bluuebunny/crossref_metadata_2025 huggingface dataset:
Pathways in Beta-Cell Stimulus-Secretion Coupling as Targets for Therapeutic Insulin Secretagogues
Authors: Henquin Jean-Claude
Published: March 2007

Physiologically, insulin secretion is subject to a dual, hierarchal control by triggering and amplifying pathways. By closing ATP-sensitive K+ channels (KATP channels) in the plasma membrane, glucose and other metabolized nutrients depolarize β-cells, stimulate Ca2+ influx, and increase the cytosolic concentration of free Ca2+ ([Ca2+]i), which constitutes the indispensable triggering signal to induce exocytosis of insulin granules. The increase in β-cell metabolism also generates amplifying signals that augment the efficacy of Ca2+ on the exocytotic machinery. Stimulatory hormones and neurotransmitters modestly increase the triggering signal and strongly activate amplifying pathways biochemically distinct from that set into operation by nutrients. Many drugs can increase insulin secretion in vitro, but only few have a therapeutic potential. This review identifies six major pathways or sites of stimulus-secretion coupling that could be aimed by potential insulin-secreting drugs and describes several strategies to reach these targets. It also discusses whether these perspectives are realistic or theoretical only. These six possible β-cell targets are 1) stimulation of metabolism, 2) increase of [Ca2+]i by closure of K+ATP channels, 3) increase of [Ca2+]i by other means, 4) stimulation of amplifying pathways, 5) action on membrane receptors, and 6) action on nuclear receptors. The theoretical risk of inappropriate insulin secretion and, hence, of hypoglycemia linked to these different approaches is also envisaged.
Impact of Adenoviral Transduction With SREBP1c or AMPK on Pancreatic Islet Gene Expression Profile
Authors: Diraison Frederique , Motakis Efthimios , Parton Laura E. , Nason Guy P. , Leclerc Isabelle , Rutter Guy A.
Published: March 2007

Accumulation of triglyceride in islets may contribute to the loss of glucose-stimulated insulin secretion (GSIS) in some forms of type 2 diabetes (Diraison et al., Biochem J 373:769–778, 2004). Here, we use adenoviral vectors and oligonucleotide microarrays to determine the effects of the forced expression of SREBP1c on the gene expression profile of rat islets. Sterol regulatory element binding protein-1c (SREBP1c) overexpression led to highly significant (P < 0.1 with respect to null adenovirus) changes in the expression of 1,238 genes or expressed sequence tags, of which 1,180 (95.3%) were upregulated. By contrast, overexpression of constitutively active AMP-activated protein kinase (AMPK), expected to promote lipolysis, altered the expression of 752 genes, of which 702 (93%) were upregulated. To identify specific targets for SREBP1c or AMPK, we eliminated messages that were 1) affected in the same direction by the expression of either protein, 2) changed by less than twofold, or 3) failed a positive false discovery test; 206 SREBP1c-regulated genes (195; 95% upregulated) and 48 AMPK-regulated genes (33; 69% upregulated) remained. As expected, SREBP1c-induced genes included those involved in cholesterol (6), fatty acid (3), and eicosanoid synthesis. Interestingly, somatostatin receptor (sstr1) expression was increased by SREBP1c, whereas AMPK induced the expression of peptide YY, the early endocrine pancreas marker.
The Effect of Vitamin E on Endothelial Function of Micro- and Macrocirculation and Left Ventricular Function in Type 1 and Type 2 Diabetic Patients
Authors: Economides Panayiotis A. , Khaodhiar Lalita , Caselli Antonella , Caballero A. Enrique , Keenan Hillary , Bursell Sven-Erik , King George L. , Johnstone Michael T. , Horton Edward S. , Veves Aristidis
Published: March 2007

We examined the effects of high-dosage vitamin E treatment over a 12-month period on the vascular reactivity of micro- and macrocirculation and left ventricular function in diabetic patients. Subjects (n = 89) were randomized to vitamin E (1,800 IU daily) or placebo and were followed for 12 months. High-resolution ultrasound images were used to measure the flow-mediated dilation (FMD; endothelium dependent) and nitroglycerin-induced dilation (NID; endothelium independent) of the brachial artery. Laser Doppler perfusion imaging was used to measure vascular reactivity in the forearm skin. Left ventricular function was evaluated using transthoracic echocardiogram. At the end of the 6-month period, a worsening in endothelium-dependent skin vasodilation (P = 0.02) and rise in endothelin levels (P = 0.01) were found in the vitamin E compared with the placebo group. At the end of the 12-month period, a worsening was observed in NID (P = 0.02) and a marginal worsening was seen in systolic blood pressure (P = 0.04) and FMD (P = 0.04) in the vitamin E compared with the placebo group. In addition C-reactive protein levels decreased marginally in the vitamin E compared with the placebo group (P = 0.05). No changes were observed in left ventricular function. We concluded that long-term treatment with 1,800 IU of vitamin E has no beneficial effects on endothelial or left ventricular function in diabetic patients. Because vitamin E–treated patients had a worsening in some vascular reactivity measurements when compared with control subjects, the use of high dosages of vitamin E cannot be recommended.
Nitrergic Neurodegeneration in Cerebral Arteries of Streptozotocin-Induced Diabetic Rats
Authors: Cellek Selim , Anderson Patrick N. , Foxwell Neale A.
Published: March 2007

