bluuebunny/crossref_metadata_2025 huggingface dataset:
Characterization of the T-Cell Response to Coxsackievirus B4
Authors: Varela-Calvino Ruben , Ellis Richard , Sgarbi Gianluca , Dayan Colin M. , Peakman Mark
Published: March 2007

Most of the evidence linking enterovirus (EV) infection with the development and/or acceleration of type 1 diabetes is indirect. Few studies have examined T-cell responses to these viruses, and therefore the nature of the viral targets and the immune cells involved in antiviral responses remain unclear. In the present study, we examined the characteristics of the T-cell response to the EV Coxsackievirus B4 (CVB4) in patients with type 1 diabetes and healthy control subjects. We find that CVB4-specific T-cells preferentially target the envelope proteins VP1, VP2, and VP3, and that the response to these and other CVB4 proteins differs markedly in type 1 diabetic patients compared with nondiabetic control subjects. The frequency of T-cell proliferative responses against VP2 was significantly reduced in type 1 diabetic patients compared with control subjects, especially in patients tested near to diagnosis (P < 0.001). In contrast, median levels of γ-interferon (IFN-γ) production by T-cells in response to the CVB4 antigens tested were generally high in new-onset type 1 diabetic patients, who produced significantly higher levels in response to VP3 compared with healthy subjects (P < 0.05) and patients with long-standing disease (P < 0.05). New-onset type 1 diabetic patients also had higher levels in response to P2C compared with healthy subjects (P < 0.005) and to VP2 compared with patients with long-standing disease (P < 0.05). These results suggest that the quality of the immune response to CVB4 antigens differs significantly between type 1 diabetic patients and control subjects, with a predominance of primed effector (IFN-γ–producing) memory cells near to disease diagnosis. The data are consistent with the notion that the diagnosis of type 1 diabetes is associated with recent or persistent exposure to EV antigens.
Expression of the Vesicular Inhibitory Amino Acid Transporter in Pancreatic Islet Cells
Authors: Chessler Steven D. , Simonson William T. , Sweet Ian R. , Hammerle Lisa P.
Published: March 2007

γ-Aminobutyric acid (GABA) is stored in microvesicles in pancreatic islet cells. Because GAD65 and GAD67, which catalyze the formation of GABA, are cytoplasmic, the existence of an islet vesicular GABA transporter has been postulated. Here, we test the hypothesis that the putative transporter is the vesicular inhibitory amino acid transporter (VIAAT), a neuronal transmembrane transporter of GABA and glycine. We sequenced the human VIAAT gene and determined that the human and rat proteins share over 98% sequence identity. In vitro expression of VIAAT and immunoblotting of brain and islet lysates revealed two forms of the protein: an ∼52-kDa and an ∼57-kDa form. By immunoblotting and immunohistochemistry, we detected VIAAT in rat but not human islets. Immunohistochemical staining showed that in rat islets, the distribution of VIAAT expression parallels that of GAD67, with increased expression in the mantle. GABA, too, was found to be present in islet non-β-cells. We conclude that VIAAT is expressed in rat islets and is more abundant in the mantle and that expression in human islets is very low or nil. The rat islet mantle differs from rat and human β-cells in that it contains only GAD67 and relatively increased levels of VIAAT. Cells that express only GAD67 may require higher levels of VIAAT expression.
cFLIP Protein Prevents Tumor Necrosis Factor-α–Mediated Induction of Caspase-8–Dependent Apoptosis in Insulin-Secreting βTc-Tet Cells
Authors: Cottet Sandra , Dupraz Philippe , Hamburger Fabienne , Dolci Wanda , Jaquet Muriel , Thorens Bernard
Published: March 2007

Type 1 diabetes is characterized by the infiltration of activated leukocytes within the pancreatic islets, leading to β-cell dysfunction and destruction. The exact role played by interferon-γ, tumor necrosis factor (TNF)-α, and interleukin-1β in this pathogenic process is still only partially understood. To study cytokine action at the cellular level, we are working with the highly differentiated insulin-secreting cell line, βTc-Tet. We previously reported that it was susceptible to apoptosis induced by TNF-α, in combination with interleukin-1β and interferon-γ. Here, we report that cytokine-induced apoptosis was correlated with the activation of caspase-8. We show that in βTc-Tet cells, overexpression of cFLIP, the cellular FLICE (FADD-like IL-1β-converting enzyme)-inhibitory protein, completely abolished cytokine-dependent activation of caspase-8 and protected the cells against apoptosis. Furthermore, cFLIP overexpression increased the basal and interleukin-1β–mediated transcriptional activity of nuclear factor (NF)-κB, whereas it did not change cytokine-induced inducible nitric oxide synthase gene transcription and nitric oxide secretion. The presence of cFLIP prevented the weak TNF-α–induced reduction in cellular insulin content and secretion; however, it did not prevent the decrease in glucose-stimulated insulin secretion induced by the combined cytokines, in agreement with our previous data demonstrating that interferon-γ alone could induce these β-cell dysfunctions. Together, our data demonstrate that overexpression of cFLIP protects mouse β-cells against TNF-α–induced caspase-8 activation and apoptosis and is correlated with enhanced NF-κB transcriptional activity, suggesting that cFLIP may have an impact on the outcome of death receptor–triggered responses by directing the intracellular signals from β-cell death to β-cell survival.
The Composition of Dietary Fat Directly Influences Glucose-Stimulated Insulin Secretion in Rats
Authors: Dobbins Robert L. , Szczepaniak Lidia S. , Myhill Jeff , Tamura Yoshifumi , Uchino Hiroshi , Giacca Adria , McGarry J. Denis
Published: March 2007

