bluuebunny/crossref_metadata_2025 huggingface dataset:
Diabetes Due to a Progressive Defect in β-Cell Mass in Rats Transgenic for Human Islet Amyloid Polypeptide (HIP Rat)
Authors: Butler Alexandra E. , Jang Jennifer , Gurlo Tatyana , Carty Maynard D. , Soeller Walter C. , Butler Peter C.
Published: March 2007

The islet in type 2 diabetes is characterized by a deficit in β-cell mass, increased β-cell apoptosis, and impaired insulin secretion. Also, islets in type 2 diabetes often contain deposits of islet amyloid derived from islet amyloid polypeptide (IAPP), a 37–amino acid protein cosecreted with insulin by β-cells. Several lines of evidence suggest that proteins with a capacity to develop amyloid fibrils may also form small toxic oligomers that can initiate apoptosis. The amino acid sequence of IAPP in rats and mice is identical and differs from that in humans by substitution of proline residues in the amyloidogenic sequence so that the protein no longer forms amyloid fibrils or is cytotoxic. In the present study, we report a novel rat model for type 2 diabetes: rats transgenic for human IAPP (the HIP rat). HIP rats develop diabetes between 5 and 10 months of age, characterized by an ∼60% deficit in β-cell mass that is due to an increased frequency of β-cell apoptosis. HIP rats develop islet amyloid, but the extent of amyloid was not related to the frequency of β-cell apoptosis (r = 0.10, P = 0.65), whereas the fasting blood glucose was (r = 0.77, P < 0.001). The frequency of β-cell apoptosis was related to the frequency of β-cell replication (r = 0.97, P < 0.001) in support of the hypothesis that replicating cells are more vulnerable to apoptosis than nondividing cells. The HIP rat provides additional evidence in support of the potential role of IAPP oligomer formation toward the increased frequency of apoptosis in type 2 diabetes, a process that appears to be compounded by glucose toxicity when hyperglycemia supervenes.
Regulation of Rat Pancreatic CCKB Receptor and Somatostatin Expression by Insulin
Authors: Julien Sophie , Laine Jean , Morisset Jean
Published: March 2007

The cholecystokinin B receptor (CCKBR) is localized on pancreatic endocrine somatostatin δ-cells. Pancreatic somatostatin content was increased in diabetic rats. The mechanisms involved in this phenomenon are unknown, and we believe insulin is involved. In this study, four groups of rats were used: controls, streptozotocin-induced diabetic, streptozotocin-induced diabetic with insulin, and streptozotocin-induced diabetic with insulin and its cessation. Rats were killed after 7–28 days of treatment for diabetes, and somatostatin mRNA expression and pancreatic somatostatin content, CCKBR mRNA and protein expression evaluation in total pancreas and purified islets, and the cellular localization of somatostatin and CCKBR in islets was measured. Data indicate that diabetes is established after 7 days, is controlled by insulin, and reappears after treatment cessation. Pancreatic somatostatin mRNA expression and somatostatin content were increased during diabetes, normalized during insulin treatment, and reaugmented after treatment cessation. Gland and islet CCKBR mRNA and protein almost disappeared during diabetes; CCKB mRNA reappeared in response to insulin, but the protein did not. Confocal microscopy confirmed data obtained on somatostatin and CCKBR as established biochemically in the course of the treatments. In conclusion, these data strongly suggest that insulin can negatively control pancreatic somatostatin mRNA and hormone content and positively control CCKBR mRNA; the CCKBR protein appears to be delayed.
Influence of Autonomic Neuropathy on QTc Interval Lengthening During Hypoglycemia in Type 1 Diabetes
Authors: Lee Stuart P. , Yeoh Lishan , Harris Nigel D. , Davies Catherine M. , Robinson Robert T. , Leathard Andrew , Newman Christopher , Macdonald Ian A. , Heller Simon R.
Published: March 2007