Although autonomic neuropathy is recognized as an independent risk factor for stroke in diabetes, the mechanism by which autonomic nerves are involved in this pathology is unknown. Parasympathetic (cholinergic) nerves of the autonomic nervous system are known to innervate and to cause relaxation of cerebral arteries by releasing nitric oxide (NO); hence, they are called nitrergic nerves. However, the effect of diabetes on nitrergic nerves is unknown. Here, we show that perivascular nitrergic nerves around the cerebral arteries degenerate in two phases in streptozotocin-induced diabetic rats. In the first phase, perivascular nitrergic nerve fibers remain intact while they lose their neuronal NO synthase content. This phase is reversible with insulin treatment. In the second phase, nitrergic cell bodies in the ganglia are lost via apoptosis in an irreversible manner. Throughout the two phases, irreversible thickening of the smooth muscle layer of cerebral arteries is observed. This is the first demonstration of nitrergic degeneration in diabetic cerebral arteries, which could elucidate the link between diabetic autonomic neuropathy and stroke.
Altered Transcapillary Escape of Albumin and Microalbuminuria Reflects Two Different Pathogenetic Mechanisms
Authors: Nosadini Romano , Velussi Mario , Brocco Enrico , Abaterusso Cataldo , Piarulli Francesco , Morgia Giuseppe , Satta Andrea , Faedda Rossana , Abhyankar Avinash , Luthman Holger , Tonolo Giancarlo
Published: March 2007

We studied the following in normo- and microalbuminuric hypertensive type 2 diabetic patients: 1) transcapillary escape rate of albumin (TERalb) and 2) expression of mRNA slit diaphragm and podocyte proteins in renal biopsies. Normoalbuminuric subjects had renal cancer, and kidney biopsy was performed during surgery. TERalb was evaluated by clearance of 125I-albumin. Real-time PCR of mRNA slit diaphragm was measured in kidney specimens. Albumin excretion rate (AER) was by definition lower in normoalbuminuric subjects than in microalbuminuric subjects with typical diabetic glomerulopathy (group 1), in microalbuminuric subjects with normal or near-normal glomerular structure (group 2), and in microalbuminuric subjects with atypical diabetic nephropathy (group 3). This classification was based on light microscopy analysis of renal tissue. TERalb (%/h) was similar in normoalbuminuric and microalbuminuric group 1, 2, and 3 diabetic patients (medians: 14.1 vs. 14.4 vs. 15.7 vs. 14.9, respectively) (ANOVA, NS). mRNA expression of slit diaphragm proteins CD2AP, FAT, Actn 4, NPHS1, and NPHS2 was higher in normoalbuminuric patients than in microalbuminuric patients (groups 1, 2, and 3) (ANOVA, P < 0.001). All diabetic patients had greater carotid artery intimal thickness than normal control subjects using ultrasound technique (ANOVA, P < 0.01). In conclusion, the present study suggests that microalbuminuria identifies a subgroup of hypertensive type 2 diabetic patients who have altered mRNA expression of slit diaphragm and podocyte proteins, even before glomerular structure shows abnormalities using light microscopy analysis. On the contrary, altered TERalb and increased carotid artery intimal thickness are shown by all hypertensive type 2 diabetic patients, both with normal and altered patterns of AER.
Genetic Linkage and Association of the Growth Hormone Secretagogue Receptor (Ghrelin Receptor) Gene in Human Obesity
Authors: Baessler Andrea , Hasinoff Michael J. , Fischer Marcus , Reinhard Wibke , Sonnenberg Gabriele E. , Olivier Michael , Erdmann Jeanette , Schunkert Heribert , Doering Angela , Jacob Howard J. , Comuzzie Anthony G. , Kissebah Ahmed H. , Kwitek Anne E.
Published: March 2007

The growth hormone secretagogue receptor (GHSR) (ghrelin receptor) plays an important role in the regulation of food intake and energy homeostasis. The GHSR gene lies on human chromosome 3q26 within a quantitative trait locus strongly linked to multiple phenotypes related to obesity and the metabolic syndrome. Because the biological function and location of the GHSR gene make it an excellent candidate gene, we tested the relation between common single nucleotide polymorphisms (SNPs) in the GHSR gene and human obesity. We performed a comprehensive analysis of SNPs, linkage disequilibrium (LD), and haplotype structure across the entire GHSR gene region (99.3 kb) in 178 pedigrees with multiple obese members (DNA of 1,095 Caucasians) and in an independent sample of the general population (MONICA Augsburg left ventricular hypertrophy substudy; DNA of 1,418 Caucasians). The LD analysis revealed a disequilibrium block consisting of five SNPs, consistent in both study cohorts. We found linkage among all five SNPs, their haplotypes, and BMI. Further, we found suggestive evidence for transmission disequilibrium for the minor SNP alleles (P < 0.05) and the two most common haplotypes with the obesity affection status (“susceptible” P = 0.025, “nonsusceptible” P = 0.045) in the family cohort using the family-based association test program. Replication of these findings in the general population resulted in stronger evidence for an association of the SNPs (best P = 0.00001) and haplotypes with the disease (“susceptible” P = 0.002, “nonsusceptible” P = 0.002). To our knowledge, these data are the first to demonstrate linkage and association of SNPs and haplotypes within the GHSR gene region and human obesity. This linkage, together with significant transmission disequilibrium in families and replication of this association in an independent population, provides evidence that common SNPs and haplotypes within the GHSR region are involved in the pathogenesis of human obesity.
Defective Suppressor Function in CD4+CD25+ T-Cells From Patients With Type 1 Diabetes
Authors: Lindley Shelley , Dayan Colin M. , Bishop Amanda , Roep Bart O. , Peakman Mark , Tree Timothy I.M.
Published: March 2007

Type 1 diabetes is a T-cell–mediated disease that is associated with loss of immunological tolerance to self-antigens. The mechanisms involved in maintenance of peripheral tolerance include a specialized subset of regulatory T-cells (Treg) within the CD4+CD25+ T-cell population, but the function and phenotype of these cells in type 1 diabetes have not been investigated. We hypothesized that a deficiency in the CD4+CD25+ Treg population or its function could contribute to the lack of self-tolerance evident in patients with type 1 diabetes. We show that although levels of CD4+CD25+ T-cells are normal in patients with recent-onset adult type 1 diabetes, the ability of the Tregs in this population to suppress T-cell proliferation during in vitro cocultures is markedly reduced compared with control subjects (P = 0.007). Moreover, in patients with type 1 diabetes, these cocultures display a more proinflammatory phenotype, with increased secretion of interferon-γ (P = 0.005) and decreased interleukin-10 production (P = 0.03). These deficiencies may reflect a disturbance in the balance of the CD4+CD25+ population, because in patients with type 1 diabetes, a higher proportion of these cells coexpress the early activation marker CD69 (P = 0.007) and intracellular CTLA-4 (P = 0.01). These data demonstrate deficiency in function of the CD4+CD25+ Treg population that may influence the pathogenesis of type 1 diabetes.
Signals From the Embryonic Mouse Pancreas Induce Differentiation of Human Embryonic Stem Cells Into Insulin-Producing β-Cell–Like Cells
Authors: Brolén Gabriella K.C. , Heins Nico , Edsbagge Josefina , Semb Henrik
Published: March 2007