Acute elevations of plasma free fatty acid (FFA) levels augment glucose-stimulated insulin secretion (GSIS). Prolonged elevations of FFA levels reportedly impair GSIS, but no one has previously compared GSIS after prolonged exposure to saturated or unsaturated fat. Rats received a low-fat diet (Low-Fat) or one enriched with either saturated (Lard) or unsaturated fat (Soy) for 4 weeks. Insulin responses during hyperglycemic clamps were augmented by saturated but not unsaturated fat (580 ± 25, 325 ± 30, and 380 ± 50 pmol · l−1 · min−1 in Lard, Soy, and Low-Fat groups, respectively). Despite hyperinsulinemia, the amount of glucose infused was lower in the Lard compared with the Low-Fat group. Separate studies measured GSIS from the perfused pancreas. Without fatty acids in the perfusate, insulin output in the Lard group (135 ± 22 ng/30 min) matched that of Low-Fat rats (115 ± 13 ng/30 min), but exceeded that of Soy rats (80 ± 7 ng/30 min). When FFAs in the perfusate mimicked the quantity and composition of plasma FFAs in intact animals, in vivo insulin secretory patterns were restored. Because the GSIS of rats consuming Lard diets consistently exceeded that of the Soy group, we also assessed responses after 48-h infusions of lard or soy oil. Again, lard oil exhibited greater insulinotropic potency. These data indicate that prolonged exposure to saturated fat enhances GSIS (but this does not entirely compensate for insulin resistance), whereas unsaturated fat, given in the diet or by infusion, impairs GSIS. Inferences regarding the impact of fatty acids on GSIS that are based on models using unsaturated fat may not reflect the effects of saturated fat.
Gene Expression Profile in Skeletal Muscle of Type 2 Diabetes and the Effect of Insulin Treatment
Authors: Sreekumar Raghavakaimal , Halvatsiotis Panagiotis , Schimke Jill Coenen , Nair K. Sreekumaran
Published: March 2007

Type 2 diabetes is characterized by muscle insulin resistance. Nondiabetic first-degree relatives of type 2 diabetic patients have also been reported to have insulin resistance. A polygenic basis for pathogenesis of type 2 diabetes has been proposed. A gene expression profile was evaluated in the skeletal muscle of patients with type 2 diabetes while not on treatment for 2 weeks and after 10 days of intensive insulin treatment. Comparison of gene expression pattern with age-, sex-, and BMI-matched people with no family history of diabetes was performed using a microarray technique (Hu6800 arrays; Affymetrix, Santa Clara, CA). Only those gene transcripts showing ≥1.9-fold changes and an average difference in fluorescence intensity of ≥1,000 in all subjects are reported. Insulin sensitivity (SI) was measured using an intravenous glucose tolerance test. Of 6,451 genes surveyed, transcriptional patterns of 85 genes showed alterations in the diabetic patients after withdrawal of treatment, when compared with patterns in the nondiabetic control subjects. Insulin treatment reduced the difference in patterns between diabetic and nondiabetic control subjects (improved) in all but 11 gene transcripts, which included genes involved in structural and contractile functions, growth and tissue development, stress response, and energy metabolism. These improved transcripts included genes involved in insulin signaling, transcription factors, and mitochondrial maintenance. However, insulin treatment altered the transcription of 29 additional genes involved in signal transduction; structural and contractile functions; growth and tissue development; and protein, fat, and energy metabolism. Type 2 diabetic patients had elevated circulating insulin during the insulin-treated phase, although their blood glucose levels (98.8 ± 6.4 vs. 90.0 ± 2.9 mg/dl for diabetic vs. control) were similar to those of the control subjects. In contrast, after withdrawal of treatment, the diabetic patients had reduced SI and elevated blood glucose (224.0 ± 26.2 mg/dl), although their insulin levels were similar to those of the nondiabetic control subjects. This study identified several candidate genes for muscle insulin resistance, complications associated with poor glycemic control, and effects of insulin treatment in people with type 2 diabetes.
Interleukin-10 Protects Nitric Oxide–Dependent Relaxation During Diabetes
Authors: Gunnett Carol A. , Heistad Donald D. , Faraci Frank M.
Published: March 2007

Interleukin (IL)-10, an anti-inflammatory cytokine, preserves endothelial function during acute inflammation. We tested the hypotheses that IL-10 plays a protective role in blood vessels during diabetes by suppressing impairment of endothelium-dependent relaxation and that protection by IL-10 is mediated by effects on superoxide (O2−). Streptozotocin (150 mg/kg i.p.) or citrate buffer was injected into IL-10–deficient (IL-10−/−) mice and wild-type controls (IL-10+/+). In IL-10+/+ and IL-10−/− mice, blood glucose levels were ∼120 mg/dl after citrate administration and ∼400 mg/dl after streptozotocin administration. Vasorelaxation was examined in arteries in vitro 12–16 weeks later. Maximum relaxation to acetylcholine (30 μmol/l) was 88 ± 3% (means ± SE) in nondiabetic mice and 84 ± 3% in diabetic IL-10+ /+ mice (P > 0.05). Thus, at this time point, diabetes did not impair endothelium-dependent relaxation in vessels in wild-type mice. In contrast, maximum relaxation in vessels from diabetic IL-10−/− mice was significantly decreased (74 ± 5%) compared with nondiabetic IL-10−/− mice (93 ± 2%, P < 0.05). Superoxide dismutase with polyethylene glycol (PEG-SOD) restored impaired responses to acetylcholine to levels seen in controls. Responses to acetylcholine also were improved by allopurinol (an inhibitor of xanthine oxidase) in vessels from diabetic IL-10− /− mice. Thus, diabetes produces greater impairment of relaxation to acetylcholine in IL-10−/− mice than in IL-10+/ + mice. These findings provide direct evidence that IL-10 impedes mechanisms of endothelial dysfunction during diabetes. Restoration of vasorelaxation with PEG-SOD or allopurinol suggests that the mechanism(s) by which IL-10 preserves endothelium-dependent vasorelaxation involves O2−, perhaps by reducing production of O2− by xanthine oxidase.
Thrombomodulin Deficiency in Human Diabetic Nerve Microvasculature
Authors: Hafer-Macko Charlene E. , Ivey Frederick M. , Gyure Kymberly A. , Sorkin John D. , Macko Richard F.
Published: March 2007

