bluuebunny/crossref_metadata_2025 huggingface dataset:
Resistance to Exercise-Induced Increase in Glucose Uptake During Hyperinsulinemia in Insulin-Resistant Skeletal Muscle of Patients With Type 1 Diabetes
Authors: Peltoniemi Pauliina , Yki-Järvinen Hannele , Oikonen Vesa , Oksanen Airi , Takala Teemu O. , Rönnemaa Tapani , Erkinjuntti Matti , Knuuti M. Juhani , Nuutila Pirjo
Published: March 2007

Insulin and exercise have been shown to activate glucose transport at least in part via different signaling pathways. However, it is unknown whether insulin resistance is associated with a defect in the ability of an acute bout of exercise to enhance muscle glucose uptake in vivo. We compared the abilities of insulin and isometric exercise to stimulate muscle blood flow and glucose uptake in 12 men with type 1 diabetes (age 24 ± 1 years, BMI 23.0 ± 0.4 kg/m2) and in 11 age- and weight-matched nondiabetic men (age 25 ± 1 years, BMI 22.3 ± 0.6 kg/m2) during euglycemic hyperinsulinemia (1 mU · kg−1 · min−1 insulin infusion for 150 min). One-legged exercise was performed at an intensity of 10% of maximal isometric force for 105 min (range 45–150). Rates of muscle blood flow, oxygen consumption, and glucose uptake were quantitated simultaneously in both legs using [15O]water, [15O]oxygen, [18F]-2-fluoro-2-deoxy-d-glucose, and positron emission tomography. Resting rates of oxygen consumption were similar during hyperinsulinemia between the groups (2.4 ± 0.3 vs. 2.0 ± 0.5 ml · kg−1 muscle · min−1; normal subjects versus patients with type 1 diabetes, NS), and exercise increased oxygen consumption similarly in both groups (25.3 ± 4.3 vs. 20.1 ± 3.0 ml · kg−1 muscle · min−1, respectively, NS). Rates of insulin-stimulated muscle blood flow and the increments in muscle blood flow induced by exercise were also similar in normal subjects (129 ± 14 ml · kg−1 · min−1) and in patients with type 1 diabetes (115 ± 12 ml · kg−1 · min−1). The patients with type 1 diabetes exhibited resistance to both insulin stimulation of glucose uptake (34 ± 6 vs. 76 ± 9 μmol · kg−1 muscle · min−1, P < 0.001) and also to the exercise-induced increment in glucose uptake (82 ± 15 vs. 162 ± 29 μmol · kg−1 muscle · min−1, P < 0.05). We conclude that the ability of exercise to increase insulin-stimulated glucose uptake in vivo is blunted in patients with insulin-resistant type 1 diabetes compared with normal subjects. This could be caused by either separate or common defects in exercise- and insulin-stimulated pathways.
Effects of Novel Polymorphisms in the RAGE Gene on Transcriptional Regulation and Their Association With Diabetic Retinopathy
Authors: Hudson Barry I. , Stickland Max H. , Futers T. Simon , Grant Peter J.
Published: March 2007

Interactions between advanced glycation end products (AGEs) and the receptor for AGE (RAGE) are implicated in the vascular complications in diabetes. We have identified eight novel polymorphisms, of which the −1420 (GGT)n, −1393 G/T, −1390 G/T, and −1202 G/A were in the overlapping PBX2 3′ untranslated region (UTR), and the −429 T/C (66.5% TT, 33.5% TC/CC), −407 to –345 deletion (99% I, 1% I/D, 0% D), −374 T/A (66.4% TT, 33.6% TA/AA), and +20 T/A were in the RAGE promoter. To evaluate the effects on transcriptional activity, we measured chloramphenicol acetyl transferase (CAT) reporter gene expression, driven by variants of the –738 to +49 RAGE gene fragment containing the four polymorphisms identified close to the transcriptional start site. The –429 C, −374 A, and 63-bp deletion alleles resulted in a mean increase of CAT expression of twofold (P < 0.0001), threefold (P < 0.001), and fourfold (P < 0.05), respectively, with the –374 T and A alleles yielding highly differential binding of nuclear protein extract from both monocyte- and hepatocyte-derived cell lines. The prevalence of the functional polymorphisms were investigated in subjects with type 2 diabetes (106 with and 109 without retinopathy), with the –429 C allele showing an increase in the retinopathy group (P < 0.05). These data suggest that the polymorphisms involved in differences in RAGE gene regulation may influence the pathogenesis of diabetic vascular complications.
Epidermal Growth Factor Increases Undifferentiated Pancreatic Embryonic Cells In Vitro
Authors: Cras-Méneur Corentin , Elghazi Lynda , Czernichow Paul , Scharfmann Raphael
Published: March 2007

During embryonic life, the development of a proper mass of mature pancreatic tissue is thought to require the proliferation of precursor cells, followed by their differentiation into endocrine or acinar cells. We investigated whether perturbing the proliferation of precursor cells in vitro could modify the final mass of endocrine tissue that develops. For that purpose, we used activators or inhibitors of signals mediated by receptor tyrosine kinases. We demonstrated that when embryonic day 13.5 rat pancreatic epithelium is cultured in the presence of PD98059, an inhibitor of the mitogen-activated protein (MAP) kinase, epithelial cell proliferation is decreased, whereas endocrine cell differentiation is activated. On the other hand, in the presence of epidermal growth factor (EGF), an activator of the MAP kinase pathway, the mass of tissue that develops is increased, whereas the absolute number of endocrine cells that develops is decreased. Under this last condition, a large number of epithelial cells proliferate but remain undifferentiated. In a second step, when EGF is removed from the pool of immature pancreatic epithelial cells, the cells differentiate en masse into insulin-expressing cells. The total number of insulin-expressing cells that develop can thus be increased by first activating the proliferation of immature epithelial cells with growth factors, thus allowing an increase in the pool of precursor cells, and next allowing their differentiation into endocrine cells by removing the growth factor. This strategy suggests a possible tissue engineering approach to expanding β-cells.
Gamma Globulin Levels Predict Type 2 Diabetes in the Pima Indian Population
Authors: Lindsay Robert S. , Krakoff Jonathan , Hanson Robert L. , Bennett Peter H. , Knowler William C.
Published: March 2007