The recent success in restoring normoglycemia in type 1 diabetes by islet cell transplantation indicates that cell replacement therapy of this severe disease is achievable. However, the severe lack of donor islets has increased the demand for alternative sources of β-cells, such as adult and embryonic stem cells. Here, we investigate the potential of human embryonic stem cells (hESCs) to differentiate into β-cells. Spontaneous differentiation of hESCs under two-dimensional growth conditions resulted in differentiation of Pdx1+/Foxa2+ pancreatic progenitors and Pdx1+/Isl1+ endocrine progenitors but no insulin-producing cells. However, cotransplantation of differentiated hESCs with the dorsal pancreas, but not with the liver or telencephalon, from mouse embryos resulted in differentiation of β-cell–like cell clusters. Comparative analysis of the basic characteristics of hESC-derived insulin+ cell clusters with human adult islets demonstrated that the insulin+ cells share important features with normal β-cells, such as synthesis (proinsulin) and processing (C-peptide) of insulin and nuclear localization of key β-cell transcription factors, including Foxa2, Pdx1, and Isl1.
Tumor Necrosis Factor-α Induces Skeletal Muscle Insulin Resistance in Healthy Human Subjects via Inhibition of Akt Substrate 160 Phosphorylation
Authors: Plomgaard Peter , Bouzakri Karim , Krogh-Madsen Rikke , Mittendorfer Bettina , Zierath Juleen R. , Pedersen Bente K.
Published: March 2007

Most lifestyle-related chronic diseases are characterized by low-grade systemic inflammation and insulin resistance. Excessive tumor necrosis factor-α (TNF-α) concentrations have been implicated in the development of insulin resistance, but direct evidence in humans is lacking. Here, we demonstrate that TNF-α infusion in healthy humans induces insulin resistance in skeletal muscle, without effect on endogenous glucose production, as estimated by a combined euglycemic insulin clamp and stable isotope tracer method. TNF-α directly impairs glucose uptake and metabolism by altering insulin signal transduction. TNF-α infusion increases phosphorylation of p70 S6 kinase, extracellular signal–regulated kinase-1/2, and c-Jun NH2-terminal kinase, concomitant with increased serine and reduced tyrosine phosphorylation of insulin receptor substrate-1. These signaling effects are associated with impaired phosphorylation of Akt substrate 160, the most proximal step identified in the canonical insulin signaling cascade regulating GLUT4 translocation and glucose uptake. Thus, excessive concentrations of TNF-α negatively regulate insulin signaling and whole-body glucose uptake in humans. Our results provide a molecular link between low-grade systemic inflammation and the metabolic syndrome.
Effects of Insulin on Methionine and Homocysteine Kinetics in Type 2 Diabetes With Nephropathy
Authors: Tessari Paolo , Coracina Anna , Kiwanuka Edward , Vedovato Monica , Vettore Monica , Valerio Anna , Zaramella Michela , Garibotto Giacomo
Published: March 2007

Although hyperhomocysteinemia, an independent cardiovascular risk factor, is common in type 2 diabetes with nephropathy, the mechanism(s) of this alteration is not known. In healthy humans, hyperinsulinemia increases methionine transmethylation, homocysteine transsulfuration, and clearance. No such data exist in type 2 diabetes either in the fasting state or in response to hyperinsulinemia. To this purpose, seven male type 2 diabetic patients with albuminuria (1.2 ± 0.4 g/day, three with mild to moderate renal insufficiency) and seven matched control subjects were infused for 6 h with l-[methyl-2H3, 1-13C]methionine. Methionine flux, transmethylation, and disposal into proteins as well as homocysteine remethylation, transsulfuration, and clearance were determined before and after euglycemic hyperinsulinemia (∼1,000 pmol/l). In type 2 diabetic subjects, homocysteine concentration was twofold greater (P < 0.01) and methionine transmethylation and homocysteine clearance lower (from ∼15 to >50% and from ∼40 to >100%, respectively; P < 0.05) than in control subjects. The insulin-induced increments of methionine transmethylation, homocysteine transsulfuration, and clearance were markedly reduced in type 2 diabetic subjects (by more than threefold, P < 0.05 or less vs. control subjects). In contrast, methionine methyl and carbon flux were not increased in the patients. In conclusion, pathways of homocysteine disposal are impaired in type 2 diabetes with nephropathy, both in postabsorptive and insulin-stimulated states, possibly accounting for the hyperhomocysteinemia of this condition.
Statins Ameliorate Endothelial Barrier Permeability Changes in the Cerebral Tissue of Streptozotocin-Induced Diabetic Rats
Authors: Mooradian Arshag D. , Haas Michael J. , Batejko Oksana , Hovsepyan Meri , Feman Stephen S.
Published: March 2007