Hypoglycemia produces electrocardiographic QTc lengthening, a predictor of arrhythmia risk and sudden death. This results from both sympatho-adrenal activation and a lowered serum potassium. It has been suggested that cardiac autonomic neuropathy (CAN) might indicate those who are at particular risk. We tested this hypothesis in 28 adults with type 1 diabetes and 8 nondiabetic control subjects. After standard tests of autonomic function and baroreflex sensitivity (BRS) measurement, diabetic participants were divided into three groups: 1) CAN− with normal BRS (BRS+; n = 10), 2) CAN− with impaired BRS (BRS−; n = 9), and 3) CAN+ (n = 9). QTc was then measured during controlled hypoglycemia (2.5 mmol/l) using a hyperinsulinemic clamp. Mean (±SE) QTc lengthened from 377 ± 9 ms (baseline) to a maximum during hypoglycemia of 439 ± 13 ms in BRS+ subjects and from 378 ± 5 to 439 ± 10 ms in control subjects. Peak QTc tended to be lower in CAN+ (baseline, 383 ± 6; maximum, 408 ± 10) and BRS− groups (baseline, 380 ± 8; maximum, 421 ± 11; F = 1.7, P = 0.18). Peak epinephrine concentrations (nmol/l) were 3.1 ± 0.8 (BRS+), 2.6 ± 0.5 (BRS−), 1.4 ± 0.3 (CAN+), and 5.7 ± 0.8 (control subjects). These data do not indicate that those with CAN are at particular risk for abnormal cardiac repolarization during hypoglycemia. Indeed, they suggest that such patients may be relatively protected, perhaps as a result of attenuated sympatho-adrenal responses.
Molecular Alterations Underlie Nodal and Paranodal Degeneration in Type 1 Diabetic Neuropathy and Are Prevented by C-Peptide
Authors: Sima Anders A.F. , Zhang Weixian , Li Zhen-Guo , Murakawa Yuichi , Pierson Christopher R.
Published: March 2007

To explore the molecular abnormalities underlying the degeneration of the node of Ranvier, a characteristic aberration of type 1 diabetic neuropathy, we examined in type 1 BB/Wor and type 2 BBZDR/Wor rats changes in expression of key molecules that make up the nodal and paranodal apparatus of peripheral nerve. Their posttranslational modifications were examined in vitro. Their responsiveness to restored insulin action was examined in type 1 animals replenished with proinsulin C-peptide. In sciatic nerve, the expression of contactin, receptor protein tyrosine phosphatase β, and the Na+-channel β1 subunit, paranodal caspr and nodal ankyrinG was unaltered in 2-month type 1 diabetic BB/Wor rats but significantly decreased after 8 months of diabetes. These abnormalities were prevented by C-peptide administered to type 1 BB/Wor rats and did not occur in duration- and hyperglycemia-matched type 2 BBZDR/Wor rats. The expression of the α-Na+-channel subunit was unaltered. In SH-SY5Y cells, only the combination of insulin and C-peptide normalized posttranslational O-linked N-acetylglucosamine modifications and maximized serine phosphorylation of ankyrinG and p85 binding to caspr. The beneficial effects of C-peptide resulted in significant normalization of the nerve conduction deficits. These data describe for the first time the progressive molecular aberrations underlying nodal and paranodal degenerative changes in type 1 diabetic neuropathy and demonstrate that they are preventable by insulinomimetic C-peptide.
Interaction and Association Analysis of a Type 1 Diabetes Susceptibility Locus on Chromosome 5q11-q13 and the 7q32 Chromosomal Region in Scandinavian Families
Authors: Holm Pernilla , Rydlander Berit , Luthman Holger , Kockum Ingrid ,   for the European Consortium for IDDM Genome Studies
Published: March 2007