It has been proposed that inflammation or infection may contribute to the development of type 2 diabetes. We examined whether serum gamma globulin, a nonspecific measure of the humoral immune system, predicted changes in glucose tolerance in 2,530 members of the Pima Indian population, a group with a marked predisposition to type 2 diabetes. Cross-sectionally, gamma globulin was positively related to age (r = 0.08, P < 0.0005), BMI (r = 0.09; P < 0.0001), and female sex (P < 0.0001). Gamma globulin concentrations were familial, being positively correlated among siblings (r = 0.23; P < 0.0001) and between parents and their children (mother/child: r = 0.17, P < 0.0001; father/child: r = 0.25, P < 0.0001). Gamma globulin concentrations were higher with greater degrees of American Indian heritage (P < 0.004, with adjustment for age, sex, and BMI) and in the presence of a family history of type 2 diabetes (P < 0.04). Higher gamma globulin levels predicted risk of diabetes. In univariate analysis, a 1 SD difference in gamma globulin was associated with a 20% higher incidence of diabetes in those who were normal glucose tolerant at baseline (hazard rate ratio 1.20 [CI 1.11–1.30]; P < 0.0001) and remained as a significant predictor of diabetes, even when controlled for effects of sex, BMI, and 2-h glucose as additional predictors (hazard rate ratio for 1 SD difference in gamma globulin, 1.14 [1.05–1.24]; P = 0.002). Gamma globulin was also associated in univariate analysis with later development of impaired glucose tolerance (IGT) (hazard rate ratio 1.15 [1.07–1.23]; P < 0.0001), but not with the transition from IGT to diabetes (hazard rate ratio 1.04 [0.90–1.20]; P = 0.6). Thus, gamma globulin levels predict increased risk of diabetes in the Pima population. We suggest that immune function or activation may play a role in the development of type 2 diabetes.
Glucose Induces β-Cell Apoptosis Via Upregulation of the Fas Receptor in Human Islets
Authors: Maedler Kathrin , Spinas Giatgen A. , Lehmann Roger , Sergeev Pavel , Weber Markus , Fontana Adriano , Kaiser Nurit , Donath Marc Y.
Published: March 2007

In autoimmune type 1 diabetes, Fas–to–Fas-ligand (FasL) interaction may represent one of the essential pro-apoptotic pathways leading to a loss of pancreatic β-cells. In the advanced stages of type 2 diabetes, a decline in β-cell mass is also observed, but its mechanism is not known. Human islets normally express FasL but not the Fas receptor. We observed upregulation of Fas in β-cells of type 2 diabetic patients relative to nondiabetic control subjects. In vitro exposure of islets from nondiabetic organ donors to high glucose levels induced Fas expression, caspase-8 and -3 activation, and β-cell apoptosis. The effect of glucose was blocked by an antagonistic anti-Fas antibody, indicating that glucose-induced apoptosis is due to interaction between the constitutively expressed FasL and the upregulated Fas. These results support a new role for glucose in regulating Fas expression in human β-cells. Upregulation of the Fas receptor by elevated glucose levels may contribute to β-cell destruction by the constitutively expressed FasL independent of an autoimmune reaction, thus providing a link between type 1 and type 2 diabetes.
A Preprandial Rise in Plasma Ghrelin Levels Suggests a Role in Meal Initiation in Humans
Authors: Cummings David E. , Purnell Jonathan Q. , Frayo R. Scott , Schmidova Karin , Wisse Brent E. , Weigle David S.
Published: March 2007

The recently discovered orexigenic peptide ghrelin is produced primarily by the stomach and circulates in blood at levels that increase during prolonged fasting in rats. When administered to rodents at supraphysiological doses, ghrelin activates hypothalamic neuropeptide Y/agouti gene–related protein neurons and increases food intake and body weight. These findings suggest that ghrelin may participate in meal initiation. As a first step to investigate this hypothesis, we sought to determine whether circulating ghrelin levels are elevated before the consumption of individual meals in humans. Ghrelin, insulin, and leptin were measured by radioimmunoassay in plasma samples drawn 38 times throughout a 24-h period in 10 healthy subjects provided meals on a fixed schedule. Plasma ghrelin levels increased nearly twofold immediately before each meal and fell to trough levels within 1 h after eating, a pattern reciprocal to that of insulin. Intermeal ghrelin levels displayed a diurnal rhythm that was exactly in phase with that of leptin, with both hormones rising throughout the day to a zenith at 0100, then falling overnight to a nadir at 0900. Ghrelin levels sampled during the troughs before and after breakfast correlated strongly with 24-h integrated area under the curve values (r = 0.873 and 0.954, respectively), suggesting that these convenient, single measurements might serve as surrogates for 24-h profiles to estimate overall ghrelin levels. Circulating ghrelin also correlated positively with age (r = 0.701). The clear preprandial rise and postprandial fall in plasma ghrelin levels support the hypothesis that ghrelin plays a physiological role in meal initiation in humans.
T-Cell Epitope Analysis on the Autoantigen Phogrin (IA-2β) in the Nonobese Diabetic Mouse
Authors: Kelemen Katalin , Wegmann Dale R. , Hutton John C.
Published: March 2007