Human diabetic neuropathy is multifactorial in etiology, with ischemia as a final common pathology. Although impaired vascular endothelial cell function in diabetic microvascular injury is established, the role of thrombomodulin (TM)-dependent protein C antithrombotic mechanism in the pathogenesis of neuropathy is unclear. This neuropathologic case-control study investigated whether vascular endothelial TM expression is deficient in peripheral nerve microvessels in diabetic neuropathy. Sural nerve biopsies from 7 patients with diabetic neuropathy and 10 with axonal neuropathy without vasculopathy were immunostained with anti-TM and anti–von Willebrand factor (vWF; an endothelial cell marker) antibodies. The proportion of TM-positive microvessels was expressed relative to total vWF-staining vessels, according to vessel caliber and regional distribution within the nerve. In diabetic nerves compared with reference controls, the proportion of TM-positive endoneurial microvessels was 15-fold lower (0.02 vs. 0.30 in diabetic nerves vs. controls, P < 0.004), and the proportion of small-caliber epineurial microvessels was 10-fold lower (0.04 vs. 0.43, P < 0.001). No TM expression was detected at the perineurium in diabetic or control nerves. We demonstrate a substantial reduction of vascular endothelial TM expression throughout human diabetic neuropathy. These findings suggest that an impaired native TM-dependent protein C antithrombotic mechanism may contribute to microvascular ischemia in the pathogenesis of diabetic neuropathy.
Insertion/Deletion Polymorphism of the ACE Gene Is Associated With Type 2 Diabetes
Authors: Feng Yan , Niu Tianhua , Xu Xin , Chen Changzhong , Li Qiongfang , Qian Rongli , Wang Guoying , Xu Xiping
Published: March 2007

In an attempt to examine the role of an ACE gene insertion/deletion (I/D) polymorphism in type 2 diabetes, we conducted a case-control association study among 132 couple-pairs from northern China. The genotype frequencies for II, ID, and DD were 39.8, 39.8, and 20.3%, respectively, in the case group and 44.8, 44.8, and 10.4% in the control group. The DD frequency was significantly higher in the case group than in the control group (χ21 = 4.77, P = 0.03), suggesting that the DD genotype is associated with an increased susceptibility to type 2 diabetes in our study population.
An X-Chromosome Scan Reveals a Locus for Fat Distribution in Chromosome Region Xp21–22
Authors: Price R. Arlen , Li Wei-Dong , Kilker Robin
Published: March 2007

Several groups have completed autosomal genome scans for human obesity, but only two have examined the X chromosome. A French group reported linkage of BMI to Xp and Xq markers, and a Finnish group reported linkage of BMI to Xq. We scanned the X chromosome in two cohorts, 190 European-American families (940 members) and 43 African-American families (208 members). We examined five correlated obesity phenotypes, BMI, body fat percentage, hip and waist circumferences, and plasma leptin concentration. We also examined leptin resistance (leptin/BMI) and fat patterning (waist-to-hip ratio [WHR]). Variables were adjusted for age within generation, race, and sex. We genotyped 20 markers with average spacing of 10 cM and no interval >22 cM and conducted nonparametric analyses. Suggestive linkage was found for WHR only. Linkage was supported in both family sets, and support was especially strong for females. Z scores for analyses of female phenotypes were 2.69, 1.73, and 2.37 (P = 0.0036, 0.0418, and 0.0089) for African-Americans, European-Americans, and the combined sample, respectively. The peaks were 51–73 cM from the p terminus, 14–34 cM distal of the French report in Xp22. Our results suggest that a quantitative trait locus influencing fat distribution in women may lie in chromosome region Xp21–22; however, the linked interval is large and differs substantially from that of the French and Finnish groups. Given the positive but divergent results, it would be worthwhile for others to examine the X chromosome.
Lipid-Induced Insulin Resistance in Human Muscle Is Associated With Changes in Diacylglycerol, Protein Kinase C, and IκB-α
Authors: Itani Samar I. , Ruderman Neil B. , Schmieder Frank , Boden Guenther
Published: March 2007

The possibility that lipid-induced insulin resistance in human muscle is related to alterations in diacylglycerol (DAG)/protein kinase C (PKC) signaling was investigated in normal volunteers during euglycemic-hyperinsulinemic clamping in which plasma free fatty acid (FFA) levels were increased by a lipid/heparin infusion. In keeping with previous reports, rates of insulin-stimulated glucose disappearance (GRd) were normal after 2 h but were reduced by 43% (from 52.7 ± 8.2 to 30.0 ± 5.3 μmol · kg–1 · min–1, P < 0.05) after 6 h of lipid infusion. No changes in PKC activity or DAG mass were seen in muscle biopsy samples after 2 h of lipid infusion; however, at ∼6 h, PKC activity and DAG mass were increased approximately fourfold, as were the abundance of membrane-associated PKC-βII and -δ. A threefold increase in membrane-associated PKC-βII was also observed at ∼2 h but was not statistically significant (P = 0.058). Ceramide mass was not changed at either time point. To evaluate whether the fatty acid–induced insulin activation of PKC was associated with a change in the IkB kinase (IKK)/nuclear factor (NF)-κB pathway, we determined the abundance in muscle of IκB-α, an inhibitor of NF-κB that is degraded after its phosphorylation by IKK. In parallel with the changes in DAG/PKC, no change in IκB-α mass was observed after 2 h of lipid infusion, but at ∼6 h, IκB-α was diminished by 70%. In summary, the results indicated that the insulin resistance observed in human muscle when plasma FFA levels were elevated during euglycemic-hyperinsulinemic clamping was associated with increases in DAG mass and membrane-associated PKC-βII and -δ and a decrease in IκB-α. Whether acute FFA-induced insulin resistance in human skeletal muscle is caused by the activation of these specific PKC isoforms and the IKK-β/IκB/NFκB pathway remains to be established.
The Gene INPPL1, Encoding the Lipid Phosphatase SHIP2, Is a Candidate for Type 2 Diabetes In Rat and Man
Authors: Marion Evelyne , Kaisaki Pamela Jane , Pouillon Valérie , Gueydan Cyril , Levy Jonathan C. , Bodson André , Krzentowski Georges , Daubresse Jean-Claude , Mockel Jean , Behrends Jens , Servais Geneviève , Szpirer Claude , Kruys Véronique , Gauguier Dominique , Schurmans Stéphane
Published: March 2007