Statins may have favorable effects on endothelial barrier function. The effect of rosuvastatin and simvastatin therapy (10 mg/kg) for 5 weeks on blood-brain barrier (BBB), blood-retinal barrier (BRB), and cardiac muscle permeability of streptozotocin-induced diabetic rats was studied. The size-selective permeability of different vascular beds to a group of fluorescein isothiocyanate dextrans of varying molecular weights was measured. The volume of distribution of 250-, 70-, and 40-kDa dextrans in the cerebral tissue of diabetic rats were significantly increased. The volume of distribution of these dextrans in cerebral tissue was normalized by both statins. Diabetes did not significantly alter the BRB, but both statins decreased the volume of distribution of 70- and 40-kDa dextrans in the retina. The volume of distribution of 40 kDa in cardiac muscle was increased in diabetes, and this change was prevented with statin treatment. Treatment with rosuvastatin and mevalonate (150 mg/kg in drinking water for 5 weeks) did not alter the volume of distribution measurements. We concluded that 1) diabetes in rats is associated with significant changes in the BBB permeability; 2) statin treatment improves the endothelial barrier function in cerebral tissue, retina, and cardiac muscle; and 3) this statin effect could not be attributed to HMGCoA reductase inhibition.
A −30G>A Polymorphism of the β-Cell–Specific Glucokinase Promoter Associates With Hyperglycemia in the General Population of Whites
Authors: Rose Christian S. , Ek Jakob , Urhammer Søren A. , Glümer Charlotte , Borch-Johnsen Knut , Jørgensen Torben , Pedersen Oluf , Hansen Torben
Published: March 2007

A graded relationship has been reported between fasting and postprandial plasma glucose levels and the subsequent risk of cardiovascular morbidity and mortality. We hypothesized that the GCK −30G>A promoter polymorphism is associated with elevated glycemia in the middle-aged general population of whites, as well as with features of the World Health Organization (WHO)-defined metabolic syndrome. The GCK −30G>A polymorphism was genotyped in the population-based Inter99 study cohort (5,965 subjects) and in 332 nondiabetic subjects and 1,063 patients with type 2 diabetes. In the Inter99 cohort, the GCK −30A allele was associated with increased fasting (P < 0.001) and post–oral glucose tolerance test (OGTT) plasma glucose levels (P < 0.001), and in the same cohort, the GCK −30A allele was more frequent among 1,325 subjects with the metabolic syndrome than among 1,679 subjects without any components of the metabolic syndrome (P = 0.002). Moreover, the GCK −30A allele frequency was higher among 2,587 subjects with impaired glucose regulation (IGR) than among 4,773 glucose-tolerant subjects (17.3% [95% CI 16.2–18.3] vs. 15.0% [14.3–15.7], P < 0.001, odds ratio GG vs. GA 1.21 [1.08–1.36], GG vs. AA 1.62 [1.17–2.24]). In conclusion, the GCK −30G>A polymorphism associates with elevated fasting and post-OGTT glycemia in the middle-aged general population of whites, as well as with IGR and other features of the WHO-defined metabolic syndrome.
Association of Melanin-Concentrating Hormone Receptor 1 5′ Polymorphism With Early-Onset Extreme Obesity
Authors: Bell Christopher G. , Meyre David , Samson Chantal , Boyle Cliona , Lecoeur Cécile , Tauber Maïte , Jouret Béatrice , Jaquet Delphine , Levy-Marchal Claire , Charles Marie Aline , Weill Jacques , Gibson Fernando , Mein Charles A. , Froguel Philippe , Walley Andrew J.
Published: March 2007

Murine models have been highly effective in identifying the monogenic forms of human obesity discovered to date. Melanin-concentrating hormone receptor 1 (MCHR1) has been shown to be significant in the downstream orexigenic activity of the leptin-melanocortin pathway by such models. In this study, the human MCHR1 gene was extensively characterized by sequencing 3.5 kb of coding, untranslated and intronic regions plus 1 kb of putative promoter region in 180 morbidly obese adults and 87 morbidly obese children, a total of >2.4 Mb of sequencing. Thirty-nine single nucleotide polymorphisms (SNPs) were found, seven of which encode an amino acid change. One mutation, R248Q, appeared to cosegregate with the obesity trait in one pedigree but was also found to be a rare polymorphism in control samples. To investigate the possible polygenic role of MCHR1, the six common SNPs (minor allele frequency >5%) found in the sequenced regions were then screened in 557 morbidly obese adults, 552 obese children, and 1,195 nonobese nondiabetic control subjects. The plausible promoter SNP, rs133068, was found to be associated with protection against obesity in obese children only (allele frequency P = 0.006 and genotype frequency P = 0.004). Most significant results were found when using a dominant model (P = 0.001, odds ratio 0.695 [95% CI 0.560–0.863]). However, similar associations were found when both adults and children were analyzed together (P = 0.006, 0.783 [0.658–0.930]), suggesting that severe forms of obesity with early onset may be associated with SNPs in MCHR1.
Inositol (1,4,5)-Trisphosphate Dynamics and Intracellular Calcium Oscillations in Pancreatic β-Cells
Authors: Tamarina Natalia A. , Kuznetsov Andrey , Rhodes Christopher J. , Bindokas Vytautas P. , Philipson Louis H.
Published: March 2007

Glucose-stimulated insulin secretion is associated with transients of intracellular calcium concentration ([Ca2+]i) in the pancreatic β-cell. We tested the hypothesis that inositol (1,4,5)-trisphosphate [Ins(1,4,5)P3] [Ca2+]i release is incorporated in glucose-induced [Ca2+]i oscillations in mouse islets and MIN6 cells. We found that depletion of intracellular Ca2+ stores with thapsigargin increased the oscillation frequency by twofold and inhibited the slow recovery phase of [Ca2+]i oscillations. We employed a pleckstrin homology domain–containing fluorescent biosensor, phospholipase C∂ pleckstrin homology domain–enhanced green fluorescent protein, to visualize Ins(1,4,5)P3 dynamics in insulin-secreting MIN6 cells and mouse islets in real time using a video-rate confocal system. In both types of cells, stimulation with carbamoylcholine (CCh) and depolarization with KCl results in an increase in Ins(1,4,5)P3 accumulation in the cytoplasm. When stimulated with glucose, the Ins(1,4,5)P3 concentration in the cytoplasm oscillates in parallel with oscillations of [Ca2+]i. Maximal accumulation of Ins(1,4,5)P3 in these oscillations coincides with the peak of [Ca2+]i and tracks changes in frequencies induced by the voltage-gated K+ channel blockade. We show that Ins(1,4,5)P3 release in insulin-secreting cells can be stimulated by depolarization-induced Ca2+ flux. We conclude that Ins(1,4,5)P3 concentration oscillates in parallel with [Ca2+]i in response to glucose stimulation, but it is not the driving force for [Ca2+]i oscillations.
Glycation and Carboxymethyllysine Levels in Skin Collagen Predict the Risk of Future 10-Year Progression of Diabetic Retinopathy and Nephropathy in the Diabetes Control and Complications Trial and Epidemiology of Diabetes Interventions and Complications Participants With Type 1 Diabetes
Authors: Genuth Saul , Sun Wanjie , Cleary Patricia , Sell David R. , Dahms William , Malone John , Sivitz William , Monnier Vincent M. ,   for the DCCT Skin Collagen Ancillary Study Group
Published: March 2007