The protein tyrosine phosphatases (PTPs) IA-2 and phogrin (IA-2β) are major autoantigens in type 1 diabetes that possess common serological epitopes in their COOH termini. The epitopes recognized by the T-cells that cause the disease, however, remain to be defined. Eight phogrin-specific T-cell clones were generated from NOD mice, and their epitopes were mapped. The mapping was performed initially with recombinant gluthathione S-transferase–phogrin COOH deletion constructs and ultimately with overlapping synthetic peptides. Two dominant epitopes were identified: one (aa 629–649) immediately adjacent to the transmembrane domain (aa 604–628) and the second (aa 755–777) lying in the NH2-terminal region of the conserved PTP domain. T-cells that are specific to either of these peptides and that could destroy islet tissue in vivo though spontaneous T-cell proliferative responses were observed in prediabetic female NOD splenocytes only to the aa 755–777 epitope. In NOD female mice immunized with the epitope peptide, intramolecular determinant spreading occurred from the aa 629–649 epitope to the aa 755–777 epitope but not in the opposite direction. We concluded that the initial T-cell response to phogrin is restricted to a small number of dominant peptides and that it subsequently spreads to other regions of the molecule, including those containing the major humoral epitopes that are highly conserved between IA-2 and phogrin.
Linear Correlation Between the Total Islet Mass and the Volume-Weighted Mean Islet Volume
Authors: Skau Maren , Pakkenberg Bente , Buschard Karsten , Bock Troels
Published: March 2007

To understand the dynamics of islet population, especially during conditions with growth of the total islet mass, it is important to have reliable estimators of parameters describing the quantitative appearance of the islet population. We describe a stereological estimator of the volume-weighted mean islet volume based on unbiased assumption-free stereological principles. The volume-weighted mean islet volume is the mean islet volume if the islets are weighted (sampled) proportional to their volume. This method allows simultaneously unbiased estimation of the total islet mass. With use of this method, 22 male Sprague-Dawley rats within the age span of 34–102 days old were investigated. We found a linear correlation (P < 0.001) between total islet mass and the volume-weighted mean islet volume. The results support models demonstrating that the physiological growth of the total islet mass in the period studied is totally or mainly caused by proportional growth of existing islets. The functional meaning of the volume-weighted mean islet volume is discussed, and previous methods to study the mean islet volume and islet number are critically evaluated. We propose the volume-weighted mean islet volume to be a biologically useful parameter when describing the mean volume of the pancreatic islets and investigating the differences between experimental groups.
Glycerol-Stimulated Proinsulin Biosynthesis in Isolated Pancreatic Rat Islets via Adenoviral-Induced Expression of Glycerol Kinase Is Mediated via Mitochondrial Metabolism
Authors: Skelly Robert H. , Wicksteed Barton , Antinozzi Peter A. , Rhodes Christopher J.
Published: March 2007

In this study, we examined whether adenoviral-mediated glycerol kinase (AdV-CMV-GlyK) expression in isolated rat pancreatic islets could introduce glycerol-induced proinsulin biosynthesis. In AdV-CMV-GlyK–infected islets, specific glycerol-induced proinsulin biosynthesis translation and insulin secretion were observed in parallel from the same islets. The threshold concentration of glycerol required to stimulate proinsulin biosynthesis was lower (0.25–0.5 mmol/l) than that for insulin secretion (1.0–1.5 mmol/l), reminiscent of threshold differences for glucose-stimulated proinsulin biosynthesis versus insulin secretion. The dose-dependent glycerol-induced proinsulin biosynthesis correlated with the rate of glycerol oxidation in AdV-CMV-GlyK–infected islets, indicating that glycerol metabolism was required for the response. However, glycerol did not significantly increase lactate output from AdV-CMV-GlyK–infected islets, but the dihydroxyacetone phosphate (DHAP) to α-glycerophosphate (α-GP) ratio significantly increased in AdV-CMV-GlyK–infected islets incubated at 2 mmol/l glycerol compared with that at a basal level of 2.8 mmol/l glucose (P ≤ 0.05). The DHAP:α-GP ratio was unaffected in AdV-CMV-GlyK–infected islets incubated at 2 mmol/l glycerol in the added presence of α-cyanohydroxycinnaminic acid (α-CHC), an inhibitor of the plasma membrane and mitochondrial lactate/pyruvate transporter. However, α-CHC inhibited glycerol-induced proinsulin biosynthesis and insulin secretion in AdV-CMV-GlyK–infected islets (>75%; P = 0.05), similarly to glucose-induced proinsulin biosynthesis and insulin secretion in AdV-CMV-GlyK–infected and control islets. These data indicated that in AdV-CMV-GlyK–infected islets, the importance of mitochondrial metabolism of glycerol was required to generate stimulus–response coupling signals to induce proinsulin biosynthesis and insulin secretion.
Treatment of Type 2 Diabetes by Adenoviral-Mediated Overexpression of the Glucokinase Regulatory Protein
Authors: Slosberg Eric D. , Desai Urvi J. , Fanelli Barbara , St. Denny Irene , Connelly Sheila , Kaleko Michael , Boettcher Brian R. , Caplan Shari L.
Published: March 2007

The enzyme glucokinase (GK) plays a central role in glucose homeostasis. Hepatic GK activity is acutely controlled by the action of the GK regulatory protein (GKRP). In vitro evidence suggests that GKRP reversibly binds to GK and inhibits its activity; however, less is known about the in vivo function of GKRP. To further explore the physiological role of GKRP in vivo, we used an E1/E2a/E3-deficient adenoviral vector containing the cDNA encoding human GKRP (Av3hGKRP). High fat diet–induced diabetic mice were administered Av3hGKRP or a control vector lacking a transgene (Av3Null). Surprisingly, the Av3hGKRP-treated mice showed a significant improvement in glucose tolerance and had lower fasting blood glucose levels than Av3Null-treated mice. A coincident decrease in insulin levels indicated that the Av3hGKRP-treated mice had sharply improved insulin sensitivity. These mice also exhibited lower leptin levels, reduced body weight, and decreased liver GK activity. In vitro experiments indicated that GKRP was able to increase both GK protein and enzymatic activity levels, suggesting that another role for GKRP is to stabilize and/or protect GK. These data are the first to indicate the ability of GKRP to treat type 2 diabetes and therefore have significant implications for future therapies of this disease.
Small Increases in Insulin Inhibit Hepatic Glucose Production Solely Caused by an Effect on Glycogen Metabolism
Authors: Edgerton Dale S. , Cardin Sylvain , Emshwiller Maya , Neal Doss , Chandramouli Visvanathan , Schumann William C. , Landau Bernard R. , Rossetti Luciano , Cherrington Alan D.
Published: March 2007