Genetic susceptibility to type 2 diabetes involves many genes, most of which are still unknown. The lipid phosphatase SHIP2 is a potent negative regulator of insulin signaling and sensitivity in vivo and is thus a good candidate gene. Here we report the presence of SHIP2 gene mutations associated with type 2 diabetes in rats and humans. The R1142C mutation specifically identified in Goto-Kakizaki (GK) and spontaneously hypertensive rat strains disrupts a potential class II ligand for Src homology (SH)-3 domain and slightly impairs insulin signaling in cell culture. In humans, a deletion identified in the SHIP2 3′ untranslated region (UTR) of type 2 diabetic subjects includes a motif implicated in the control of protein synthesis. In cell culture, the deletion results in reporter messenger RNA and protein overexpression. Finally, genotyping of a cohort of type 2 diabetic and control subjects showed a significant association between the deletion and type 2 diabetes. Altogether, our results show that mutations in the SHIP2 gene contribute to the genetic susceptibility to type 2 diabetes in rats and humans.
Increased Lipolysis and Decreased Leptin Production by Human Omental as Compared With Subcutaneous Preadipocytes
Authors: van Harmelen Vanessa , Dicker Andrea , Rydén Mikael , Hauner Hans , Lönnqvist Fredrik , Näslund Erik , Arner Peter
Published: March 2007

Site differences in adipose tissue function may have implications for insulin-resistant conditions. In mature adipose tissue, subcutaneous adipocytes have higher leptin secretion, similar tumor necrosis factor (TNF)-α secretion, and lower catecholamine-stimulated lipolysis as compared with omental adipocytes. In this study, lipolysis and leptin and TNF-α secretion were compared between human omental and subcutaneous preadipocytes. After 16 days of incubation in a minimal differentiation medium, leptin mRNA and secretion were found to be two to eight times higher in subcutaneous than omental preadipocytes (P < 0.05). On the other hand, norepinephrine-induced lipolysis was about two times higher in the omental than in the subcutaneous preadipocytes, whereas basal lipolysis did not differ between the two regions. TNF-α secretion was marginally but significantly higher in the omental than in the subcutaneous preadipocytes. Preadipocyte differentiation was equal in both regions and was augmented to the same extent by different thiazolidinediones (rosiglitazone, pioglitazone, or troglitazone) in the two depots. In the presence of rosiglitazone, leptin secretion remained about three times higher and norepinephrine-induced lipolysis about six times lower in subcutaneous as compared with omental preadipocytes (P < 0.05), whereas TNF-α secretion and basal lipolysis were similar in preadipocytes from the two regions. These findings remained unaltered even if rosiglitazone was removed from the medium. However, leptin mRNA showed no regional differences in rosiglitazone-treated cells. Thus, regional differences in adipocyte leptin secretion as well as in norepinephrine-induced lipolysis are marked and present during different stages of preadipocyte differentiation and seem to be determined by intrinsic (i.e., primary) factors.
Isomer-Dependent Metabolic Effects of Conjugated Linoleic Acid
Authors: Roche Helen M. , Noone Enda , Sewter Ciaran , Mc Bennett Siobhan , Savage David , Gibney Michael J. , O’Rahilly Stephen , Vidal-Puig Antonio J.
Published: March 2007

Conjugated linoleic acid (CLA) is a heterogeneous group of positional and geometric isomers of linoleic acid. This study demonstrates the divergent effects of the cis-9 trans-11 (c9,t11-CLA) and trans-10 cis-12 (t10,c12-CLA) isomers of CLA on lipid metabolism and nutrient regulation of gene expression in ob/ob mice. The c9, t11-CLA diet decreased serum triacylglycerol (P = 0.01) and nonesterified fatty acid (NEFA) (P = 0.05) concentrations, and this was associated with reduced hepatic sterol regulatory element-binding protein-1c (SREBP-1c; P = 0.0045) mRNA expression, coupled with reduced levels of both the membrane-bound precursor and the nuclear forms of the SREBP-1 protein. C9,t11-CLA significantly reduced hepatic LXRα (P = 0.019) mRNA expression, a novel regulator of SREBP-1c. In contrast, c9,t11-CLA increased adipose tissue SREBP-1c mRNA expression (P = 0.0162) proportionally to the degree of reduction of tumor necrosis factor α (TNF-α) mRNA (P = 0.012). Recombinant TNF-α almost completely abolished adipose tissue SREBP-1c mRNA expression in vivo. The t10,c12-CLA diet promoted insulin resistance and increased serum glucose (P = 0.025) and insulin (P = 0.01) concentrations. T10, c12-CLA induced profound weight loss (P = 0.0001) and increased brown and white adipose tissue UCP-2 (P = 0.001) and skeletal muscle UCP-3 (P = 0.008) mRNA expression. This study highlights the contrasting molecular and metabolic effect of two isomers of the same fatty acids. The ameliorative effect of c9,t11-CLA on lipid metabolism may be ascribed to reduced synthesis and cleavage of hepatic SREBP-1, which in turn may be regulated by hepatic LXRα expression.
Metformin Increases AMP-Activated Protein Kinase Activity in Skeletal Muscle of Subjects With Type 2 Diabetes
Authors: Musi Nicolas , Hirshman Michael F. , Nygren Jonas , Svanfeldt Monika , Bavenholm Peter , Rooyackers Olav , Zhou Gaochao , Williamson Joanne M. , Ljunqvist Olle , Efendic Suad , Moller David E. , Thorell Anders , Goodyear Laurie J.
Published: March 2007

