bluuebunny/crossref_metadata_2025 huggingface dataset:
Insulin Sensitivity, Insulin Secretion, and Abdominal Fat
Authors: Wagenknecht Lynne E. , Langefeld Carl D. , Scherzinger Ann L. , Norris Jill M. , Haffner Steven M. , Saad Mohammed F. , Bergman Richard N.
Published: March 2007

The relationship between insulin sensitivity and overall obesity is well established. However, there remains debate as to which of the fat depots, visceral abdominal tissue (VAT) or subcutaneous abdominal tissue (SAT), is of greater importance. Also, the relationship between fat distribution and insulin secretion is largely unknown. We studied SI, acute insulin response (AIR), and disposition index (DI), as obtained by minimal model analysis, in 999 Hispanic and 458 African-American men and women as part of the Insulin Resistance Atherosclerosis Study (IRAS) Family Study. VAT and SAT were measured from computed tomography scans performed at the L4/L5 vertebral region. A mixed-model approach was used to determine the relationship between each of the glucose homeostasis measures (SI, AIR, and DI) versus abdominal fat measures. Mean values were as follows: age, 41 years; SI, 1.98 10−4 · min−1 · μU−1 · ml−1; AIR, 840 pmol · ml−1 · min−1; BMI, 28.5 kg/m2; VAT, 100 cm2; and SAT, 333 cm2. SAT, VAT, and their joint interaction were each inversely and significantly associated with SI, adjusting for age, sex, ethnicity, and BMI. SAT, but not VAT, was positively associated with AIR, except when additionally adjusting for SI, in which case VAT was inversely associated with AIR. VAT and the joint interaction of VAT and SAT were inversely associated with DI. The fat measures explained 27% of the model R2 for SI, 16% for AIR, and 16% for DI. Thus, fat distribution is an important determinant of both insulin resistance and insulin secretion.
Variations in IB1/JIP1 Expression Regulate Susceptibility of β-Cells to Cytokine-Induced Apoptosis Irrespective of C-Jun NH2-Terminal Kinase Signaling
Authors: Ling Zhidong , Van de Casteele Mark , Dong Jing , Heimberg Harry , Haefliger Jacques-Antoine , Waeber Gérard , Schuit Frans , Pipeleers Daniel
Published: March 2007

We previously reported that interleukin-1β (IL-1β) alone does not cause apoptosis of β-cells, whereas when combined with γ-interferon (IFN-γ) and tumor necrosis factor-α (TNF-α), it exerts a distinct apoptotic effect. Studies in β-cell lines indicated that IL-1β reduced expression of islet brain (IB)-1/JNK interacting protein (JIP)-1, a JNK scaffold protein with antiapoptotic action. We examined whether variations in IB1/JIP-1 expression in purified primary β-cells affect their susceptibility to cytokine-induced apoptosis. Exposure to IL-1β for 24 h decreased cellular IB1/JIP-1 content by 66 ± 17%; this IL-1β effect was maintained in the presence of TNF-α + IFN-γ, which did not influence IB1/JIP-1 levels by themselves. Addition of IL-1β to TNF-α + IFN-γ increased apoptosis from 20 ± 2% to 59 ± 5%. A similar increase in TNF-α + IFN-γ-induced apoptosis was produced by adenoviral expression of antisense IB1/JIP-1 and was not further enhanced by addition of IL-1β, indicating that IL-1β-mediated suppression of IB1/JIP-1 in β-cells increases their susceptibility to cytokine-induced apoptosis. However, adenovirally mediated overexpression of IB1/JIP-1 also potentiated TNF-α + IFN-γ-induced apoptosis, suggesting that the antiapoptotic effect of IB1/JIP-1 depends on well-defined cellular levels. We conclude that the IB1/JIP-1 level in β-cells can control their susceptibility to apoptosis independent of JNK signaling.
Low Plasma Ghrelin Is Associated With Insulin Resistance, Hypertension, and the Prevalence of Type 2 Diabetes
Authors: Pöykkö Seppo M. , Kellokoski Eija , Hörkkö Sohvi , Kauma Heikki , Kesäniemi Y. Antero , Ukkola Olavi
Published: March 2007

Experimental studies have suggested that ghrelin plays a role in glucose homeostasis and in the regulation of blood pressure (BP). We therefore assessed the hypothesis that a low ghrelin concentration may be a risk factor for type 2 diabetes and hypertension. We also characterized the effect of the ghrelin Arg51Gln and Leu72Met mutations on ghrelin concentrations in the population-based hypertensive (n = 519) and control (n = 526) cohorts of our OPERA (Oulu Project Elucidating Risk of Atherosclerosis) study. The fasting plasma ghrelin concentrations of 1,040 subjects were analyzed using the radioimmunoassay method. Insulin sensitivity was assessed using the quantitative insulin sensitivity check index (QUICKI). Ghrelin concentrations were negatively associated with fasting insulin (P < 0.001), systolic (P = 0.026) and diastolic BP (P = 0.018), and the prevalence of type 2 diabetes (P = 0.015) and insulin resistance (P < 0.001) in the multivariate models. In the control cohort, low ghrelin was associated with hypertension (BP >140/90 mmHg) (P = 0.031). The subjects with the ghrelin 51Gln allele had lower ghrelin concentrations than the Arg51Arg homozygotes (P = 0.001). We conclude that low ghrelin is independently associated with type 2 diabetes, insulin concentration, insulin resistance, and elevated BP. Therefore, it might have some role in the etiology of type 2 diabetes and the regulation of BP. The ghrelin Arg51Gln mutation is associated with low plasma ghrelin concentrations.
Insulin Affects Vascular Smooth Muscle Cell Phenotype and Migration Via Distinct Signaling Pathways
Authors: Wang Cecilia C. Low , Gurevich Inga , Draznin Boris
Published: March 2007