We have previously reported suggestive linkage to chromosome 5p13-q13 in type 1 diabetic families. ISL1, a transcription factor involved in pancreas development, maps to this region. Sequencing of the ISL1 gene in patients and control subjects identified seven single nucleotide polymorphisms (SNPs) and one microsatellite in noncoding regions. Four haplotypes formed by six of these SNPs and one microsatellite were associated with type 1 diabetes in Swedish families (P < 0.04). To identify possible interactions with the 5q11-q13 region, we applied pathway-restricted linkage analysis by analyzing for effects from regions encoding other transcription factors that are active during pancreas development and maintenance of insulin production. Linkage analysis allowing for interaction between 5q11-q13 and 7q32 resulted in an increase of logarithm of odds from 2.2 to 5.3. This increase was estimated to correspond to a P value <0.0016 using permutation. The transcription factor PAX4 is located at 7q32 and participates downstream of ISL1 in the transcription factor cascade critical to β-cell development. Association with type 1 diabetes was also observed using the transmission disequilibrium test for two haplotypes at the PAX4 locus (P < 0.05). We conclude that pathway-restricted linkage analysis assists in the identification of possible gene-gene interactions and that 5q11-q13 and 7q32 together constitute a significant susceptibility factor for type 1 diabetes.
Diabetes-Associated Mutations in Insulin Identify Invariant Receptor Contacts
Authors: Xu Bin , Hu Shi-Quan , Chu Ying-Chi , Wang Shuhua , Wang Run-ying , Nakagawa Satoe H. , Katsoyannis Panayotis G. , Weiss Michael A.
Published: March 2007

Mutations in human insulin cause an autosomal-dominant syndrome of diabetes and fasting hyperinsulinemia. We demonstrate by residue-specific photo cross-linking that diabetes-associated mutations occur at receptor-binding sites. The studies use para-azido-phenylalanine, introduced at five sites by total protein synthesis. Because two such sites (ValA3 and PheB24) are largely buried in crystal structures of the free hormone, their participation in receptor binding is likely to require a conformational change to expose a hidden functional surface. Our results demonstrate that this surface spans both chains of the insulin molecule and includes sites of rare human mutations that cause diabetes.
Lower Within-Subject Variability of Insulin Detemir in Comparison to NPH Insulin and Insulin Glargine in People With Type 1 Diabetes
Authors: Heise Tim , Nosek Leszek , Rønn Birgitte Biilmann , Endahl Lars , Heinemann Lutz , Kapitza Christoph , Draeger Eberhard
Published: March 2007

The aim of this randomized double-blind study was to compare the within-subject variability of the glucose-lowering effect of a novel insulin analog, insulin detemir, with that of insulin glargine and NPH insulin in people with type 1 diabetes. Fifty-four subjects (32 males and 22 females, age 38 ± 10 years [mean ± SD], BMI 24 ± 2 kg/m2, HbA1c 7.5 ± 1.2%, diabetes duration 18 ± 9 years) participated in this parallel group comparison. Each subject received four single subcutaneous doses of 0.4 units/kg of either insulin detemir (n = 18), insulin glargine (n = 16), or human NPH insulin (n = 17) under euglycemic glucose clamp conditions (target blood glucose concentration 5.5 mmol/l) on four identical study days. The pharmacodynamic (glucose infusion rates [GIRs]) and pharmacokinetic (serum concentrations of insulin detemir, human insulin, and insulin glargine) properties of the basal insulin preparations were recorded for 24 h postdosing. Insulin detemir was associated with significantly less within-subject variability than both NPH insulin and insulin glargine, as assessed by the coefficient of variation (CV) for the pharmacodynamic end points studied [GIR-AUC(0–12 h) 27% (detemir) vs. 59% (NPH) vs. 46% (glargine); GIR-AUC(0–24 h) 27 vs. 68 vs. 48%; GIRmax 23 vs. 46 vs. 36%; P < 0.001 for all comparisons]. Insulin detemir also provided less within-subject variability in the pharmacokinetic end points: maximal concentration (Cmax) 18 vs. 24 vs. 34%; INS-AUC(0–∞) 14 vs. 28 vs. 33%. The results suggest that insulin detemir has a significantly more predictable glucose-lowering effect than both NPH insulin and insulin glargine.
In Vitro–Deranged Intestinal Immune Response to Gliadin in Type 1 Diabetes
Authors: Auricchio Renata , Paparo Francesco , Maglio Maria , Franzese Adriana , Lombardi Francesca , Valerio Giuliana , Nardone Gerardo , Percopo Selvaggia , Greco Luigi , Troncone Riccardo
Published: March 2007