Metformin is an effective hypoglycemic drug that lowers blood glucose concentrations by decreasing hepatic glucose production and increasing glucose disposal in skeletal muscle; however, the molecular site of metformin action is not well understood. AMP-activated protein kinase (AMPK) activity increases in response to depletion of cellular energy stores, and this enzyme has been implicated in the stimulation of glucose uptake into skeletal muscle and the inhibition of liver gluconeogenesis. We recently reported that AMPK is activated by metformin in cultured rat hepatocytes, mediating the inhibitory effects of the drug on hepatic glucose production. In the present study, we evaluated whether therapeutic doses of metformin increase AMPK activity in vivo in subjects with type 2 diabetes. Metformin treatment for 10 weeks significantly increased AMPK α2 activity in the skeletal muscle, and this was associated with increased phosphorylation of AMPK on Thr172 and decreased acetyl-CoA carboxylase-2 activity. The increase in AMPK α2 activity was likely due to a change in muscle energy status because ATP and phosphocreatine concentrations were lower after metformin treatment. Metformin-induced increases in AMPK activity were associated with higher rates of glucose disposal and muscle glycogen concentrations. These findings suggest that the metabolic effects of metformin in subjects with type 2 diabetes may be mediated by the activation of AMPK α2.
Regulation of Circulating Soluble Leptin Receptor Levels By Gender, Adiposity, Sex Steroids, and Leptin
Authors: Chan Jean L. , Blüher Susann , Yiannakouris Nikos , Suchard Marc A. , Kratzsch Jurgen , Mantzoros Christos S.
Published: March 2007

Leptin is an adipocyte-secreted hormone important in energy homeostasis and diverse physiological processes. A circulating soluble form of the leptin receptor [soluble leptin receptor (sOB-R)] is the main leptin-binding protein and determinant of free leptin index (FLI), the presumed biologically active form of leptin. We performed observational and interventional studies to elucidate the regulation of sOB-R and FLI in humans. In a cross-sectional study (n = 118), leptin, gender, and adiposity were significant determinants of sOB-R. By multivariate analysis, estradiol (E2) and testosterone predict sOB-R, whereas insulin predicts leptin and FLI. In a frequent-sampling study (n = 6), sOB-R followed a significant circadian rhythm inverse to that of leptin, suggesting that leptin’s biological activity may have an even more pronounced diurnal variation than originally thought. A 72-h fast in eight men decreased leptin levels by 80% and increased lymphocyte expression of leptin receptor mRNA and serum sOB-R levels by 100%. Physiological and pharmacological doses of recombinant-methionyl human leptin (rhLeptin) administered to fasted men prevented the fasting-induced increase of sOB-R levels, and pharmacological doses resulted in a decrease in sOB-R levels. These studies provide evidence that sOB-R is regulated by gender, adiposity, hormones, and rhLeptin administration. This may have important implications for the biological activity of leptin in disease states associated with abnormal leptin levels (e.g., obesity and anorexia nervosa).
Adrenalectomy Improves Diabetes in A-ZIP/F-1 Lipoatrophic Mice by Increasing Both Liver and Muscle Insulin Sensitivity
Authors: Haluzik Martin , Dietz Kelly R. , Kim Jason K. , Marcus-Samuels Bernice , Shulman Gerald I. , Gavrilova Oksana , Reitman Marc L.
Published: March 2007

The virtually fatless A-ZIP/F-1 mouse is profoundly insulin resistant, diabetic, and a good model for humans with severe generalized lipoatrophy. Like a number of other mouse models of diabetes, the A-ZIP/F-1 mouse has elevated serum corticosterone levels. Leptin infusion lowers the corticosterone levels, suggesting that leptin deficiency contributes to the hypercorticosteronemic state. To test the hypothesis that the increased glucocorticoids contribute to the diabetes and insulin resistance, we examined the effect of adrenalectomy on A-ZIP/F-1 mice. Adrenalectomy significantly decreased the blood glucose, serum insulin, and glycated hemoglobin levels. Hyperinsulinemic-euglycemic clamps were performed to characterize the changes in whole-body and tissue insulin sensitivity. The adrenalectomized A-ZIP/F-1 mice displayed a marked improvement in insulin-induced suppression of endogenous glucose production, indicating increased hepatic insulin sensitivity. Adrenalectomy also increased muscle glucose uptake and glycogen synthesis. These results suggest that the chronically increased serum corticosterone levels contribute to the diabetes of the A-ZIP/F-1 mice and that removal of the glucocorticoid excess improves the insulin sensitivity in both muscle and liver.
Immunological Characterization and Therapeutic Activity of an Altered-Peptide Ligand, NBI-6024, Based on the Immunodominant Type 1 Diabetes Autoantigen Insulin B-Chain (9–23) Peptide
Authors: Alleva David G. , Gaur Amitabh , Jin Liping , Wegmann Dale , Gottlieb Peter A. , Pahuja Anil , Johnson Eric B. , Motheral Theresa , Putnam Amy , Crowe Paul D. , Ling Nicholas , Boehme Stefen A. , Conlon Paul J.
Published: March 2007