Insulin maintains vascular smooth muscle cell (VSMC) quiescence yet can also promote VSMC migration. The mechanisms by which insulin exerts these contrasting effects were examined using α-smooth muscle actin (α-SMA) as a marker of VSMC phenotype because α-SMA is highly expressed in quiescent but not migratory VSMC. Insulin alone maintained VSMC quiescence and modestly stimulated VSMC migration. Wortmannin, a phosphatidylinositol 3-kinase (PI3K) inhibitor, decreased insulin-stimulated expression of α-SMA mRNA by 26% and protein by 48% but had no effect on VSMC migration. PD98059, a mitogen-activated protein kinase (MAPK) kinase inhibitor, decreased insulin-induced VSMC migration by 52% but did not affect α-SMA levels. Platelet-derived growth factor (PDGF) promoted dedifferentiation of VSMC, and insulin counteracted this effect. Furthermore, insulin increased α-SMA mRNA and protein levels to 111 and 118%, respectively, after PDGF-induced dedifferentiation, an effect inhibited by wortmannin. In conclusion, insulin’s ability to maintain VSMC quiescence and reverse the dedifferentiating influence of PDGF is mediated via the PI3K pathway, whereas insulin promotes VSMC migration via the MAPK pathway. Thus, with impaired PI 3-kinase signaling and intact MAPK signaling, as seen in insulin resistance, insulin may lose its ability to maintain VSMC quiescence and instead promote VSMC migration.
KIR in Type 1 Diabetes
Authors: van der Slik Arno R. , Koeleman Bobby P.C. , Verduijn Willem , Bruining G. Jan , Roep Bart O. , Giphart Marius J.
Published: March 2007

Killer cell immunoglobulin-like receptors (KIRs) modulate natural killer cell and T-cell function by interacting with HLA class 1 ligands on target cells. Both KIR and HLA are highly polymorphic. We studied the influence of KIR and HLA class 1 genes on the susceptibility to develop type 1 diabetes. The results showed increased numbers of activating KIR genes in patients compared with control subjects (P = 0.049). The combination of the activating KIR2DS2 gene, together with its putative HLA ligand, was present more frequently in patients than in diabetes high-risk HLA-matched control subjects (P = 0.030). Moreover, our results imply that an increase in activating KIR2DS2-HLA ligand pairs combined with a lack of inhibitory KIR-HLA ligand pairs is associated with an additional risk to develop type 1 diabetes in individuals with diabetes high-risk HLA alleles (P = 0.035). We propose that the genetic imbalance between KIR and their HLA class 1 ligands may enhance the activation of T-cells with a low affinity for pancreatic self-antigens, thereby contributing to the pathogenesis of type 1 diabetes.
Recombinant Fabs of Human Monoclonal Antibodies Specific to the Middle Epitope of GAD65 Inhibit Type 1 Diabetes–Specific GAD65Abs
Authors: Padoa Carolyn J. , Banga J. Paul , Madec Anne-Marie , Ziegler Manfred , Schlosser Michael , Ortqvist Eva , Kockum Ingrid , Palmer Jerry , Rolandsson Olov , Binder Katherine A. , Foote Jefferson , Luo Dong , Hampe Christiane S.
Published: March 2007

Autoantibodies to the 65-kDa isoform of GAD (GAD65Abs) are associated with type 1 diabetes development, but the conformational nature of the GAD65Ab epitopes complicates the evaluation of disease risk. Six GAD65-specific recombinant Fabs (rFabs) were cloned from monoclonal antibodies b96.11, DP-C, DP-A, DPD, 144, and 221–442. The binding of GAD65Abs in 61 type 1 diabetic patients to GAD65 was analyzed by competitive radioimmunoassays with the six rFabs to ascertain disease-specific GAD65Ab binding specificities. The median binding was reduced significantly by rFab b96.11 (72%) (P < 0.0001), DP-A (84%) (P < 0.0001), DP-C (84%) (P < 0.0001), 221–442 (79%) (P < 0.0001), and DP-D (80%) (P < 0.0001). The competition pattern in type 1 diabetic patients differed from that in GAD65Ab-positive late autoimmune diabetes in adults (LADA) patients (n = 44), first-degree relatives (n = 38), and healthy individuals (n = 14). Whereas 87 and 72% of the type 1 diabetic sera were competed by rFab b96.11 and DP-C, respectively, only 34 and 26% of LADA patients, 18 and 25% of first-degree relatives, and 7 and 28% of healthy individuals showed competition (P < 0.0001). These findings support the view that type 1 diabetes is associated with disease- and epitope-specific GAD65Abs and supports the notion that the middle epitope is disease associated. These GAD65-specific rFabs should prove useful in predicting type 1 diabetes and in the study of conformational GAD65Ab epitopes.
Discovery of Gene Networks Regulating Cytokine-Induced Dysfunction and Apoptosis in Insulin-Producing INS-1 Cells
Authors: Kutlu Burak , Cardozo Alessandra K. , Darville Martine I. , Kruhøffer Mogens , Magnusson Nils , Ørntoft Torben , Eizirik Décio L.
Published: March 2007

Locally released cytokines contribute to β-cell dysfunction and apoptosis in type 1 diabetes. In vitro exposure of insulin-producing INS-1E cells to the cytokines interleukin (IL)-1β + interferon (IFN)-γ leads to a significant increase in apoptosis. To characterize the genetic networks implicated in β-cell dysfunction and apoptosis and its dependence on nitric oxide (NO) production, we performed a time-course microarray analysis of cytokine-induced genes in insulin-producing INS-1E cells. INS-1E cells were exposed in duplicate to IL-1β + IFN-γ for six different time points (1, 2, 4, 8, 12, and 24 h) with or without the inducible NO synthase (iNOS) blocker NG-monomethyl-l-arginine (NMA). The microarray analysis identified 698 genes as cytokine modified (≥2.5-fold change compared with control) in at least one time point. Based on their temporal pattern of variation, the cytokine-regulated genes were classified into 15 clusters by the k-means method. These genes were further classified into 14 different groups according to their putative function. Changes in the expression of genes related to metabolism, signal transduction, and transcription factors at all time points studied indicate β-cell attempts to adapt to the effects of continuous cytokine exposure. Notably, several apoptosis-related genes were modified at early time points (2–4 h) preceding iNOS expression. On the other hand, 46% of the genes modified by cytokines after 8–24 h were NO dependent, indicating the important role of this radical for the late effects of cytokines. The present results increase by more than twofold the number of known cytokine-modified genes in insulin-producing cells and yield comprehensive information on the role of NO for these modifications in gene expression. These data provide novel and detailed insights into the gene networks activated in β-cells facing a prolonged immune assault.
Contribution of Elevated Free Fatty Acid Levels to the Lack of Glucose Effectiveness in Type 2 Diabetes
Authors: Hawkins Meredith , Tonelli Julia , Kishore Preeti , Stein Daniel , Ragucci Enzo , Gitig Alon , Reddy Kalpana
Published: March 2007