Based on our earlier work, a 2.5-fold increase in insulin secretion should completely inhibit hepatic glucose production through the hormone’s direct effect on hepatic glycogen metabolism. The aim of the present study was to test the accuracy of this prediction and to confirm that gluconeogenic flux, as measured by three independent techniques, was unaffected by the increase in insulin. A 40-min basal period was followed by a 180-min experimental period in which an increase in insulin was induced, with euglycemia maintained by peripheral glucose infusion. Arterial and hepatic sinusoidal insulin levels increased from 10 ± 2 to 19 ± 3 and 20 ± 4 to 45 ± 5 μU/ml, respectively. Net hepatic glucose output decreased rapidly from 1.90 ± 0.13 to 0.23 ± 0.16 mg · kg−1 · min−1. Three methods of measuring gluconeogenesis and glycogenolysis were used: 1) the hepatic arteriovenous difference technique (n = 8), 2) the [14C]phosphoenolpyruvate technique (n = 4), and 3) the 2H2O technique (n = 4). The net hepatic glycogenolytic rate decreased from 1.72 ± 0.20 to −0.28 ± 0.15 mg · kg−1 · min−1 (P < 0.05), whereas none of the above methods showed a significant change in hepatic gluconeogenic flux (rate of conversion of phosphoenolpyruvate to glucose-6-phosphate). These results indicate that liver glycogenolysis is acutely sensitive to small changes in plasma insulin, whereas gluconeogenic flux is not.
Blood-to-Brain Glucose Transport, Cerebral Glucose Metabolism, and Cerebral Blood Flow Are Not Increased After Hypoglycemia
Authors: Segel Scott A. , Fanelli Carmine G. , Dence Carmen S. , Markham Joanne , Videen Tom O. , Paramore Deanna S. , Powers William J. , Cryer Philip E.
Published: March 2007

Recent antecedent hypoglycemia has been found to shift glycemic thresholds for autonomic (including adrenomedullary epinephrine), symptomatic, and other responses to subsequent hypoglycemia to lower plasma glucose concentrations. This change in threshold is the basis of the clinical syndromes of hypoglycemia unawareness and, in part, defective glucose counterregulation and the unifying concept of hypoglycemia-associated autonomic failure in type 1 diabetes. We tested in healthy young adults the hypothesis that recent antecedent hypoglycemia increases blood-to-brain glucose transport, a plausible mechanism of this phenomenon. Eight subjects were studied after euglycemia, and nine were studied after ∼24 h of interprandial hypoglycemia (∼55 mg/dl, ∼3.0 mmol/l). The latter were shown to have reduced plasma epinephrine (P = 0.009), neurogenic symptoms (P = 0.009), and other responses to subsequent hypoglycemia. Global bihemispheric blood-to-brain glucose transport and cerebral glucose metabolism were calculated from rate constants derived from blood and brain time-activity curves—the latter determined by positron emission tomography (PET)—after intravenous injection of [1-11C]glucose at clamped plasma glucose concentrations of 65 mg/dl (3.6 mmol/l). For these calculations, a model was used that includes a fourth rate constant to account for egress of [11C] metabolites. Cerebral blood flow was measured with intravenous [15O]water using PET. After euglycemia and after hypoglycemia, rates of blood-to-brain glucose transport (24.6 ± 2.3 and 22.4 ± 2.4 μmol · 100 g−1 · min−1, respectively), cerebral glucose metabolism (16.8 ± 0.9 and 15.9 ± 0.9 μmol · 100 g−1 · min−1, respectively) and cerebral blood flow (56.8 ± 3.9 and 53.3 ± 4.4 ml · 100 g−1 · min−1, respectively) were virtually identical. These data do not support the hypothesis that recent antecedent hypoglycemia increases blood-to-brain glucose transport during subsequent hypoglycemia. They do not exclude regional increments in blood-to-brain glucose transport. Alternatively, the fundamental alteration might lie beyond the blood-brain barrier.
Angiotensin II Promotes Glucose-Induced Activation of Cardiac Protein Kinase C Isozymes and Phosphorylation of Troponin I
Authors: Malhotra Ashwani , Kang Barinder P.S. , Cheung Simon , Opawumi David , Meggs Leonard G.
Published: March 2007

Activation of the protein kinase C (PKC) family is a potential signaling mechanism by which high ambient glucose concentration modulates the phenotype and physiological function of cells. Recently, the cardiac renin angiotensin system (RAS) has been reported to promote PKC translocation in the diabetic heart via the angiotensin (ANG) II type 1 receptor (AT-1R). To evaluate the molecular events coupled with high glucose−induced PKC translocation and to examine the role of endogenously released ANG II in myocyte PKC signaling, primary cultures of adult rat ventricular myocytes were exposed to normal (5 mmol/l) or high (25 mmol/l) glucose for 12–24 h. Western blot analysis indicated that adult rat ventricular myocytes coexpress six PKC isozymes (α, β1, β2, δ, ε, and ζ). Translocation of five PKC isozymes (β1, β2, δ, ε, and ζ) was detected in response to 25 mmol/l glucose. Inhibition of phospholipase C with tricyclodecan-9-yl-xanthogenate blocked glucose-induced translocation of PKC-β2, -δ, and -ζ. Inhibition of tyrosine kinase with genistein blocked glucose-induced translocation of PKC-β1 and -δ, whereas chelation of intracellular Ca2+ with 1,2-bis(2-aminophenoxy)ethane N,N,N,’N′-tetraacetic acid blocked translocation of PKC-β1 and -β2. Enzyme-linked immunosorbent assay performed on culture media from myocytes maintained in 25 mmol/l glucose detected a twofold increase in ANG II. Addition of an AT-1R antagonist (losartan; 100 nmol/l) to myocyte cultures blocked translocation of PKC-β1, -β2, -δ, and -ε. Phosphorylation of troponin (Tn) I was increased in myocytes exposed to 25 mmol/l glucose. Losartan selectively inhibited Tn I serine phosphorylation but did not affect phosphorylation at threonine residues. We concluded that 1) 25 mmol/l glucose triggers the release of ANG II by myocytes, resulting in activation of the ANG II autocrine pathway; 2) differential translocation of myocyte PKC isozymes occurs in response to 25 mmol/l glucose and ANG II; and 3) AT-1R−dependent PKC isozymes (β1, β2, δ, and ε) target Tn I serine residues.
Effect of Antioxidant Treatment of Streptozotocin-Induced Diabetic Rats on Endoneurial Blood Flow, Motor Nerve Conduction Velocity, and Vascular Reactivity of Epineurial Arterioles of the Sciatic Nerve
Authors: Coppey Lawrence J. , Gellett Jill S. , Davidson Eric P. , Dunlap Joyce A. , Lund Donald D. , Yorek Mark A.
Published: March 2007