The nonobese diabetic (NOD) mouse is a good model for human type 1 diabetes, which is characterized by autoreactive T-cell-mediated destruction of insulin-producing islet β-cells of the pancreas. The 9–23 amino acid region of the insulin B-chain [B(9–23)] is an immunodominant T-cell target antigen in the NOD mouse that plays a critical role in the disease process. By testing a series of B(9–23) peptide analogs with single or double alanine substitutions, we identified a set of altered peptide ligands (APLs) capable of inhibiting B(9–23)-induced proliferative responses of NOD pathogenic T-cell clones. These APLs were unable to induce proliferation of these clones. However, vaccinations with the APLs induced strong cellular responses, as measured by in vitro lymphocyte proliferation and Th2 cytokine production (i.e., interleukin [IL]-4 and IL-10, but not γ-interferon [IFN-γ]). These responses were cross-reactive with the native antigen, B(9–23), suggesting that the APL-induced Th2 responses may provide protection by controlling endogenous B(9–23)-specific Th1 (i.e., IFN-γ-producing) pathogenic responses. One of these APLs that contained alanine substitutions at residues 16 and 19 (16Y→A, 19C→A; NBI-6024) was further characterized for its therapeutic activity because it consistently induced T-cell responses (e.g., T-cell lines and clones) that were of the Th2 type and that were cross-reactive with B(9–23). Subcutaneous injections of NBI-6024 to NOD mice administered either before or after the onset of disease substantially delayed the onset and reduced the incidence of diabetes. This study is the first to report therapeutic activity of an APL derived from an islet β-cell-specific antigen in type 1 diabetes.
An Interval Tightly Linked to but Distinct From the H2 Complex Controls Both Overt Diabetes (Idd16) and Chronic Experimental Autoimmune Thyroiditis (Ceat1) in Nonobese Diabetic Mice
Authors: Boulard Olivier , Damotte Diane , Deruytter Nathalie , Fluteau Guy , Carnaud Claude , Garchon Henri-Jean
Published: March 2007

The major histocompatibility complex (MHC) has long been associated with predisposition to several autoimmune diseases, including type 1 diabetes and autoimmune thyroiditis. In type 1 diabetes, a primary role has been assigned to class II genes, both in humans and in the nonobese diabetic (NOD) mouse model. However, an involvement of other tightly linked genes is strongly suspected. Here, through two independent sets of experiments, we provide solid evidence for the existence of at least one such gene. First, using a new recombinant congenic NOD strain, R114, we definitively individualized the Idd16 locus from the MHC in a 6-cM interval proximal to H2-K. It affords almost complete protection against diabetes and is associated with delayed insulitis. Second, by genome scan, we mapped non-H2 genes associated with the highly penetrant form of chronic experimental autoimmune thyroiditis (EAT) that is elicited in NOD and NOD.H2k mice by immunization with thyroglobulin. We identified one major dominant locus, Ceat1, on chromosome 17, overlapping with Idd16. Most importantly, R114 recombinant congenic mice challenged with thyroglobulin did not develop chronic EAT. This new major region defined by both Idd16 and Ceat1 might thus concur to the unique strength of the MHC in autoimmune susceptibility of NOD mice.
Glycemic Control Determines Hepatic and Peripheral Glucose Effectiveness in Type 2 Diabetic Subjects
Authors: Hawkins Meredith , Gabriely Ilan , Wozniak Robert , Reddy Kalpana , Rossetti Luciano , Shamoon Harry
Published: March 2007

Glucose effectiveness is impaired in type 2 diabetes. We hypothesize that chronic hyperglycemia and hyperlipidemia contribute importantly to this defect. To test this hypothesis, we compared the effect of acute hyperglycemia on glucose turnover in type 2 diabetic subjects in good control (GC) (n = 14, age 51.7 ± 3.7 years, BMI 28.4 ± 1.0 kg/ m2, HbA1c 5.9 ± 0.2%) and poor control (PC) (n = 10, age 50.0 ± 2.5 years, BMI 27.9 ± 1.5 kg/m2, HbA1c 9.9 ± 0.6%) with age- and weight-matched nondiabetic subjects (ND) (n = 11, age 47.0 ± 4.4 years, BMI 28.5 ± 1.0 kg/m2, HbA1c 5.1 ± 0.2%). Fixed hormonal conditions were attained by infusing somatostatin for 6 h with replacement of basal insulin, glucagon, and growth hormone. Glucose fluxes ([3-3H]glucose) were compared during euglycemic (5 mmol/l, t = 180–240 min) and hyperglycemic (Hy) (10 mmol/l, t = 300–360 min, variable glucose infusion) clamp intervals. Acute hyperglycemia suppressed hepatic glucose production (GP) by 43% and increased peripheral glucose uptake (GU) by 86% in the ND subjects. Conversely, GP failed to suppress (−7%) and GU was suboptimally increased (+34%) in response to Hy in the PC group. However, optimal glycemic control was associated with normal glucose effectiveness in GC subjects (GP −38%, GU +72%; P > 0.05 for GC vs. ND). To determine whether short-term correction of hyperglycemia and/or hyperlipidemia is sufficient to reverse the impairment in glucose effectiveness, five PC subjects were restudied after 72 h of normoglycemia (∼100 mg/dl; variable insulin infusions). These subjects regained normal effectiveness of glucose to suppress GP and stimulate GU and in response to Hy (GP −47%, GU + 71%; P > 0.05 vs. baseline studies). Thus, chronic hyperglycemia and/or hyperlipidemia contribute to impaired effectiveness of glucose in regulating glucose fluxes in type 2 diabetes and hence to worsening of the overall metabolic condition. Short-term normalization of plasma glucose might break the vicious cycle of impaired glucose effectiveness in type 2 diabetes.
TNF-α Is a Predictor of Insulin Resistance in Human Pregnancy
Authors: Kirwan John P. , Hauguel-De Mouzon Sylvie , Lepercq Jacques , Challier Jean-Claude , Huston-Presley Larraine , Friedman Jacob E. , Kalhan Satish C. , Catalano Patrick M.
Published: March 2007