Increased circulating free fatty acids (FFAs) inhibit both hepatic and peripheral insulin action. Because the loss of effectiveness of glucose to suppress endogenous glucose production and stimulate glucose uptake contributes importantly to fasting hyperglycemia in type 2 diabetes, we examined whether the approximate twofold elevations in FFA characteristic of poorly controlled type 2 diabetes contribute to this defect. Glucose levels were raised from 5 to 10 mmol/l while maintaining fixed hormonal conditions by infusing somatostatin with basal insulin, glucagon, and growth hormone. Each individual was studied at two FFA levels: with (NA+) and without (NA−) infusion of nicotinic acid in nine individuals with poorly controlled type 2 diabetes (HbA1c = 10.1 ± 0.7%) and with (LIP+) and without (LIP−) infusion of lipid emulsion in nine nondiabetic individuals. Elevating FFA to ∼500 μmol/l blunted the ability of glucose to suppress endogenous glucose production (LIP− = −48% vs. LIP+ = −28%; P < 0.01) and increased glucose uptake (LIP− = 97% vs. LIP+ = 51%; P < 0.01) in nondiabetic individuals. Raising FFA also blunted the endogenous glucose production response in 10 individuals with type 2 diabetes in good control (HbA1c = 6.3 ± 0.3%). Conversely, normalizing FFA nearly restored the endogenous glucose production (NA− = −7% vs. NA+ = −41%; P < 0.001) and glucose uptake (NA− = 26% vs. NA+ = 64%; P < 0.001) responses to hyperglycemia in individuals with poorly controlled type 2 diabetes. Thus, increased FFA levels contribute substantially to the loss of glucose effectiveness in poorly controlled type 2 diabetes.
Extracellular Glucose in Rat Ventromedial Hypothalamus During Acute and Recurrent Hypoglycemia
Authors: de Vries Martin G. , Arseneau Linda M. , Lawson Marcus E. , Beverly Joseph L.
Published: March 2007

The activity of neurons in the ventromedial hypothalamus (VMH) important for initiating compensatory responses to hypoglycemia is influenced by ambient glucose concentration. In the present study, we used in vivo microdialysis to evaluate interstitial glucose concentrations in rat VMH under various glycemic conditions. Using the zero-net-flux method, steady-state glucose concentration in the VMH was ∼20% of blood glucose (∼1.4 mmol/l) in fed rats but ∼14% of blood glucose (∼0.7 mmol/l) in overnight-fasted rats. During moderate hypoglycemia VMH glucose declined in parallel with blood glucose; however, VMH glucose decreased to a greater degree than blood glucose during a more severe hypoglycemic episode, falling to 10 ± 1.2% of blood levels (P < 0.01). To determine whether VMH glucose concentrations were influenced by recurrent episodes of hypoglycemia a second zero-net-flux study was conducted. Steady-state glucose concentrations in the VMH were ∼20% lower after three episodes of recurrent hypoglycemia, a value 17.8 ± 0.8% of blood glucose, although the relative change in VMH glucose levels during the first and fourth hypoglycemic episodes were similar. From these results, we conclude that interstitial glucose concentrations in the VMH are not maintained at a constant level and are more dynamic than previously proposed.
Leptin and Wound Inflammation in Diabeticob/obMice
Authors: Goren Itamar , Kämpfer Heiko , Podda Maurizio , Pfeilschifter Josef , Frank Stefan
Published: March 2007

In this study, we investigated the role of leptin for the inflammatory response in diabetes-impaired skin repair. We demonstrated, that systemic treatment of diabetic ob/ob mice with leptin blunted polymorphonuclear neutrophil (PMN), but not macrophage influx into the wound site. Closed wounds of leptin-administered mice were characterized by tremendous numbers of macrophage within the granulation tissue. In line, leptin supplementation potently attenuated epithelium-derived CXC- but not CC-chemokine expression. PMNs were preferentially located in the scab, but macrophages predominantly resided within the wound stroma of the animals. The scabs of nonhealing wounds were most likely to serve as sinks for bioactive inflammatory mediators, which were still capable to drive gene expression in keratinocytes in vitro. Differential effects of leptin on PMN and macrophage axes of inflammation must be indirect, as topical administration of leptin onto wounds of ob/ob mice did not reduce PMN influx into the wounded areas. Moreover, caloric-restricted, pair-fed ob/ob mice were characterized by impaired healing conditions that were associated with persisting PMNs. Interestingly, we documented the absence of leptin receptor expression in human diabetic foot ulcers. Thus, we show that leptin might function as a regulatory link between the endocrine and the immune system in the context of skin repair.
Effect of Type 1 Diabetes on the Gender Difference in Coronary Artery Calcification: a Role for Insulin Resistance?
Authors: Dabelea Dana , Kinney Gregory , Snell-Bergeon Janet K. , Hokanson John E. , Eckel Robert H. , Ehrlich James , Garg Satish , Hamman Richard F. , Rewers Marian
Published: March 2007