We have shown that diabetes-induced reduction in endoneurial blood flow (EBF) and impaired endothelium-dependent vascular relaxation precede slowing of motor nerve conduction velocity (MNCV) and decreased sciatic nerve Na+/K+ ATPase activity. Furthermore, vascular dysfunction was accompanied by an accumulation of superoxide in arterioles that provide circulation to the sciatic nerve. In the present study, we examined the effect that treatment of streptozotocin-induced diabetic rats with antioxidants has on vascular and neural function. Diabetic rats were treated with 0.5% α-lipoic acid as a diet supplement or with hydroxyethyl starch deferoxamine (HES-DFO) by weekly intravenous injections at a dose of 75 mg/kg. The treatments significantly improved diabetes-induced decrease in EBF, acetylcholine-mediated vascular relaxation in arterioles that provide circulation to the region of the sciatic nerve, and MNCV. The treatments also reduced the production of superoxide by the aorta and superoxide and peroxynitrite by arterioles that provide circulation to the region of the sciatic nerve. Treating diabetic rats with α-lipoic acid prevented the diabetes-induced increase in thiobarbituric acid–reactive substances in serum and significantly improved lens glutathione levels. In contrast, treating diabetic rats with HES-DFO did not prevent diabetes-induced changes of either of these markers of oxidative stress. Diabetes-induced increase in sciatic nerve conjugated diene levels was not improved by treatment with either α-lipoic acid or HES-DFO. Treating diabetic rats with α-lipoic acid but not HES-DFO partially improved sciatic nerve Na+/K+ ATPase activity and myo-inositol content. The increase in sciatic nerve sorbitol levels in diabetic rats was unchanged by either treatment. These studies suggest that diabetes-induced oxidative stress and the generation of superoxide may be partially responsible for the development of diabetic vascular and neural complications.
Abnormalities of Retinal Metabolism in Diabetes and Experimental Galactosemia
Authors: Kowluru Renu A. , Tang Jie , Kern Timothy S.
Published: March 2007

Antioxidants were administered to diabetic rats and experimentally galactosemic rats to evaluate the ability of these agents to inhibit the development of diabetic retinopathy. Alloxan diabetic rats and nondiabetic rats that were fed 30% galactose randomly received standard diets or the diets supplemented with ascorbic acid and α-tocopherol (vitamins C+E diet) or a more comprehensive mixture of antioxidants (multi-antioxidant diet), including Trolox, α-tocopherol, N-acetyl cysteine, ascorbic acid, β-carotene, and selenium. Diabetes or galactose feeding of at least 12 months resulted in pericyte loss, acellular capillaries, and basement membrane thickening. Compared with diabetic controls, the development of acellular capillaries was inhibited by 50% (P < 0.05) in diabetic rats that received supplemental vitamins C+E, and the number of pericyte ghosts tended to be reduced. The vitamins C+E supplement had no beneficial effect in galactosemic rats, but these rats consumed only approximately half as much of the antioxidants as the diabetic rats. The multi-antioxidant diet significantly inhibited (∼55–65%) formation of both pericyte ghosts and acellular capillaries in diabetic rats and galactosemic rats (P < 0.05 vs. controls), without affecting the severity of hyperglycemia. Parameters of retinal oxidative stress, protein kinase C activity, and nitric oxides remained elevated for at least 1 year of hyperglycemia, and these abnormalities were normalized by multi-antioxidant therapy. Thus, long-term administration of antioxidants can inhibit the development of the early stages of diabetic retinopathy, and the mechanism by which this action occurs warrants further investigation. Supplementation with antioxidants can offer an achievable and inexpensive adjunct therapy to help inhibit the development of retinopathy in diabetes.
Susceptibility and Negative Epistatic Loci Contributing to Type 2 Diabetes and Related Phenotypes in a KK/Ta Mouse Model
Authors: Shike Toshihide , Hirose Sachiko , Kobayashi Michimasa , Funabiki Kazuhiko , Shirai Toshikazu , Tomino Yasuhiko
Published: March 2007