Several mechanistic pathways linking hyperglycemia to diabetes complications, including glycation of proteins and formation of advanced glycation end products (AGEs), have been proposed. We investigated the hypothesis that skin collagen glycation and AGEs predict the risk of progression of microvascular disease. We measured glycation products in the skin collagen of 211 Diabetes Control and Complications Trial (DCCT) volunteers in 1992 who continued to be followed in the Epidemiology of Diabetes Interventions and Complications study for 10 years. We determined whether the earlier measurements of glycated collagen and AGE levels correlated with the risk of progression of retinopathy and nephropathy from the end of the DCCT to 10 years later. In multivariate analyses, the combination of furosine (glycated collagen) and carboxymethyllysine (CML) predicted the progression of retinopathy (χ2 = 59.4, P < 0.0001) and nephropathy (χ2 = 18.2, P = 0.0001), even after adjustment for mean HbA1c (A1C) (χ2 = 32.7, P < 0.0001 for retinopathy) and (χ2 = 12.8, P = 0.0016 for nephropathy). The predictive effect of A1C vanished after adjustment for furosine and CML (χ2 = 0.0002, P = 0.987 for retinopathy and χ2 = 0.0002, P = 0.964 for nephropathy). Furosine explained more of the variation in the 10-year progression of retinopathy and nephropathy than did CML. These results strengthen the role of glycation of proteins and AGE formation in the pathogenesis of retinopathy and nephropathy. Glycation and subsequent AGE formation may explain the risk of these complications associated with prior A1C and provide a rational basis for the phenomenon of “metabolic memory” in the pathogenesis of these diabetes complications.
Partial Gene Deletion of Heart-Type Fatty Acid–Binding Protein Limits the Severity of Dietary-Induced Insulin Resistance
Authors: Shearer Jane , Fueger Patrick T. , Bracy Deanna P. , Wasserman David H. , Rottman Jeffrey N.
Published: March 2007

The aim of this study was to determine the contribution of heart-type fatty acid–binding protein (H-FABP) to glucose and long-chain fatty acid (LCFA) utilization in dietary-induced insulin resistance. We tested the hypothesis that H-FABP facilitates increases in LCFA flux present in glucose-intolerant states and that a partial reduction in the amount of this protein would compensate for all or part of the impairment. Transgenic H-FABP heterozygotes (HET) and wild-type (WT) littermates were studied following chow diet (CHD) or high-fat diet (HFD) for 12 weeks. Catheters were surgically implanted in the carotid artery and jugular vein for sampling and infusions, respectively. Following 5 days of recovery, mice received either a saline infusion or underwent a euglycemic insulin clamp (4 mU · kg–1 · min–1) for 120 min. At 90 min, a bolus of 2-deoxyglucose and [125I]-15-(ρ-iodophenyl)-3-R,S-methylpentadecanoic acid were administered to obtain indexes of glucose and LCFA utilization. At 120 min, skeletal muscles were excised for tracer determination. All HFD mice were obese and hyperinsulinemic; however, only HFD-WT mice were hyperglycemic. Glucose infusion rates during insulin clamps were 49 ± 4, 59 ± 4, 16 ± 4, and 33 ± 4 mg · kg–1 · min–1 for CHD-WT, CHD-HET, HFD-WT, and HFD-HET mice, respectively, showing that HET limited the severity of whole-body insulin resistance with HFD. Insulin-stimulated muscle glucose utilization was attenuated in HFD-WT but unaffected in HFD-HET mice. Conversely, rates of LCFA clearance were increased with HFD feeding in HFD-WT but not in HFD-HET mice. In conclusion, a partial reduction in H-FABP protein normalizes fasting glucose levels and improves whole-body insulin sensitivity in HFD-fed mice despite obesity.
The Stimulatory Effect of Globular Adiponectin on Insulin-Stimulated Glucose Uptake and Fatty Acid Oxidation Is Impaired in Skeletal Muscle From Obese Subjects
Authors: Bruce Clinton R. , Mertz Valerie A. , Heigenhauser George J.F. , Dyck David J.
Published: March 2007

Adiponectin is an adipose-derived hormone that plays an important role in regulating insulin sensitivity in rodents. However, little is known regarding the effect of adiponectin on metabolism in human skeletal muscle. Therefore, we examined whether the globular head of adiponectin, gAcrp30, acutely activates fatty acid oxidation and glucose uptake in isolated human skeletal muscle. Furthermore, we aimed to determine whether these effects would differ in muscle from lean versus obese individuals. Treatment with gAcrp30 (2.5 μg/ml) increased fatty acid oxidation in lean muscle (70%, P < 0.0001) and to a lesser extent in obese muscle (30%, P < 0.01). In the absence of insulin, gAcrp30 increased glucose uptake 37% in lean (P < 0.05) and 33% in obese muscle (P < 0.05). Combined exposure of insulin and gAcrp30 demonstrated an additive effect on glucose uptake in lean and obese individuals, but this effect was reduced by 50% in obese muscle (P < 0.05). These metabolic effects were attributable to an increase in AMP-activated protein kinase-α1 (AMPKα1) and AMPKα2 activity. However, in obese muscle the activation of AMPKα2 by gAcrp30 was blunted. This study provides evidence that gAcrp30 plays a role in regulating fatty acid and glucose metabolism in human skeletal muscle. However, the effects are blunted in obesity, indicating the possible development of adiponectin resistance.
Impaired Generation of Reactive Oxygen Species in Leprechaunism Through Downregulation of Nox4
Authors: Park Hye Sun , Jin Dong Kyu , Shin Sang Min , Jang Mi Kyung , Longo Nicholas , Park Ji Won , Bae Duk Soo , Bae Yun Soo
Published: March 2007