The objective of this is study was to examine whether estimated insulin resistance and insulin resistance-related factors are associated with coronary artery calcification (CAC) in 1,420 asymptomatic participants in the Coronary Artery Calcification in Type 1 Diabetes (CACTI) study. A total of 656 patients with type 1 diabetes and 764 control subjects aged 20–55 years were examined. CAC was assessed by electron-beam computed tomography. Insulin resistance was computed with linear regression based on an equation previously validated in clamp studies on type 1 diabetic adults. Insulin resistance was associated with CAC (OR 1.6 in type 1 diabetes and 1.4 in control subjects, P < 0.001), independent of coronary artery disease risk factors. There was a male excess of CAC in control subjects (OR 2.7, adjusted for age, smoking, and LDL and HDL cholesterol levels) and in type 1 diabetic patients (OR 2.2, adjusted for the same factors and diabetes duration). After adjusting for insulin resistance, the CAC male excess in diabetic patients decreased from OR 2.2 (P < 0.001) to 1.8 (P = 0.04). After adjustment for waist-to-hip ratio, waist circumference, or visceral fat, the gender difference in CAC was not significant in diabetic subjects. In conclusion, gender differences in insulin resistance-associated fat distribution may explain why type 1 diabetes increases coronary calcification in women relatively more than in men.
Association of Solute Carrier Family 12 (Sodium/Chloride) Member 3 With Diabetic Nephropathy, Identified by Genome-Wide Analyses of Single Nucleotide Polymorphisms
Authors: Tanaka Nobue , Babazono Tetsuya , Saito Susumu , Sekine Akihiro , Tsunoda Tatsuhiko , Haneda Masakazu , Tanaka Yasushi , Fujioka Tomoaki , Kaku Kohei , Kawamori Ryuzou , Kikkawa Ryuichi , Iwamoto Yasuhiko , Nakamura Yusuke , Maeda Shiro
Published: March 2007

To identify genetic elements that might confer susceptibility to diabetic nephropathy, we performed a genome-wide analysis of gene-based single nucleotide polymorphisms (SNPs) in a large cohort of Japanese patients with diabetes. In case-control association studies, patients with type 2 diabetes were divided into two groups, one having retinopathy as well as overt nephropathy and the other (the control group) having diabetic retinopathy but with no signs of renal involvement. Genotyping of these patients at >55,000 SNP loci indicated a gene encoding solute carrier family 12 member 3 (SLC12A3) to be a good candidate for the susceptibility to diabetic nephropathy, in view of a significant association of one landmark SNP located in the 24th intron (χ2 = 15.4, P = 0.000087, odds ratio = 2.53 [95% CI 1.57–4.09]). Subsequent analysis of additional genetic variations in this gene identified several SNPs that were significantly associated with nephropathy, especially one in exon 23 (+78 G to A: Arg913Gln, χ2 = 18.5, P = 0.00002, odds ratio = 2.53 [95% CI 1.64–3.90]). The results implicated that substitution of Arg913 to Gln in the SLC12A3 gene might reduce the risk to develop diabetic nephropathy and suggested that the gene product might be a potential target for the prevention or treatment of this disease.
Oxidative Stress Induces Nucleo-Cytoplasmic Translocation of Pancreatic Transcription Factor PDX-1 Through Activation of c-Jun NH2-terminal Kinase
Authors: Kawamori Dan , Kajimoto Yoshitaka , Kaneto Hideaki , Umayahara Yutaka , Fujitani Yoshio , Miyatsuka Takeshi , Watada Hirotaka , Leibiger Ingo B. , Yamasaki Yoshimitsu , Hori Masatsugu
Published: March 2007

Oxidative stress is induced in pancreatic β-cells under diabetic conditions and causes β-cell dysfunction. Antioxidant treatment of diabetic animals leads to recovery of insulin biosynthesis and increases the expression of its controlling transcription factor, pancreatic duodenal homeobox-1 (PDX-1), in pancreatic β-cells. Here, we show that PDX-1 is translocated from the nuclei to the cytoplasm of pancreatic β-cells in response to oxidative stress. When oxidative stress was charged upon β-cell-derived HIT-T15 cells, both endogenous PDX-1 and exogenously introduced green fluorescent protein-tagged PDX-1 moved from the nuclei to the cytoplasm. The addition of a dominant negative form of c-Jun NH2-terminal kinase (JNK) inhibited oxidative stress-induced PDX-1 translocation, suggesting an essential role of JNK in mediating this phenomenon. Whereas the nuclear localization signal (NLS) in PDX-1 was not affected by oxidative stress, leptomycin B, a specific inhibitor of the classical leucine-rich nuclear export signal (NES), inhibited nucleo-cytoplasmic translocation of PDX-1 induced by oxidative stress. Moreover, we identified an NES at position 82-94 of the mouse PDX-1 protein. Thus, our present results revealed a novel mechanism that negatively regulates PDX-1 function. The identification of the NES, which overrides the function of the NLS in an oxidative stress-responsive, JNK-dependent manner, supports the complicated regulation of PDX-1 function in vivo and may further the understanding of β-cell pathophysiology in diabetes.
Hepatic Akt Activation Induces Marked Hypoglycemia, Hepatomegaly, and Hypertriglyceridemia With Sterol Regulatory Element Binding Protein Involvement
Authors: Ono Hiraku , Shimano Hitoshi , Katagiri Hideki , Yahagi Naoya , Sakoda Hideyuki , Onishi Yukiko , Anai Motonobu , Ogihara Takehide , Fujishiro Midori , Viana Amelia Y.I. , Fukushima Yasushi , Abe Miho , Shojima Nobuhiro , Kikuchi Masatoshi , Yamada Nobuhiro , Oka Yoshitomo , Asano Tomoichiro
Published: March 2007