The KK/Ta mouse strain serves as a suitable polygenic model for human type 2 diabetes. Using 93 microsatellite markers in 208 KK/Ta × (BALB/c × KK/Ta)F1 male backcross mice, we carried out a genome-wide linkage analysis of KK/Ta alleles contributing to type 2 diabetes and related phenotypes, such as obesity and dyslipidemia. We identified three major chromosomal intervals significantly contributing to impaired glucose metabolism: one quantitative trait locus for impaired glucose tolerance on chromosome 6 and two loci for fasting blood glucose levels on chromosomes 12 and 15. The latter two loci appeared to act in a complementary fashion. Two intervals showed significant linkages for serum triglyceride levels, one on chromosome 4 and the other on chromosome 8. The KK allele on chromosome 8 acts to promote serum triglyceride levels, whereas the KK allele on chromosome 4 acts to suppress this effect in a recessive fashion. In addition, it is suggested that the chromosome 4 locus also acts to downregulate body weight and that the chromosome 8 locus acts to upregulate serum insulin levels. Our data clearly showed that each disease phenotype of type 2 diabetes and related disorders in KK/Ta mice is under the control of separate genetic mechanisms. However, there appear to be common genes contributing to different disease phenotypes. There are potentially important candidate genes that may be relevant to the disease.
Heme Oxygenase-1 Induction in Islet Cells Results in Protection From Apoptosis and Improved In Vivo Function After Transplantation
Authors: Pileggi Antonello , Molano R. Damaris , Berney Thierry , Cattan Pierre , Vizzardelli Caterina , Oliver Robert , Fraker Christopher , Ricordi Camillo , Pastori Ricardo L. , Bach Fritz H. , Inverardi Luca
Published: March 2007

Transplantation of islets of Langerhans represents a viable therapeutic approach for the treatment of type 1 diabetes. Unfortunately, transplanted islets are susceptible to allogeneic recognition and rejection, recurrence of autoimmunity, and destruction by local inflammation at the site of implantation. The last of these phenomena might not only result in functional impairment and death of islet cells but could also contribute to amplifying the subsequent specific immune response. Induction of islet cell protection against inflammation could therefore be postulated to be a powerful means to improve overall graft fate. Heme oxygenase-1 (HO-1) has been described as an inducible protein capable of cytoprotection via radical scavenging and apoptosis prevention. The purpose of the present study was to analyze whether HO-1 upregulation in a β-cell line and in freshly isolated murine islets could result in protection from apoptosis and improve in vivo functional performance. HO-1 upregulation was induced reproducibly with protoporphyrins and was correlated with protection from apoptosis induced in vitro with proinflammatory cytokines or Fas engagement. Furthermore, in vivo HO-1 upregulation resulted in improved islet function in a model of marginal mass islet transplantation in rodents. Strategies aimed at inducing HO-1 upregulation might result in improved success in islet transplantation.
Autoreactive Diabetogenic T-Cells in NOD Mice Can Efficiently Expand From a Greatly Reduced Precursor Pool
Authors: Serreze David V. , Johnson Ellis A. , Chapman Harold D. , Graser Robert T. , Marron Michele P. , DiLorenzo Teresa P. , Silveira Pablo , Yoshimura Yoshitaka , Nathenson Stanley G. , Joyce Sebastian
Published: March 2007

A broad repertoire of pancreatic β-cell autoreactive T-cells normally contributes to the development of type 1 diabetes in NOD mice. However, it has been unknown if a large reduction in the precursor pool from which autoreactive T-cells are drawn would inhibit the development of type 1 diabetes. To address this issue, we reduced the precursor frequency of autoreactive T-cells in NOD mice through allelic exclusion induced by transgenic expression of an H2-Db class I−restricted T-cell receptor (TCR) specific for a pathologically irrelevant lymphocytic choriomeningitis virus (LCMV) peptide. TCR allelic exclusion greatly reduced the pool of T-cells from which diabetogenic effectors could be derived in these NOD.LCMV TCR Tg mice. Surprisingly, this did not impair their type 1 diabetes susceptibility. Furthermore, a diabetogenic CD8 T-cell population that is prevalent in standard NOD mice was present at essentially equivalent levels in pancreatic islets of NOD.LCMV TCR Tg mice. Other data indicated that the antigenic specificity of these CD8 T-cells is primarily the function of a shared TCR-α chain. Although the percentage of TCR transgenic T-cells decreased in NOD versus B6,D2 control mice, much higher total numbers of both the TCR transgenic and the nontransgenic T-cells accumulated in the NOD strain. This transgenic T-cell accumulation in the absence of the cognate peptide indicated that the NOD genetic background preferentially promotes a highly efficient antigen-independent T-cell expansion. This might allow diabetogenic T-cells in NOD mice to undergo an efficient expansion before encountering antigen, which would represent an important and previously unconsidered aspect of pathogenesis.
A Low-Affinity Ca2+-Dependent Association of Calmodulin With the Rab3A Effector Domain Inversely Correlates With Insulin Exocytosis
Authors: Kajio Hiroshi , Olszewski Scott , Rosner Philip J. , Donelan Matthew J. , Geoghegan Kieran F. , Rhodes Christopher J.
Published: March 2007

The stimulus-response coupling pathway for glucose-regulated insulin secretion has implicated a rise in cytosolic [Ca2+]i as a key factor to induce insulin exocytosis. However, it is unclear how elevated [Ca2+]i communicates with the pancreatic β-cell’s exocytotic apparatus. As Rab3A is a model protein involved in regulated exocytosis, we have focused on its role in regulating insulin exocytosis. By using a photoactivatable cross-linking synthetic peptide that mimics the effector domain of Rab3A and microsequence analysis, we found calmodulin to be a major Rab3A target effector protein in pancreatic β-cells. Coimmunoprecipitation analysis from pancreatic islets confirmed a Rab3A-calmodulin interaction in vivo, and that it inversely correlated with insulin exocytosis. Calmodulin affected neither GTPase nor guanine nucleotide exchange activity of Rab3A. The calmodulin-Rab3A interaction was pH- and Ca2+-dependent, and it was preferential for GTP-bound Rab3A. However, Rab3A affinity for calmodulin was relatively low (Kd = 18–22 μmol/l at 10−5 mol/l [Ca2+]) and competed by other calmodulin-binding proteins that had higher affinity (e.g., Ca2+/calmodulin-dependent protein kinase-2 [CaMK-2] {Kd = 300–400 nmol/l at 10−5 mol/l [Ca2+]}). Moreover, the Ca2+ dependence of the calmodulin-Rab3A interaction (K0.5 = 15–18 μmol/l [Ca2+], maximal at 100 μmol/l [Ca2+]) was significantly lower compared with that of the calmodulin–CaMK-2 association (K0.5 = 40 μmol/l [Ca2+], maximal at 1 mmol/l [Ca2+]). The data suggested that a transient Rab3A-calmodulin interaction might represent a means of directing calmodulin to the cytoplasmic face of a β-granule, where it can be subsequently transferred for activation of other β-granule–associated calmodulin-binding proteins as local [Ca2+]i rises to promote insulin exocytosis.
Human Fructosamine-3-Kinase
Authors: Szwergold Benjamin S. , Howell Scott , Beisswenger Paul J.
Published: March 2007