Historically, insulin resistance during pregnancy has been ascribed to increased production of placental hormones and cortisol. The purpose of this study was to test this hypothesis by correlating the longitudinal changes in insulin sensitivity during pregnancy with changes in placental hormones, cortisol, leptin, and tumor necrosis factor (TNF)-α. Insulin resistance was assessed in 15 women (5 with gestational diabetes mellitus [GDM] and 10 with normal glucose tolerance) using the euglycemic-hyperinsulinemic clamp procedure, before pregnancy (pregravid) and during early (12–14 weeks) and late (34–36 weeks) gestation. Body composition, plasma TNF-α, leptin, cortisol, and reproductive hormones (human chorionic gonadotropin, estradiol, progesterone, human placental lactogen, and prolactin) were measured in conjunction with the clamps. Placental TNF-α was measured in vitro using dually perfused human placental cotyledon from five additional subjects. Compared with pregravid, insulin resistance was evident during late pregnancy in all women (12.4 ± 1.2 vs. 8.1 ± 0.8 10−2 mg · kg−1 fat-free mass · min−1 · μU−1 · ml−1). TNF-α, leptin, cortisol, all reproductive hormones, and fat mass were increased in late pregnancy (P < 0.001). In vitro, most of the placental TNF-α (94%) was released into the maternal circulation; 6% was released to the fetal side. During late pregnancy, TNF-α was inversely correlated with insulin sensitivity (r = −0.69, P < 0.006). Furthermore, among all of the hormonal changes measured in this study, the change in TNF-α from pregravid to late pregnancy was the only significant predictor of the change in insulin sensitivity (r = −0.60, P < 0.02). The placental reproductive hormones and cortisol did not correlate with insulin sensitivity in late pregnancy. Multivariate stepwise regression analysis revealed that TNF-α was the most significant independent predictor of insulin sensitivity (r = −0.67, P < 0.0001), even after adjustment for fat mass by covariance (r = 0.46, P < 0.01). These observations challenge the view that the classical reproductive hormones are the primary mediators of change in insulin sensitivity during gestation and provide the basis for including TNF-α in a new paradigm to explain insulin resistance in pregnancy.
Alterations in the Activity of Placental Amino Acid Transporters in Pregnancies Complicated by Diabetes
Authors: Jansson Thomas , Ekstrand Ylva , Björn Caroline , Wennergren Margareta , Powell Theresa L.
Published: March 2007

Alterations in placental transport may contribute to accelerated fetal growth in pregnancies complicated by diabetes. We studied the activity of the syncytiotrophoblast amino acid transporter system A and the transport of the essential amino acids leucine, lysine, and taurine. Syncytiotrophoblast microvillous plasma membranes (MVMs) and basal plasma membranes (BMs) were isolated from placentas obtained from normal pregnancies and pregnancies complicated by gestational diabetes mellitus (GDM) and type 1 diabetes, with and without large-for-gestational-age (LGA) fetuses. Amino acid transport was assessed using radio-labeled substrates and rapid filtration techniques. System A activity in MVM was increased (65–80%, P < 0.05) in all groups with diabetes independent of fetal overgrowth. However, MVM system A activity was unaffected in placentas of normal pregnancies with LGA fetuses. MVM leucine transport was increased in the GDM/LGA group. In BMs, amino acid transport was unaffected by diabetes. In conclusion, diabetes in pregnancy is associated with an increased system A activity in MVM, and MVM leucine transport is increased in the GDM/LGA group. We suggest that these changes result in an increased uptake of neutral amino acids across MVM, which may be used in placental metabolism or be delivered to the fetus. The increased MVM leucine uptake in the GDM/LGA group may contribute to accelerated fetal growth in these patients.
Vitamin D Receptor (VDR) mRNA and VDR Protein Levels in Relation to Vitamin D Status, Insulin Secretory Capacity, and VDR Genotype in Bangladeshi Asians
Authors: Ogunkolade Babatunji-William , Boucher Barbara J. , Prahl Jean M. , Bustin Stephen A. , Burrin Jacky M. , Noonan Kate , North Bernard V. , Mannan Nassima , McDermott Michael F. , DeLuca Hector F. , Hitman Graham A.
Published: March 2007