Akt is critical in insulin-induced metabolism of glucose and lipids. To investigate functions induced by hepatic Akt activation, a constitutively active Akt, NH2-terminally myristoylation signal-attached Akt (myr-Akt), was overexpressed in the liver by injecting its adenovirus into mice. Hepatic myr-Akt overexpression resulted in a markedly hypoglycemic, hypoinsulinemic, and hypertriglyceridemic phenotype with fatty liver and hepatomegaly. To elucidate the sterol regulatory element binding protein (SREBP)-1c contribution to these phenotypic features, myr-Akt adenovirus was injected into SREBP-1 knockout mice. myr-Akt overexpression induced hypoglycemia and hepatomegaly with triglyceride accumulation in SREBP-1 knockout mice to a degree similar to that in normal mice, whereas myr-Akt-induced hypertriglyceridemia in knockout mice was milder than that in normal mice. The myr-Akt-induced changes in glucokinase, phosphofructokinase, glucose-6-phosphatase, and PEPCK expressions were not affected by knocking out SREBP-1, whereas stearoyl-CoA desaturase 1 induction was completely inhibited in knockout mice. Constitutively active SREBP-1-overexpressing mice had fatty livers without hepatomegaly, hypoglycemia, or hypertriglyceridemia. Hepatic acetyl-CoA carboxylase, fatty acid synthase, stearoyl-CoA desaturase 1, and glucose-6-phosphate dehydrogenase expressions were significantly increased by overexpressing SREBP-1, whereas glucokinase, phospho-fructokinase, glucose-6-phosphatase, and PEPCK expressions were not or only slightly affected. Thus, SREBP-1 is not absolutely necessary for the hepatic Akt-mediated hypoglycemic effect. In contrast, myr-Akt-induced hypertriglyceridemia and hepatic triglyceride accumulation are mediated by both Akt-induced SREBP-1 expression and a mechanism involving fatty acid synthesis independent of SREBP-1.
Adrenalectomy Alters the Sensitivity of the Central Nervous System Melanocortin System
Authors: Drazen Deborah L. , Wortman Matthew D. , Schwartz Michael W. , Clegg Deborah J. , van Dijk Gertjan , Woods Stephen C. , Seeley Randy J.
Published: March 2007

Removal of adrenal steroids by adrenalectomy (ADX) reduces food intake and body weight in rodents and prevents excessive weight gain in many genetic and dietary models of obesity. Thus, glucocorticoids appear to play a key role to promote positive energy balance in normal and pathological conditions. By comparison, central nervous system melanocortin signaling provides critical inhibitory tone to regulate energy balance. The present experiments sought to test whether glucocorticoids influence energy balance by altering the sensitivity to melanocortin receptor ligands. Because melanocortin-producing neurons are hypothesized to be downstream of leptin in a key weight-reducing circuit, we tested rats for their sensitivity to leptin and confirmed reports that the hypophagic response to third ventricular (i3vt) leptin is increased in ADX rats and is normalized by glucocorticoid replacement. Next we tested rats for their sensitivity to the melanocortin agonist melanotan II and found that, as for leptin, ADX enhanced the hypophagic response via a glucocorticoid-dependent mechanism. The central nervous system melanocortin system is unique in that it includes the endogenous melanocortin receptor antagonist, AgRP. The orexigenic effect of i3vt AgRP was absent in ADX rats and restored by glucocorticoid replacement. We conclude that the potent weight-reducing effects of ADX likely involve heightened responsiveness to melanocortin receptor stimulation.
Activation of Vascular Endothelial Growth Factor Receptor-1 Sustains Angiogenesis and Bcl-2 Expression Via the Phosphatidylinositol 3-Kinase Pathway in Endothelial Cells
Authors: Cai Jun , Ahmad Shakil , Jiang Wen G. , Huang Jianhua , Kontos Christopher D. , Boulton Mike , Ahmed Asif
Published: March 2007

Vascular insufficiency and retinal ischemia precede many proliferative retinopathies and stimulate secretion of various vasoactive growth factors, including vascular endothelial growth factor (VEGF) and placenta growth factor (PlGF). It is unclear, however, how PlGF, which is elevated in proliferative diabetic retinopathy and is a VEGF homolog that binds only to VEGF receptor (VEGFR)-1, promotes pathological angiogenesis. When primary microvascular endothelial cells were grown on collagen gels, PlGF-containing ligands upregulated Bcl-2 expression and stimulated the formation of capillary-like tube networks that were retained for up to 14 days in culture. The inhibition of VEGFR-1 results in a dramatic decrease in the number of capillary connections, indicating that VEGFR-1 ligands promote branching angiogenesis. In contrast, VEGF-induced tube formations and Bcl-2 expression were significantly decreased at the end of this period. Flow cytometry analysis of annexin-V/propidium iodide–stained cells revealed that PlGF and PlGF/VEGF heterodimer inhibited apoptosis in serum-deprived endothelial cells. These two growth factors stimulated a survival signaling pathway phosphatidylinositol 3-kinase (PI3K), as identified by increased Akt phosphorylation and because blocking PI3K signalling by adenovirus-mediated overexpression of wild-type phosphatase and tensin homolog on chromosome 10 (PTEN) disrupted angiogenesis and decreased Bcl-2 expression by PlGF and PlGF/VEGF heterodimer, whereas a dominant-negative PTEN mutant enhanced endothelial sprout formation and Bcl-2 expression. Together, these findings indicate that PlGF-containing ligands contribute to pathological angiogenesis by prolonging cell survival signals and maintaining vascular networks.
Autosomal-Dominant Mode of Inheritance of a Melanocortin-4 Receptor Mutation in a Patient with Severe Early-Onset Obesity Is Due to a Dominant-Negative Effect Caused by Receptor Dimerization
Authors: Biebermann Heike , Krude Heiko , Elsner Andrea , Chubanov Vladimir , Gudermann Thomas , Grüters Annette
Published: March 2007

Mutations in the melanocortin-4 receptor (MC4R) gene are the most frequent monogenic causes of severe obesity. Most mutations have been described as heterozygous with loss of function, suggesting that haploinsufficiency is the most likely mechanism of dominant inheritance. We detected a heterozygous mutation, D90N, in a patient with severe early-onset obesity. Functional characterization of the mutant receptor revealed normal cell-surface expression and binding properties but loss of signal transduction activity. In coexpression studies of wild-type (WT)-MC4R and D90N, the mutant receptor had a dominant-negative effect on WT-receptor function. Further investigation of this effect with sandwich enzyme-linked immunosorbent assays and fluorescence resonance energy transfer studies showed that the WT-MC4R and the D90N mutant form homodimers and heterodimers. We hypothesize that the dominant-negative effect of the D90N mutation is caused by a functionally altered WT-MC4R/D90N receptor heterodimer. These findings necessitate the reinvestigation of other heterozygous MC4R missense mutations to discriminate between haploinsufficiency and a dominant-negative effect. The finding of receptor dimerization highlights a more complex hypothalamic signaling network governing the regulation of body weight.
Increased Intramyocellular Lipid Concentration Identifies Impaired Glucose Metabolism in Women With Previous Gestational Diabetes
Authors: Kautzky-Willer Alexandra , Krssak Martin , Winzer Christine , Pacini Giovanni , Tura Andrea , Farhan Serdar , Wagner Oswald , Brabant Georg , Horn Rüdiger , Stingl Harald , Schneider Barbara , Waldhäusl Werner , Roden Michael
Published: March 2007