Nonenzymatic glycation appears to be an important factor in the pathogenesis of diabetic complications. Key early intermediates in this process are fructosamines, such as protein-bound fructoselysines. In this report, we describe the purification and characterization of a mammalian fructosamine-3-kinase (FN3K), which phosphorylates fructoselysine (FL) residues on glycated proteins, to FL-3-phosphate (FL3P). This phosphorylation destablilizes the FL adduct and leads to its spontaneous decomposition, thereby reversing the nonenzymatic glycation process at an early stage. FN3K was purified to homogeneity from human erythrocytes and sequenced by means of electrospray tandem mass spectrometry. The protein thus identified is a 35-kDa monomer that appears to be expressed in all mammalian tissues. It has no significant homology to other known proteins and appears to be encoded by genomic sequences located on human chromosomes 1 and 17. The lability of FL3P, the high affinity of FN3K for FL, and the wide distribution of FN3K suggest that the function of this enzyme is deglycation of nonenzymatically glycated proteins. Because the condensation of glucose and lysine residues is an ubiquitous and unavoidable process in homeothermic organisms, a deglycation system mediated by FN3K may be an important factor in protecting cells from the deleterious effects of nonenzymatic glycation. Our sequence data of FN3K are in excellent agreement with a recent report on this enzyme by Delpierre et al. (Diabetes 49:1627–1634, 2000).
Functional Significance of the UCSNP-43 Polymorphism in the CAPN10 Gene for Proinsulin Processing and Insulin Secretion in Nondiabetic Germans
Authors: Stumvoll Michael , Fritsche Andreas , Madaus Alexander , Stefan Norbert , Weisser Melanie , Machicao Fausto , Häring Hans
Published: March 2007

Recently, an association of the G allele in UCSNP-43 of calpain 10 with type 2 diabetes and decreased glucose disposal was reported. Calpain 10 is also expressed in pancreatic islets. It is not known, however, whether and how this polymorphism contributes to the biological variation of β-cell function. We studied 73 nondiabetic subjects from the southwest region of Germany (G/G, n = 41; G/A, n = 29; and A/A, n = 3) using a modified hyperglycemic clamp (10 mmol/l glucose, added glucagon-like peptide 1, final arginine bolus). The genotype distribution was not different between subjects with normal glucose tolerance (n = 56) and those with impaired glucose tolerance (n = 17; P = 0.74, χ2 test). First-phase insulin secretion (adjusted for sex and insulin sensitivity from hyperglycemic clamp) was greater in G/G (2,747 ± 297 pmol/min) than in G/A + A/A (1,612 ± 156 pmol/min, P = 0.003). Insulin secretion in response to arginine (adjusted for insulin sensitivity) was also greater in G/G (9,648 ± 1,186 pmol/min) than in G/A + A/A (5,686 ± 720 pmol/min, P = 0.04). The acute poststimulus proinsulin-to-insulin ratio was lower in G/G (1.6 ± 0.4% first phase; 1.6 ± 0.2% arginine) than in G/A + A/A (4.0 ± 0.5% first phase, P < 0.001; 2.5 ± 0.4% arginine, P = 0.03). In conclusion, it appears unlikely that any association of the UCSNP-43 polymorphism alone with type 2 diabetes involves impairment of insulin secretion in our population of German Caucasians. This may be entirely different with specific haplotype combinations.
Genetic Variation in the Peroxisome Proliferator–Activated Receptor-γ2 Gene (Pro12Ala) Affects Metabolic Responses to Weight Loss and Subsequent Weight Regain
Authors: Nicklas Barbara J. , van Rossum Elisabeth F.C. , Berman Dora M. , Ryan Alice S. , Dennis Karen E. , Shuldiner Alan R.
Published: March 2007

This study determined the effects of the peroxisome proliferator–activated receptor (PPAR)-γ2 Pro12Ala variant on body composition and metabolism and the magnitude of weight regain in 70 postmenopausal women (BMI 25–40 kg/m2) who completed 6 months of a hypocaloric diet. At baseline, BMI, percent body fat, intra-abdominal and subcutaneous abdominal fat areas, resting metabolic rate, substrate oxidation, and postprandial glucose and insulin responses were not different between genotypes (Pro/Pro = 56, Pro/Ala and Ala/Ala = 14). The intervention similarly decreased body weight by 8 ± 1% in women homozygous for the Pro allele and by 7 ± 1% in women with the Ala allele (P < 0.0001). Fat oxidation did not change in Pro/Pro women but decreased 19 ± 9% in women with the Ala allele (P < 0.05). Changes in glucose area were not different between groups; however, women with the Ala allele decreased their insulin area more than women homozygous for the Pro allele (P < 0.05). Weight regain during follow-up was greater in women with the Ala allele than women homozygous for the Pro allele (5.4 ± 0.9 vs. 2.8 ± 0.4 kg, P < 0.01). PPAR-γ2 genotype was the best predictor of weight regain (r = 0.50, P < 0.01), followed by the change in fat oxidation (partial r = 0.35, P < 0.05; cumulative r = 0.58). Thus, the Pro12Ala variant of the PPAR-γ2 gene may influence susceptibility for obesity.
Chronic Central Melanocortin-4 Receptor Antagonism and Central Neuropeptide-Y Infusion in Rats Produce Increased Adiposity by Divergent Pathways
Authors: Baran Katherine , Preston Elaine , Wilks Donna , Cooney Gregory J. , Kraegen Edward W. , Sainsbury Amanda
Published: March 2007