Leprechaunism features a clinical constellation characterized by extreme insulin resistance, growth retardation, and several distinct developmental abnormalities. One puzzling observation about leprechaunism is that mutations in the insulin receptor gene frequently associated with this syndrome cannot account for the aberrant responses of cultured cells to other growth factors. Here we report that the generation of reactive oxygen species (ROS) is impaired in cells from leprechaunism patients, thus shedding new light on this issue. Stimulation of patients’ skin fibroblast cells with platelet-derived growth factor (PDGF) resulted in a lower-level tyrosine phosphorylation of cytosolic proteins compared with that seen in normal cells. In addition, consistent with the hypothesis that ROS mediate the level of tyrosine phosphorylation of cytosolic proteins through inactivation of protein tyrosine phosphatases (PTPases), patient fibroblast cells showed a significantly higher phosphatase activity than normal cells. We further showed that the lower-level tyrosine phosphorylation in response to growth factors results from the downregulation of an NADPH oxidase, Nox4, which in turn results in the reduction of ROS generation. Ectopic expression of Nox4 in the patient fibroblast cells consistently restored PDGF-induced ROS production and regulation of PTPase activities. Taken together, these data provide insight into the mechanisms through which growth retardation is associated with leprechaunism syndrome.
Central Leptin Acutely Reverses Diet-Induced Hepatic Insulin Resistance
Authors: Pocai Alessandro , Morgan Kimyata , Buettner Christoph , Gutierrez-Juarez Roger , Obici Silvana , Rossetti Luciano
Published: March 2007

Voluntary overfeeding rapidly induces resistance to the effects of systemic insulin and leptin on liver glucose metabolism. To examine whether central administration of recombinant leptin can restore leptin and insulin action on liver glucose fluxes, we infused leptin in the third cerebral ventricle of conscious overfed rats during pancreatic-insulin clamp studies. The effect of leptin on the phosphorylation of the signal transducer and activator of transcription-3 in the arcuate nuclei of the hypothalamus was similar in animals fed a regular diet or a high-fat diet for 3 days. The infusion of leptin in the third cerebral ventricle markedly inhibited glucose production in rats fed a high-fat diet mainly by decreasing glycogenolysis. The inhibition of glycogenolysis was sufficient to normalize glucose production and was accompanied by leptin-induced decreases in the hepatic expression of glucose-6-phosphatase and phosphoenolpyruvate carboxykinase. Thus central administration of leptin rescues the hepatic insulin resistance induced by short-term hyperphagia.
Islet Cell Hormonal Responses to Hypoglycemia After Human Islet Transplantation for Type 1 Diabetes
Authors: Rickels Michael R. , Schutta Mark H. , Mueller Rebecca , Markmann James F. , Barker Clyde F. , Naji Ali , Teff Karen L.
Published: March 2007

Islet transplantation can eliminate severe hypoglycemic episodes in patients with type 1 diabetes; however, whether intrahepatic islets respond appropriately to hypoglycemia after transplantation has not been fully studied. We evaluated six islet transplant recipients, six type 1 diabetic subjects, and seven nondiabetic control subjects using a stepped hyperinsulinemic-hypoglycemic clamp. Also, three islet transplant recipients and the seven control subjects underwent a paired hyperinsulinemic-euglycemic clamp. In response to hypoglycemia, C-peptide was similarly suppressed in islet transplant recipients and control subjects and was not detectable in type 1 diabetic subjects. Glucagon was significantly more suppressed in type 1 diabetic subjects than in islet transplant recipients (P < 0.01), although the glucagon in islet transplant recipients failed to activate as in the control subjects (P < 0.01). Pancreatic polypeptide failed to activate in both type 1 diabetic subjects and islet transplant recipients compared with control subjects (P < 0.01). In islet transplant recipients, glucagon was suppressed normally by hyperinsulinemia during the euglycemic clamp and was significantly greater during the paired hypoglycemic clamp (P < 0.01). These results suggest that after islet transplantation and in response to insulin-induced hypoglycemia, endogenous insulin secretion is appropriately suppressed and glucagon secretion may be partially restored.
Homocysteine Metabolism in ZDF (Type 2) Diabetic Rats
Authors: Wijekoon Enoka P. , Hall Beatrice , Ratnam Shobhitha , Brosnan Margaret E. , Zeisel Steven H. , Brosnan John T.
Published: March 2007

Mild hyperhomocysteinemia is a risk factor for many diseases, including cardiovascular disease. We determined the effects of insulin resistance and of type 2 diabetes on homocysteine (Hcy) metabolism using Zucker diabetic fatty rats (ZDF/Gmi fa/fa and ZDF/Gmi fa/?). Plasma total Hcy was reduced in ZDF fa/fa rats by 24% in the pre-diabetic insulin-resistant stage, while in the frank diabetic stage there was a 59% reduction. Hepatic activities of several enzymes that play a role in the removal of Hcy:cystathionine β-synthase (CBS), cystathionine γ-lyase, and betaine:Hcy methyltransferase (BHMT) were increased as was methionine adenosyltransferase. CBS and BHMT mRNA levels and the hepatic level of S-adenosylmethionine were also increased in the ZDF fa/fa rats. Studies with primary hepatocytes showed that Hcy export and the transsulfuration flux in cells from ZDF fa/fa rats were particularly sensitive to betaine. Interestingly, liver betaine concentration was found to be significantly lower in the ZDf fa/fa rats at both 5 and 11 weeks. These results emphasize the importance of betaine metabolism in determining plasma Hcy levels in type 2 diabetes.
Celiac-Superior Mesenteric Ganglionectomy, but Not Vagotomy, Suppresses the Sympathoadrenal Response to Insulin-Induced Hypoglycemia
Authors: Fujita Satoshi , Donovan Casey M.
Published: March 2007

