitivity was measured by the euglycemic-hyperinsulinemic clamp method at 6–7 and at 22–24 weeks of age. A distinct relation between IMCL and insulin sensitivity was demonstrated as well as age dependence for both parameters. Rosiglitazone treatment caused a clear reduction of IMCL and hepatic fat despite increased body weight, and a marked improvement of insulin sensitivity. Thus, the insulin sensitizing properties of rosiglitazone were consistent with a redistribution of lipids from nonadipocytic (skeletal muscle, liver) back into fat tissue.
The Atherogenic Lipoprotein Profile Associated With Obesity and Insulin Resistance Is Largely Attributable to Intra-Abdominal Fat
Authors: Nieves Delma J. , Cnop Miriam , Retzlaff Barbara , Walden Carolyn E. , Brunzell John D. , Knopp Robert H. , Kahn Steven E.
Published: March 2007

Obesity and insulin resistance are both associated with an atherogenic lipoprotein profile. We examined the effect of insulin sensitivity and central adiposity on lipoproteins in 196 individuals (75 men and 121 women) with an average age of 52.7 years. Subjects were subdivided into three groups based on BMI and their insulin sensitivity index (SI): lean insulin sensitive (n = 65), lean insulin resistant (n = 73), and obese insulin resistant (n = 58). This categorization revealed that both obesity and insulin resistance determined the lipoprotein profile. In addition, the insulin-resistant groups had increased central adiposity. Increasing intra-abdominal fat (IAF) area, quantified by computed tomography scan and decreasing SI, were important determinants of an atherogenic profile, marked by increased triglycerides, LDL cholesterol, and apolipoprotein B and decreased HDL cholesterol and LDL buoyancy (Rf). Density gradient ultracentrifugation (DGUC) revealed that in subjects who had more IAF and were more insulin resistant, the cholesterol content was increased in VLDL, intermediate-density lipoprotein (IDL), and dense LDL fractions whereas it was reduced in HDL fractions. Multiple linear regression analysis of the relation between the cholesterol content of each DGUC fraction as the dependent variable and IAF and SI as independent variables revealed that the cholesterol concentration in the fractions corresponding to VLDL, IDL, dense LDL, and HDL was associated with IAF, and that SI additionally contributed independently to VLDL, but not to IDL, LDL, or HDL. Thus an atherogenic lipoprotein profile appears to be the result primarily of an increase in IAF, perhaps via insulin resistance.
Combinations of Nondiabetic Parental Genomes Elicit Impaired Glucose Tolerance in Mouse SMXA Recombinant Inbred Strains
Authors: Kobayashi Misato , Ohno Tamio , Tsuji Atsushi , Nishimura Masahiko , Horio Fumihiko
Published: March 2007

Type 2 diabetes in humans is not a single gene disorder but a multifactorial disease caused by the interaction of multiple genes and environmental factors. Recombinant inbred (RI) strains are a powerful tool for analyzing not only single genetic traits but also multifactorial genetic traits. By using the SMXA RI mice, we genetically dissected diabetes-related traits (BMI, nonfasting blood glucose concentration, and blood glucose concentration during intraperitoneal glucose tolerance tests). For minimizing the variation of glucose tolerance in each strain, all mice were fed the high-carbohydrate diet and subjected to phenotypic and genetic analyses. The parental strains, SM/J and A/J, were nondiabetic, and the differences of the mean values of diabetes-related traits were small. In contrast, an impaired glucose tolerance was observed in (SM × A)F1 mice, and marked differences in diabetes-related traits were observed in 19 SMXA RI strains. In particular, several SMXA RI strains showed markedly impaired glucose tolerance and hyperglycemia. Quantitative trait locus (QTL) analysis revealed a locus on chromosome (Chr) 10 contributing significant effect on nonfasting blood glucose concentration, as well as six diabetes-related loci on four chromosomes with suggestive evidence of linkage with diabetes-related phenotypes. The A/J-derived QTLs on Chr 2 and 18 and an SM/J-derived QTL on Chr 10 contributed to the impairment of glucose tolerance and/or the increase of blood glucose concentration. The present study indicates that QTLs derived from parental SM/J and A/J genomes, both of which are nondiabetic, interact in the RI genomes, leading to the development of hyperglycemia and diabetic phenotypes. Genetic dissection of this kind of diabetogenesis will increase our understanding of the complex gene-gene interaction and mode of inheritance in human type 2 diabetes.
Physical Exercise-Induced Hyperinsulinemic Hypoglycemia Is an Autosomal-Dominant Trait Characterized by Abnormal Pyruvate-Induced Insulin Release
Authors: Otonkoski Timo , Kaminen Nina , Ustinov Jarkko , Lapatto Risto , Meissner Thomas , Mayatepek Ertan , Kere Juha , Sipilä Ilkka
Published: March 2007