Increased hypothalamic neuropeptide-Y (NPY) action and disruption of the melanocortin (MC)-4 receptor both result in hyperphagia and obesity. To determine whether similar hormonal and metabolic mechanisms are involved in these two obesity syndromes, we investigated the time course of effects induced by 6-day intracerebroventricular (ICV) infusion of NPY (3.5 nmol/day) or the MC4 receptor antagonist HS014 (4.8 nmol/day) in rats pair-fed with vehicle-infused controls. The weight of white adipose tissue (WAT) deposits was increased after 6-day NPY and HS014 infusion compared with controls, and the increase was significantly greater in HS014- than in NPY-infused rats (retroperitoneal WAT: NPY 0.57 ± 0.05; HS014 0.80 ± 0.05; control 0.43 ± 0.03% body wt, n = 8–13, P < 0.05). Plasma leptin was also increased in both experimental groups (NPY 10.6 ± 1.9; HS014 4.4 ± 0.9; control 2.0 ± 0.1 ng/ml, n = 8–13, P < 0.05 for all comparisons). Basal plasma corticosterone and insulin levels were increased by ICV NPY infusion, whereas HS014-infused rats showed no significant increase in these parameters on any of 1–6 days of infusion. Both NPY and HS014 infusion potentiated intravenous glucose-induced (300 mg/kg) plasma insulin levels, and there was no difference in glycemia among groups. In NPY-infused rats, the plasma free fatty acid levels were decreased and triglyceridemia was increased compared with controls, but these parameters were unchanged in HS014-infused rats. Hepatic triglyceride content was significantly increased by HS014 but not by NPY infusion. Levels of uncoupling protein-1 mRNA in brown adipose tissue were significantly decreased after 6 days of HS014 infusion, similar to the effect of central NPY. Because ICV HS014 induced at least as great an increase in fat mass as ICV NPY and yet had divergent hormonal and metabolic effects, we conclude that MC4 receptor antagonism does not induce obesity solely by regulation of the endogenous NPY-ergic system.
Variation in the Calpain-10 Gene Affects Blood Glucose Levels in the British Population
Authors: Lynn Stephen , Evans Julie C. , White Christopher , Frayling Timothy M. , Hattersley Andrew T. , Turnbull Doug M. , Horikawa Yukio , Cox Nancy J. , Bell Graeme I. , Walker Mark
Published: March 2007

Variation in the calpain-10 gene (CAPN10) has been shown to be associated with type 2 diabetes in Mexican-Americans and in at least three Northern European populations. Studies in nondiabetic Pima Indians showed that one of the at-risk DNA polymorphisms, single-nucleotide polymorphism (SNP)-43, in CAPN10 was associated with insulin resistance, and individuals with the G/G-genotype had significantly higher fasting plasma glucose and 2-h insulin concentrations after a 75-g oral glucose tolerance test (OGTT). We have examined the effect of variation in CAPN10 on plasma glucose and insulin levels in a group of 285 nondiabetic British subjects after a 75-g OGTT. The results showed that subjects with G/G genotype at SNP-43 had higher 2-h plasma glucose levels than the combined G/A + A/A group (P = 0.05). We also examined the SNP-43, -19, and -63 haplotype combination 112/121, which is associated with an approximately threefold increased risk of diabetes. Subjects with the 112/121 haplotype combination (n = 29) had increased fasting (P = 0.004) and 2-h plasma glucose levels (P = 0.003) compared with the rest of the study population after correction for age, sex, and BMI. The 112/121 haplotype combination was also associated with a marked decrease in the insulin secretory response, adjusted for the level of insulin resistance (P = 0.002). We conclude that genetic variation in the CAPN10 gene influences blood glucose levels in nondiabetic British subjects and that this is due, at least in part, to the effects of calpain-10 on the early insulin secretory response.
Skeletal Muscle Microvascular Recruitment by Physiological Hyperinsulinemia Precedes Increases in Total Blood Flow
Authors: Vincent M.A. , Dawson D. , Clark A.D.H. , Lindner J.R. , Rattigan S. , Clark M.G. , Barrett E.J.
Published: March 2007

Supraphysiological doses of insulin enhance total limb blood flow and recruit capillaries in skeletal muscle. Whether these processes change in response to physiological hyperinsulinemia is uncertain. To examine this, we infused either saline (n = 6) or insulin (euglycemic clamp, 3.0 mU · min−1 · kg−1, n = 9) into anesthetized rats for 120 min. Femoral artery flow was monitored continuously using a Doppler flow probe, and muscle microvascular recruitment was assessed by metabolism of infused 1-methylxanthine (1-MX) and by contrast-enhanced ultrasound (CEU). Insulin infusion raised plasma insulin concentrations by ∼10-fold. Compared with saline, physiological hyperinsulinemia increased femoral artery flow (1.02 ± 0.10 vs. 0.68 ± 0.09 ml/min; P < 0.05), microvascular recruitment (measured by 1-MX metabolism [6.6 ± 0.5 vs. 4.5 ± 0.48 nmol/min; P < 0.05] as well as by CEU [167.0 ± 39.8 vs. 28.2 ± 13.8%; P < 0.01]), and microvascular flow velocity (β, 0.14 ± 0.02 vs. 0.09 ± 0.02 s−1). Subsequently, we studied the time dependency of insulin’s vascular action in a second group (n = 5) of animals. Using CEU, microvascular volume was measured at 0, 30, and 90 min of insulin infusion. Insulin augmented microvascular perfusion within 30 min (52.8 ± 14.8%), and this persisted at 90 min (64.6 ± 9.9%). Microvascular recruitment occurred without changes to femoral artery flow or β. We conclude that insulin increases tissue perfusion by recruiting microvascular beds, and at physiological concentrations this precedes increases in total muscle blood flow by 60–90 min.