Afferent innervation of the portal vein has been shown to be critical in hypoglycemic detection, but the neural pathway by which these afferents ascend remains unknown. To ascertain the role of vagal afferents versus spinal afferents in hypoglycemic detection, the catecholamine response to hypoglycemia was assessed in male Wistar rats undergoing hepatic vagotomy (HV), total subdiaphragmatic vagotomy (TSV), or celiac-superior mesenteric ganglionectomy (CSMG). After recovering from the surgery, the animals were exposed to a hyperinsulinemic-hypoglycemic clamp, with glucose infused peripherally via the jugular vein. In all animals, systemic hypoglycemia (2.64 ± 0.03 mmol/l) was induced via jugular vein insulin infusion (25 mU · kg−1 · min−1). No significant differences were observed among the groups with respect to arterial glucose or insulin concentration. When hypoglycemia was induced in sham-operated control animals, epinephrine was observed to rise from a basal value of 0.84 ± 0.10 to 25.18 ± 1.24 nmol/l. Neither HV nor TSV had any significant impact on the epinephrine response to hypoglycemia. In contrast, CSMG animals demonstrated a significant suppression in the epinephrine response to whole-body hypoglycemia (11.25 ± 1.21 vs. 22.32 ± 0.86 nmol/l in CSMG vs. controls; P < 0.05). The norepinephrine response for controls, 2.00 ± 0.22 at basal and rising to 8.95 ± 0.20 nmol/l in hypoglycemia, was not significantly different from that of the HV and TSV animals. As with epinephrine, the norepinephrine response to hypoglycemia was significantly suppressed in CSMG compared with control animals (4.72 ± 0.48 vs. 7.15 ± 0.76 nmol/l; P < 0.05). These findings are consistent with the idea that hypoglycemic detection at the portal vein is mediated by spinal, and not vagal, glucose-sensitive afferents.
Oxidative-Nitrosative Stress and Poly(ADP-Ribose) Polymerase (PARP) Activation in Experimental Diabetic Neuropathy
Authors: Obrosova Irina G. , Drel Viktor R. , Pacher Pal , Ilnytska Olga , Wang Zhong Q. , Stevens Martin J. , Yorek Mark A.
Published: March 2007

Poly(ADP-ribose) polymerase (PARP) activation, an important factor in the pathogenesis of diabetes complications, is considered a downstream effector of oxidative-nitrosative stress. However, some recent findings suggest that it is not necessarily the case and that PARP activation may precede and contribute to free radical and oxidant-induced injury. This study evaluated the effect of PARP inhibition on oxidative-nitrosative stress in diabetic peripheral nerve, vasa nervorum, aorta, and high glucose–exposed human Schwann cells. In vivo experiments were performed in control rats and streptozocin (STZ)-induced diabetic rats treated with and without the PARP inhibitor 3-aminobenzamide (ABA) (30 mg · kg−1 · day−1 i.p. for 2 weeks after 2 weeks of untreated diabetes). Human Schwann cells (HSC) (passages 7–10; ScienCell Research Labs) were cultured in 5.5 or 30 mmol/l glucose with and without 5 mmol/l ABA. Diabetes-induced increase in peripheral nerve nitrotyrosine immunoreactivity, epineurial vessel superoxide and nitrotyrosine immunoreactivities, and aortic superoxide production was reduced by ABA. PARP-1 (Western blot analysis) was abundantly expressed in HSC, and its expression was not affected by high glucose or ABA treatment. High-glucose–induced superoxide production and overexpression of nitrosylated and poly(ADP-ribosyl)ated protein, chemically reduced amino acid-(4)-hydroxynonenal adducts, and inducible nitric oxide synthase were decreased by ABA. We concluded that PARP activation contributes to superoxide anion radical and peroxynitrite formation in peripheral nerve, vasa nervorum, and aorta of STZ-induced diabetic rats and high- glucose–exposed HSC. The relations between oxidative-nitrosative stress and PARP activation in diabetes are bi- rather than unidirectional, and PARP activation cannot only result from but also lead to free radical and oxidant generation.
Free Fatty Acids Produce Insulin Resistance and Activate the Proinflammatory Nuclear Factor-κB Pathway in Rat Liver
Authors: Boden Guenther , She Pengxiang , Mozzoli Maria , Cheung Peter , Gumireddy Kiranmai , Reddy Prekumar , Xiang Xiaqin , Luo Zhijan , Ruderman Neil
Published: March 2007

To study mechanisms by which free fatty acids (FFAs) cause hepatic insulin resistance, we have used euglycemic-hyperinsulinemic clamping with and without infusion of lipid/heparin (to raise or to lower plasma FFAs) in alert male rats. FFA-induced hepatic insulin resistance was associated with increased hepatic diacylglycerol content (+210%), increased activities of two serine/threonine kinases (protein kinase C-δ and inhibitor of κB [IκB] kinase-β), increased activation of the proinflammatory nuclear factor-κB (NF-κB) pathway (IκB kinase-β, +640%; IκB-α, −54%; and NF-κB, +73%), and increased expression of inflammatory cytokines (tumor necrosis factor-α, +1,700% and interleukin-1β, +440%) and plasma levels of monocyte chemoattractant protein-1 (+220%). We conclude that FFAs caused hepatic insulin resistance, which can produce overproduction of glucose and hyperglycemia, and initiated inflammatory processes in the liver that could potentially result in the development of steatohepatitis.
Glucose Metabolism in Mice Lacking Muscle Glycogen Synthase
Authors: Pederson Bartholomew A. , Schroeder Jill M. , Parker Gretchen E. , Smith Micah W. , DePaoli-Roach Anna A. , Roach Peter J.
Published: March 2007