Women with previous gestational diabetes (pGDM) are frequently insulin-resistant, which could relate to intramyocellular lipid content (IMCL). IMCL were measured with 1H nuclear magnetic resonance spectroscopy in soleus (IMCL-S) and tibialis-anterior muscles (IMCL-T) of 39 pGDM (32 ± 2 years, waist-to-hip ratio 0.81 ± 0.01) and 22 women with normal glucose tolerance (NGT; 31 ± 1 years, 0.76 ± 0.02) at 4–6 months after delivery. Body fat mass (BFM) was assessed from bioimpedance analysis, insulin sensitivity index (SI), and glucose effectiveness (SG) from insulin-modified frequently sampled glucose tolerance tests. pGDM exhibited 45% increased BFM, 35% reduced SI and SG (P < 0.05), and 40% (P < 0.05) and 55% (P < 0.005) higher IMCL-S and IMCL-T, respectively. IMCL related to body fat (BFM P < 0.005, leptin P < 0.03), but only IMCL-T correlated (P < 0.03) with SI and glucose tolerance index independent of BMI. Insulin-resistant pGDM (n = 17) had higher IMCL-S (+66%) and IMCL-T (+86%) than NGT and insulin-sensitive pGDM (+28%). IMCL were also higher (P < 0.005, P = 0.05) in insulin-sensitive pGDM requiring insulin treatment during pregnancy and inversely related to the gestational week of GDM diagnosis. Thus, IMCL-T reflects insulin sensitivity, whereas IMCL-S relates to obesity. IMCL could serve as an additional parameter of increased diabetes risk because it identifies insulin-resistant pGDM and those who were diagnosed earlier and/or required insulin during pregnancy.
PPAR-γ Activation Mediates Adipose Depot−Specific Effects on Gene Expression and Lipoprotein Lipase Activity
Authors: Laplante Mathieu , Sell Henrike , MacNaul Karen L. , Richard Denis , Berger Joel P. , Deshaies Yves
Published: March 2007

This study sought to determine whether the adipose depot−specific (subcutaneous [SF] vs. visceral [VF]) action of peroxisome proliferator−activated receptor-γ (PPAR-γ) agonists on fat deposition extends to the expression of lipoprotein lipase (LPL) and other key adipose lipid metabolism genes, and whether changes in LPL impact triglyceridemia. Rats were fed a standard diet or an obesity-promoting diet for 3 weeks, with or without treatment with COOH, a nonthiazolidinedione PPAR-γ agonist. Treatment effects were essentially similar in both dietary cohorts. COOH did not affect weight gain, but increased SF (inguinal) fat mass twofold and reduced VF (retroperitoneal) accretion by half. Corresponding depot-specific alterations were observed in mRNA levels of the glucocorticoid-activating enzyme 11β-hydroxysteroid dehydrogenase 1 (11β-HSD-1) and the thermogenic modulator uncoupling protein 1 (UCP-1). COOH increased brown adipose tissue (BAT) weight and LPL availability by five- to eightfold. In rats refed standard diet after a 24-h fast, COOH reduced the insulin excursion by half. The agonist increased SF LPL activity and mRNA levels, but had no effect on VF LPL. The two- to threefold postprandial increase in plasma triglycerides (TGs) was abrogated in COOH-treated rats, likely in part because of increased LPL in SF and BAT. Thus PPAR-γ agonist treatment had a powerful, site-specific effect on adipose metabolism and lipid deposition, and greatly impacted the postprandial handling of TG-rich lipoproteins. These depot-specific effects may be mediated by differential regulation of key metabolic genes, including LPL, 11β-HSD-1, and UCP-1.
Role of the Fatty Acid Binding Protein mal1 in Obesity and Insulin Resistance
Authors: Maeda Kazuhisa , Uysal K. Teoman , Makowski Liza , Görgün Cem Z. , Atsumi Genichi , Parker Rex A. , Brüning Jens , Hertzel Ann Vogel , Bernlohr David A. , Hotamisligil Gökhan S.
Published: March 2007

The metabolic syndrome is a cluster of metabolic and inflammatory abnormalities including obesity, insulin resistance, type 2 diabetes, hypertension, dyslipidemia, and atherosclerosis. The fatty acid binding proteins aP2 (fatty acid binding protein [FABP]-4) and mal1 (FABP5) are closely related and both are expressed in adipocytes. Previous studies in aP2-deficient mice have indicated a significant role for aP2 in obesity-related insulin resistance, type 2 diabetes, and atherosclerosis. However, the biological functions of mal1 are not known. Here, we report the generation of mice with targeted null mutations in the mal1 gene as well as transgenic mice overexpressing mal1 from the aP2 promoter/enhancer to address the role of this FABP in metabolic regulation in the presence or absence of obesity. To address the role of the second adipocyte FABP in metabolic regulation in the presence and deficiency of obesity, absence of mal1 resulted in increased systemic insulin sensitivity in two models of obesity and insulin resistance. Adipocytes isolated from mal1-deficient mice also exhibited enhanced insulin-stimulated glucose transport capacity. In contrast, mice expressing high levels of mal1 in adipose tissue display reduced systemic insulin sensitivity. Hence, our results demonstrate that mal1 modulates adipose tissue function and contributes to systemic glucose metabolism and constitutes a potential therapeutic target in insulin resistance.
Molecular Regulation of Monocyte Chemoattractant Protein-1 Expression in Pancreatic β-Cells
Authors: Kutlu Burak , Darville Martine I. , Cardozo Alessandra K. , Eizirik Décio L.
Published: March 2007