Associations have been reported between vitamin D receptor (VDR) gene polymorphisms, type 1 diabetes, insulin secretion, and the insulin resistance syndrome. As VDR polymorphisms have no known functional significance, these findings may implicate a variant of the VDR gene or a locus in linkage disequilibrium with the VDR. We have examined VDR mRNA and VDR protein levels in relation to VDR polymorphisms (41 Bangladeshi subjects) and analyzed insulin secretory capacity (143 Bangladeshi subjects), allowing for other known determinants. Peripheral blood mononuclear cells (PBMCs) from subjects who had been genotyped for BsmI, ApaI, TaqI, and FokI VDR restriction fragment length polymorphisms were used for both total VDR mRNA quantitation (using TaqMan) and measurement of VDR protein levels (using a specific micro-immunoassay). Stepwise multiple regression analyses were used (to P < 0.05) to analyze the data. For the insulin secretion index, the best-fit model (n = 143, P < 0.0001) gave age (P = 0.002), TaqI (P < 0.0001), and BMI (P = 0.001) as independent determinants; with the inclusion of VDR mRNA and VDR protein levels, VDR mRNA was the sole independent determinant (n = 41, P = 0.024). However, the best-fit model for VDR mRNA (P = 0.004) gave FokI (P = 0.044) and TaqI (P = 0.04) genotypes and insulin secretory capacity (P = 0.042) as independent determinants. For VDR protein levels, the best-fit model (P = 0.006) gave TaqI genotype (P = 0.005) and circulating 1,25-dihydroxyvitamin-D levels (P = 0.03) as independent determinants. In conclusion, these studies confirm an association between VDR polymorphisms and insulin secretory capacity and demonstrate the VDR genotype to be a significant determinant of VDR mRNA and VDR protein levels in PBMCs, providing functional support to previously described genetic associations with the VDR gene. Furthermore, VDR expression has been shown to be a determinant of insulin secretory capacity.
A Haplotype at the Adiponectin Locus Is Associated With Obesity and Other Features of the Insulin Resistance Syndrome
Authors: Menzaghi Claudia , Ercolino Tonino , Di Paola Rosa , Berg Anders H. , Warram James H. , Scherer Philipp E. , Trischitta Vincenzo , Doria Alessandro
Published: March 2007

Adiponectin is a protein secreted by adipocytes that modulates insulin action. To assess whether variants of this gene contribute to the prevalence of insulin resistance in Caucasians, we genotyped 413 nondiabetic individuals for two single nucleotide polymorphisms (SNPs) at this locus. The two SNPs (45T→G and 276G→T) were chosen because of their association with type 2 diabetes in Japanese. Whereas each polymorphism was significantly associated with some correlate of insulin resistance, the haplotype defined by the two together was strongly associated with many components of the insulin resistance syndrome. Homozygotes for the risk haplotype had higher body weight (P = 0.03), waist circumference (P = 0.004), systolic (P = 0.01) and diastolic (P = 0.003) blood pressure, fasting glucose (P = 0.02) and insulin (P = 0.005) levels, homeostasis model assessment (HOMA) for insulin resistance (P = 0.003), and total to HDL cholesterol ratio (P = 0.01). Homozygotes also had significantly lower plasma levels of adiponectin (P = 0.03), independent of sex, age, and body weight. In an independent study group of 614 Caucasians, including 310 with type 2 diabetes, the risk haplotype was confirmed to be associated with increased body weight (P = 0.03) but not with type 2 diabetes per se. We conclude that variability at the adiponectin locus is associated with obesity and other features of the insulin resistance syndrome, but given the nature of the two SNPs, the risk haplotype is most probably a marker in linkage disequilibrium with an as yet unidentified polymorphism that affects plasma adiponectin levels and insulin sensitivity.
A Putative Functional Polymorphism in the IGF-I Gene
Authors: Frayling Timothy M. , Hattersley Andrew T. , McCarthy Anne , Holly Jeff , Mitchell Simon M.S. , Gloyn Anna L. , Owen Katharine , Davies David , Smith George Davey , Ben-Shlomo Yoav
Published: March 2007

IGF-I has a critical role in growth and metabolism. A microsatellite polymorphism 1 kb upstream to the IGF-I gene has recently been associated with several adult phenotypes. In a large Dutch cohort, the absence of the commonest allele (Z) was associated with reduced serum IGF-I levels, reduced height, and an increased risk of type 2 diabetes and myocardial infarction. This result has not been replicated, and the role of this polymorphism in these traits in U.K. subjects is not known. We sought further evidence for the involvement of this variant in type 2 diabetes using a case-control study and IGF-I and diabetes-related traits in a population cohort of 640 U.K. individuals aged 25 years. Absence of the common allele was not associated with type 2 diabetes (odds ratio 0.70, 95% CI 0.47–1.04 for X/X versus Z/Z genotype, χ2 test for trend across genotypes, P = 0.018). In the population cohort, the common allele (Z) was associated with decreased IGF-I levels (P = 0.01), contrary to the Dutch study, but not with adult height (P = 0.23), glucose tolerance (P = 0.84), oral glucose tolerance test-derived values of β-cell function (P = 0.90), or insulin resistance (P = 0.66). There was no association with measures of fetal growth, including birth weight (P = 0.17). Our results do not support the previous associations and suggest that the promoter microsatellite is unlikely to be functionally important.
Cross-Talk Between Iron Metabolism and Diabetes
Authors: Fernández-Real José Manuel , López-Bermejo Abel , Ricart Wifredo
Published: March 2007

Emerging scientific evidence has disclosed unsuspected influences between iron metabolism and type 2 diabetes. The relationship is bi-directional—iron affects glucose metabolism, and glucose metabolism impinges on several iron metabolic pathways. Oxidative stress and inflammatory cytokines influence these relationships, amplifying and potentiating the initiated events. The clinical impact of these interactions depends on both the genetic predisposition and the time frame in which this network of closely related signals acts. In recent years, increased iron stores have been found to predict the development of type 2 diabetes while iron depletion was protective. Iron-induced damage might also modulate the development of chronic diabetes complications. Iron depletion has been demonstrated to be beneficial in coronary artery responses, endothelial dysfunction, insulin secretion, insulin action, and metabolic control in type 2 diabetes. Here, we show that iron modulates insulin action in healthy individuals and in patients with type 2 diabetes. The extent of this influence should be tested in large-scale clinical trials, searching for the usefulness and cost-effectiveness of therapeutic measures that decrease iron toxicity. The study of individual susceptibility and of the mechanisms that influence tissue iron deposition and damage are proposed to be valuable in anticipating and treating diabetes complications.