Glycogen is an important component of whole-body glucose metabolism. MGSKO mice lack skeletal muscle glycogen due to disruption of the GYS1 gene, which encodes muscle glycogen synthase. MGSKO mice were 5–10% smaller than wild-type littermates with less body fat. They have more oxidative muscle fibers and, based on the activation state of AMP-activated protein kinase, more capacity to oxidize fatty acids. Blood glucose in fed and fasted MGSKO mice was comparable to wild-type littermates. Serum insulin was lower in fed but not in fasted MGSKO animals. In a glucose tolerance test, MGSKO mice disposed of glucose more effectively than wild-type animals and had a more sustained elevation of serum insulin. This result was not explained by increased conversion to serum lactate or by enhanced storage of glucose in the liver. However, glucose infusion rate in a euglycemic-hyperinsulinemic clamp was normal in MGSKO mice despite diminished muscle glucose uptake. During the clamp, MGSKO animals accumulated significantly higher levels of liver glycogen as compared with wild-type littermates. Although disruption of the GYS1 gene negatively affects muscle glucose uptake, overall glucose tolerance is actually improved, possibly because of a role for GYS1 in tissues other than muscle.
Plasma Free Fatty Acids and Peroxisome Proliferator–Activated Receptor α in the Control of Myocardial Uncoupling Protein Levels
Authors: Murray Andrew J. , Panagia Marcello , Hauton David , Gibbons Geoffrey F. , Clarke Kieran
Published: March 2007

Diabetic patients have abnormal cardiac energy metabolism associated with high plasma free fatty acid (FFA) concentrations. We investigated whether high plasma FFAs increase mitochondrial uncoupling protein (UCP) levels in the mouse heart by activating the nuclear transcription factor peroxisome proliferator–activated receptor (PPAR)α. We used Western blotting to measure UCP protein levels in isolated cardiac mitochondria from PPARα−/− and diabetic mice. Cardiac UCP2 and UCP3 were significantly lower in the PPARα−/− mouse than in the wild type. Treatment with the PPARα-specific agonist, WY-14,643, increased cardiac UCP2 and UCP3 levels in wild-type mice but did not alter UCP levels in PPARα−/− mice. Inhibition of β-oxidation with etomoxir increased cardiac UCP2 and UCP3 levels in wild-type mice and UCP2 levels in PPARα−/− mice but did not alter UCP3 levels in PPARα−/− mice. Streptozotocin treatment, which increased circulating FFAs by 91%, did not alter cardiac UCP2 levels in wild-type or PPARα−/− mice but increased UCP3 levels in wild-type, and not in PPARα−/−, mice. The diabetic db/db mouse had 50% higher plasma FFA concentrations and elevated cardiac UCP2 and UCP3 protein levels. We conclude that high plasma FFAs activated PPARα to increase cardiac UCP3 levels, but cardiac UCP2 levels changed via PPARα-dependent and -independent mechanisms.
Decreased Fat Mass in Interleukin-1 Receptor Antagonist–Deficient Mice
Authors: Somm Emmanuel , Henrichot Elvire , Pernin Agnès , Juge-Aubry Cristiana E. , Muzzin Patrick , Dayer Jean-Michel , Nicklin Martin J.H. , Meier Christoph A.
Published: March 2007

Interleukin (IL)-1 is a regulator of inflammation but is also implicated in the control of energy homeostasis. Because the soluble IL-1 receptor antagonist (IL-1Ra) is markedly increased in the serum of obese patients and is overexpressed in white adipose tissue in obesity, we studied the metabolic consequences of genetic IL-1Ra ablation in mice. We have shown that IL-1Ra−/− mice have a lean phenotype due to decreased fat mass, related to a defect in adipogenesis and increased energy expenditure. The adipocytes were smaller in these animals, and the expression of genes involved in adipogenesis was reduced. Energy expenditure as measured by indirect calorimetry was elevated, and weight loss in response to a 24-h fast was increased in IL-1Ra−/− animals compared with wild-type mice. Lipid oxidation of IL-1Ra−/− mice was higher during the light period, reflecting their reduction in diurnal food intake. Interestingly, IL-1Ra−/− and IL-1Ra+/− mice presented an attenuation in high-fat diet–induced caloric hyperphagia, indicating a better adaptation to hypercaloric alimentation, which is in line with the role of IL-1Ra as a mediator of leptin resistance. Taken together, we show that IL-1Ra is an important regulator of adipogenesis, food intake, and energy expenditure.
Insulin Resistance Syndrome in Subjects With Mutated RING Finger Protein TRIM37
Authors: Karlberg Niklas , Jalanko Hannu , Kallijärvi Jukka , Lehesjoki Anna-Elina , Lipsanen-Nyman Marita
Published: March 2007

We evaluated the glucose and lipid metabolism in 65 patients (aged 1.1–55 years) with mulibrey (muscle-liver-brain-eye) nanism (MUL), which is a monogenic disorder with prenatal-onset growth failure and typical clinical characteristics. MUL is caused by mutations in the TRIM37 gene, encoding a peroxisomal protein (TRIM37) with E3 ubiquitin-ligase activity. The subjects underwent clinical evaluation, abdominal ultrasonography, and laboratory measurements, including a 3-h oral glucose tolerance test. The results showed a dramatic change in glucose and lipid metabolism with age in MUL subjects. While the children had low fasting glucose and insulin levels, 90% of the adults had high fasting and postload insulin values (up to 1,450 mU/l). A 10-fold decrease in the fasting glucose-to-insulin ratio and a 4-fold decrease in whole-body insulin sensitivity index were observed. Insulin resistance, fatty liver, high serum leptin, hypertension, and acantosis nigricans were already evident in many slim prepubertal children. Half of the adults had type 2 diabetes, and an additional 42% showed impaired glucose tolerance. Seventy percent fulfilled the National Cholesterol Education Program criteria for metabolic syndrome. The peroxisomal targeting and the functional link of TRIM37 to the ubiquitin-proteosome pathway may provide novel clues to the development of metabolic syndrome.