Pancreatic β-cells are selectively destroyed during the course of type 1 diabetes. In the early stages of the disease, inflammatory infiltrates of mononuclear cells, containing predominantly monocytes and T-cells, are present in the islets (insulitis). Chemokines, such as monocyte chemoattractant protein-1 (MCP-1), play a key role in the recruitment and activation of these immunocytes. We have previously described cytokine-induced MCP-1 gene expression in human and rat pancreatic islets. In the present study, the transcriptional regulation by cytokines of the rat MCP-1 gene in fluorescence-activated cell sorting-purified rat β-cells, insulin-producing INS-1E cells, and RINm5F cells was investigated. Transient transfections with luciferase-reporter constructs identified an interleukin (IL)-1β-responsive enhancer region between -2,180 bp and −2,478 bp. Mutation of either of the two nuclear factor (NF)-κB sites present in this region abrogated IL-1β-induced MCP-1 promoter activity. Binding of NF-κB to the two sites was shown in vitro by gel shift assays, while supershift assays revealed the presence of p65/p50 heterodimers and p65 homodimers. In vivo binding of NF-κB was confirmed by chromatin immunoprecipitation assay. Blocking of NF-κB activation in cytokine-exposed primary β-cells by an adenovirus overexpressing a nondegradable form of IκBα or by pyrrolidine dithiocarbamate decreased IL-1β-induced MCP-1 mRNA expression. We conclude that NF-κB plays an important role for MCP-1 expression in β-cells. This transcription factor may be an interesting target for ex vivo gene therapy before islet transplantation.
Role of Endogenous Glucagon-Like Peptide-1 in Islet Regeneration After Partial Pancreatectomy
Authors: De León Diva D. , Deng Shaoping , Madani Reza , Ahima Rexford S. , Drucker Daniel J. , Stoffers Doris A.
Published: March 2007

A reduction in β-cell mass is an important causative factor in type 1 and type 2 diabetes. Glucagon-like peptide-1 (GLP-1) and the long-acting agonist exendin 4 (Ex-4) expand β-cell mass by stimulating neogenesis and proliferation. In the partial pancreatectomy (Ppx) model, exogenous Ex-4 promotes islet regeneration, leading to sustained improvement in glucose tolerance. In this study, we investigate the potential role of endogenous GLP-1 in islet growth. We examined β-cell mass regeneration after 70% Ppx in mice receiving the GLP-1 antagonist Ex9-39 and in GLP-1R−/− mice. In Ex9-39–treated sham-operated mice, persistent fasting hyperglycemia was observed, but β-cell mass was not diminished. In pancreatectomized mice, persistent glucose intolerance was noted, but this was not further exacerbated by Ex9-39. Accordingly, β-cell mass recovery of Ppx mice was not impaired by Ex9-39. In contrast, GLP-1R−/− CD1 mice showed worse glucose intolerance after Ppx compared with wild-type CD1 Ppx mice, and this correlated with a significant defect in β-cell mass regeneration. The recovery of β-cell mass differed markedly in the BALB/c and CD1 control mice, indicating a significant role of genetic background in the regulation of β-cell mass. These studies point to a role for endogenous GLP-1 in β-cell regeneration after Ppx in mice.
Differential Effects of Glucose and Glyburide on Energetics and Na+ Levels of βHC9 Cells
Authors: Doliba Nicolai M. , Vatamaniuk Marko Z. , Buettger Carol W. , Qin Wei , Collins Heather W. , Wehrli Suzanne L. , Carr Richard D. , Matschinsky Franz M.
Published: March 2007

In the present study, noninvasive 31P and 23Na+–nuclear magnetic resonance (NMR) technology and respirometry were used to compare the effect of high glucose (30 mmol/l) with the effect of the antidiabetic sulfonylurea (SU) compound glyburide (GLY) on energy metabolism, Na+ flux, insulin, and cAMP release of continuously superfused β-HC9 cells encapsulated in microscopic agarose beads. Both high glucose and GLY increased oxygen consumption in β-HC9 cells (15–30%) with a maximal effect at 8 mmol/l for glucose and at 250 nmol/l for GLY. At the same time, insulin release from β-cells increased by 15- and 25-fold with high glucose or GLY, respectively. The P-creatine (PCr) level was greatly increased and inorganic phosphate (Pi) was decreased with 30 mmol/l glucose in contrast to the decreased level of PCr and increased Pi with GLY. ATP levels remained unchanged during both interventions. Studies on isolated mitochondria of β-HC9 cells showed that GLY added to mitochondria oxidizing glutamine or glutamate abolished the stimulation of respiration by ADP (state 3) meanwhile leaving state 3 respiration unchanged during oxidation of other substrates. Exposure of β-HC9 cells to 5 mmol/l glucose decreased intracellular Na+ levels monitored by 23Na+-NMR spectroscopy and 30 mmol/l glucose resulted in a further decrease in cytosolic Na+. In contrast, Na+ increased when 1 μmol/l GLY was added to the perfusate containing 5 mmol/l glucose. These data support the hypothesis that glucose activates the β-cell through a “push mechanism” due to substrate pressure enhancing fuel flux, energy production, and extrusion of Na+ from the cells in contrast to SU receptor (SUR)-1 inhibitors, which may modify intermediary and energy metabolism secondarily through a “pull mechanism” due to higher energy demand resulting from increased ion fluxes and the exocytotic work load.
Inflammation-Sensitive Plasma Proteins, Diabetes, and Mortality and Incidence of Myocardial Infarction and Stroke
Authors: Engström Gunnar , Stavenow Lars , Hedblad Bo , Lind Peter , Eriksson Karl-Fredrik , Janzon Lars , Lindgärde Folke
Published: March 2007