We have identified patients in whom strenuous physical exercise leads to hypoglycemia caused by inappropriate insulin release (exercise-induced hyperinsulinism [EIHI]). The aim of the present study was to test the hypothesis that the increased levels of lactate and/or pyruvate during anaerobic exercise would trigger the aberrant insulin secretion in these patients. A total of 12 patients (8 women and 4 men from two families) were diagnosed with EIHI, based on hypoglycemia and a more than threefold increase in plasma insulin induced by a 10-min bicycle exercise test. The mode of inheritance was autosomal dominant in these families. The acute response of insulin release to a bolus of intravenous pyruvate (13.9 mmol/1.73 m2) was studied in the patients and eight healthy control subjects. Insulin secretion did not respond to the pyruvate bolus in healthy control subjects. However, all EIHI patients responded to pyruvate, displaying a brisk increase in plasma insulin. The 1 + 3-min peak response was 5.6-fold in the patients and 0.9-fold in the control subjects (P < 0.001). To test the hypothesis that the pathogenesis of EIHI would involve monocarboxylate transport or metabolism in the β-cell, we sequenced the genes encoding the known monocarboxylate transporter proteins and tested the transport of pyruvate into patient fibroblasts. The results revealed normal coding sequences and pyruvate transport. In conclusion, EIHI represents a new autosomal-dominant hyperinsulinemia syndrome that may be more common than has been realized. The pyruvate test provides a simple, safe, and specific diagnostic test for this condition.
Variation in Resistin Gene Promoter Not Associated With Polycystic Ovary Syndrome
Authors: Urbanek Margrit , Du Yangzhu , Silander Kaisa , Collins Francis S. , Steppan Claire M. , Strauss Jerome F. , Dunaif Andrea , Spielman Richard S. , Legro Richard S.
Published: March 2007

Polycystic ovary syndrome (PCOS) is a leading cause of anovulatory infertility and affects ∼4–7% of reproductive age women in the U.S. It is characterized by hyperandrogenemia and chronic anovulation and is associated with insulin resistance, obesity, and increased risk for type 2 diabetes. In a screen of candidate genes, a region on chromosome 19p13.3 was identified that shows significant evidence for both linkage and association with PCOS. A promising candidate gene for PCOS, resistin, maps to exactly this region. Resistin is a protein hormone thought to modulate glucose tolerance and insulin action. We tested for association between a single nucleotide polymorphism in the promoter region of the resistin gene and three phenotypes: PCOS, obesity, and insulin resistance. We did not find evidence for association with any of the phenotypes. It is therefore unlikely that variation in the resistin gene accounts for the strong association that we observe between chromosome 19p13.3 and PCOS. Instead, this association is most likely due to a gene or genetic element in this region that has not been identified.
Dietary Cod Protein Restores Insulin-Induced Activation of Phosphatidylinositol 3-Kinase/Akt and GLUT4 Translocation to the T-Tubules in Skeletal Muscle of High-Fat-Fed Obese Rats
Authors: Tremblay Frédéric , Lavigne Charles , Jacques Hélène , Marette André
Published: March 2007

Diet-induced obesity is known to cause peripheral insulin resistance in rodents. We have recently found that feeding cod protein to high-fat-fed rats prevents the development of insulin resistance in skeletal muscle. In the present study, we have further explored the cellular mechanisms behind this beneficial effect of cod protein on skeletal muscle insulin sensitivity. Rats were fed a standard chow diet or a high-fat diet in which the protein source was either casein, soy, or cod proteins for 4 weeks. Whole-body and muscle glucose disposal were reduced by ∼50% in rats fed high-fat diets with casein or soy proteins, but these impairments were not observed in animals fed cod protein. Insulin-induced tyrosine phosphorylation of the insulin receptor and insulin receptor substrate (IRS) proteins were similar in muscle of chow- and high-fat-fed rats regardless of the dietary protein source. However, IRS-1-associated phosphatidylinositol (PI) 3-kinase activity was severely impaired (−60%) in muscle of high-fat-fed rats consuming casein or soy protein. In marked contrast, feeding rats with cod protein completely prevented the deleterious effect of fat feeding on insulin-stimulated PI 3-kinase activity. The activation of the downstream kinase Akt/PKB by insulin, assessed by in vitro kinase assay and phosphorylation of GSK-3β, were also impaired in muscle of high-fat-fed rats consuming casein or soy protein, but these defects were also fully prevented by dietary cod protein. However, no effect of cod protein was observed on atypical protein kinase C activity. Normalization of PI 3-kinase/Akt activation by insulin in rats fed high-fat diets with cod protein was associated with improved translocation of GLUT4 to the T-tubules but not to the plasma membrane. Taken together, these results show that dietary cod protein is a natural insulin-sensitizing agent that appears to prevent obesity-linked muscle insulin resistance by normalizing insulin activation of the PI 3-kinase/Akt pathway and by selectively improving GLUT4 translocation to the T-tubules.