This study explores the relationship of inflammation-sensitive plasma proteins (ISPs) with the prevalence of diabetes and the interrelationships between ISPs and diabetes in the prediction of death and incidence of myocardial infarction and stroke. Plasma levels of fibrinogen, α1-antitrypsin, haptoglobin, ceruloplasmin, and orosomucoid were assessed in 6,050 men, aged 28–61 years. All-cause and cardiovascular mortality and incidence of myocardial infarction and stroke were monitored over 18.7 ± 3.7 years. Prevalence of diabetes (n = 321) was significantly associated with ISP levels among overweight and obese men but not among men with BMI <25 kg/m2. The association was similar for insulin resistance according to homeostasis model assessment. High ISP levels (two or more ISPs in the top quartile) increased the cardiovascular risk among diabetic men. The risk factor-adjusted relative risks for cardiovascular mortality, myocardial infarction, and stroke were 2.8 (CI 1.8–4.5), 2.2 (1.5–3.2), and 2.5 (1.4–4.6), respectively, for diabetic men with high ISP levels (reference: nondiabetic men with low ISP levels). The corresponding risks for diabetic men with low ISP levels were 1.8 (1.1–3.0), 1.3 (0.8–2.1), and 1.2 (0.6–2.5), respectively. In conclusion, in this population-based cohort, diabetes was associated with increased ISP levels among overweight and obese men but not among men with normal weight. High ISP levels increased the cardiovascular risk similarly in diabetic as compared with nondiabetic men.
A Role for the Polyol Pathway in the Early Neuroretinal Apoptosis and Glial Changes Induced by Diabetes in the Rat
Authors: Asnaghi Veronica , Gerhardinger Chiara , Hoehn Todd , Adeboje Abidemi , Lorenzi Mara
Published: March 2007

We tested the hypothesis that the apoptosis of inner retina neurons and increased expression of glial fibrillary acidic protein (GFAP) observed in the rat after a short duration of diabetes are mediated by polyol pathway activity. Rats with 10 weeks of streptozotocin-induced diabetes and GHb levels of 16 ± 2% (mean ± SD) showed increased retinal levels of sorbitol and fructose, attenuation of GFAP immunostaining in astrocytes, appearance of prominent GFAP expression in Müller glial cells, and a fourfold increase in the number of apoptotic neurons when compared with nondiabetic rats. The cells undergoing apoptosis were immunoreactive for aldose reductase. Sorbinil, an inhibitor of aldose reductase, prevented all abnormalities. Intensive insulin treatment also prevented most abnormalities, despite reducing GHb only to 12 ± 1%. Diabetic mice, known to have much lower aldose reductase activity in other tissues when compared with rats, did not accumulate sorbitol and fructose in the retina and were protected from neuronal apoptosis and GFAP changes in the presence of GHb levels of 14 ± 2%. This work documents discrete cellular consequences of polyol pathway activity in the retina, and it suggests that activation of the pathway and “retinal neuropathy” require severe hyperglycemia and/or high activity of aldose reductase. These findings have implications for how to evaluate the role of the polyol pathway in diabetic retinopathy.
Evidence for a Novel TGF-β1−Independent Mechanism of Fibronectin Production in Mesangial Cells Overexpressing Glucose Transporters
Authors: Weigert Cora , Brodbeck Katrin , Brosius III Frank C. , Huber Matthias , Lehmann Rainer , Friess Ulrich , Facchin Sonia , Aulwurm Steffen , Häring Hans U. , Schleicher Erwin D. , Heilig Charles W.
Published: March 2007

Recent experimental work indicates that the hyperglycemia-induced increase in mesangial matrix production, which is a hallmark in the development of diabetic nephropathy, is mediated by increased expression of GLUT1. Mesangial cells stably transfected with human GLUT1 mimic the effect of hyperglycemia on the production of the extracellular matrix proteins, particularly fibronectin, when cultured under normoglycemic conditions. Our investigation of the molecular mechanism of this effect has revealed that the enhanced fibronectin production was not mediated by the prosclerotic cytokine transforming growth factor (TGF)-β1. We found markedly increased nuclear content in Jun proteins, leading to enhanced DNA-binding activity of activating protein 1 (AP-1). AP-1 inhibition reduced fibronectin production in a dosage-dependent manner. Moreover, inhibition of classic protein kinase C (PKC) isoforms prevented both the activation of AP-1 and the enhanced fibronectin production. In contrast to mesangial cells exposed to high glucose, no activation of the hexosamine biosynthetic, p38, or extracellular signal–related kinase 1 and 2 mitogen-activated protein kinase pathways nor any increase in TGF-β1 synthesis could be detected, which could be explained by the absence of oxidative stress in cells transfected with the human GLUT1 gene. Our data indicate that increased glucose uptake and metabolism induce PKC-dependent AP-1 activation that is sufficient for enhanced fibronectin production, but not for increased TGF-β1 expression.
Microvascular Alterations in Diabetic Mice Correlate With Level of Hyperglycemia
Authors: Algenstaedt Petra , Schaefer Christian , Biermann Tanja , Hamann Andreas , Schwarzloh Britta , Greten Heiner , Rüther Wolfgang , Hansen-Algenstaedt Nils
Published: March 2007

Vascular alterations are the most common causes of morbidity and mortality in diabetic patients. Despite the impact of endothelial dysfunction on microcirculatory properties, little is known about the endothelial cell alteration during the development of diabetes and its correlation to the metabolic situation. For that reason we continuously monitored in vivo functional and morphological alterations of the microvasculature in hyperglycemic and hyperinsulinemic transgenic UCP1/DTA mice with brown fat deficiency, using a dorsal skin-fold chamber preparation and fluorescence microscopy. UCP1/DTA mice showed a dramatic decrease in vascular density due to a remarkable reduction of small vessels. Vascular permeability and leukocyte endothelial interactions (LEIs) significantly increased. The extent of vascular alteration correlated with the extent of metabolic dysfunction. Decreased tissue perfusion observed in UCP1/DTA mice might play a role in impaired wound healing observed in diabetes. The increased permeability in subcutaneous tissue may serve as predictor of vascular changes in early stages of diabetes. The increased LEI and serum tumor necrosis factor-α levels, which mirror the inflammatory process, support the growing evidence of the inflammatory component of diabetic disease. The results suggest that anti-inflammatory strategies might be able to prevent vascular deterioration in early stages of diabetes. Further investigations are required to evaluate the benefit of such therapeutic strategies.