Dietary gluten has been associated with an increased risk of type 1 diabetes. We have evaluated inflammation and the mucosal immune response to gliadin in the jejunum of patients with type 1 diabetes. Small intestinal biopsies from 17 children with type 1 diabetes without serological markers of celiac disease and from 50 age-matched control subjects were examined by immunohistochemistry. In addition, biopsies from 12 type 1 diabetic patients and 8 control subjects were cultured with gliadin or ovalbumin peptic-tryptic digest and examined for epithelial infiltration and lamina propria T-cell activation. The density of intraepithelial CD3+ and γδ+ cells and of lamina propria CD25+ mononuclear cells was higher in jejunal biopsies from type 1 diabetic patients versus control subjects. In the patients’ biopsies cultured with peptic-tryptic gliadin, there was epithelial infiltration by CD3+ cells, a significant increase in lamina propria CD25+ and CD80+ cells and enhanced expression of lamina propria CD54 and crypt HLA-DR. No such phenomena were observed in control subjects, even those with celiac disease–associated HLA haplotypes. In conclusion, signs of mucosal inflammation were present in jejunal biopsies from type 1 diabetic patients, and organ culture studies indicate a deranged mucosal immune response to gliadin.
Targeted Disruption of the IA-2β Gene Causes Glucose Intolerance and Impairs Insulin Secretion but Does Not Prevent the Development of Diabetes in NOD Mice
Authors: Kubosaki Atsutaka , Gross Steffen , Miura Junnosuke , Saeki Keiichi , Zhu Min , Nakamura Shinichiro , Hendriks Wiljan , Notkins Abner Louis
Published: March 2007

Insulinoma-associated protein (IA)-2β, also known as phogrin, is an enzymatically inactive member of the transmembrane protein tyrosine phosphatase family and is located in dense-core secretory vesicles. In patients with type 1 diabetes, autoantibodies to IA-2β appear years before the development of clinical disease. The genomic structure and function of IA-2β, however, is not known. In the present study, we determined the genomic structure of IA-2β and found that both human and mouse IA-2β consist of 23 exons and span ∼1,000 and 800 kb, respectively. With this information, we prepared a targeting construct and inactivated the mouse IA-2β gene as demonstrated by lack of IA-2β mRNA and protein expression. The IA-2β−/− mice, in contrast to wild-type controls, showed mild glucose intolerance and impaired glucose-stimulated insulin secretion. Knockout of the IA-2β gene in NOD mice, the most widely studied animal model for human type 1 diabetes, failed to prevent the development of cyclophosphamide-induced diabetes. We conclude that IA-2β is involved in insulin secretion, but despite its importance as a major autoantigen in human type 1 diabetes, it is not required for the development of diabetes in NOD mice.
HLA-DQ–Regulated T-Cell Responses to Islet Cell Autoantigens Insulin and GAD65
Authors: Tree Timothy I.M. , Duinkerken Gaby , Willemen Sabine , de Vries René R.P. , Roep Bart O.
Published: March 2007

HLA-DQ is strongly associated with genetic predisposition to type 1 diabetes. It is assumed that HLA-DQ molecules exert their effects on the disease via the presentation of peptides from islet autoantigens to CD4+ T-cells, but little information regarding HLA-DQ–restricted, islet antigen–specific, autoreactive T-cells is available. To investigate the role of HLA-DQ in the immune response to islet autoantigens, we measured T-cell proliferation to insulin and GAD65 in the presence and absence of monoclonal antibodies that block HLA-DQ–mediated antigen presentation in recent-onset type 1 diabetic patients and their siblings. Positive proliferative T-cell responses to GAD65 were observed in 60% of type 1 diabetic patients and 52% of siblings. This proliferation was significantly reduced in the presence of anti-DQ antibody, demonstrating the presence of primed, effector HLA-DQ–restricted T-cell responses to GAD65. Positive proliferative responses to insulin were observed in 25% of type 1 diabetic patients and 10% of siblings. However, blocking HLA-DQ–restricted T-cell responses led to a significant increase in proliferation to insulin, implying the presence of primed suppressive HLA-DQ–restricted T-cell responses to insulin. These results indicate that HLA-DQ acts as a restriction element for both proliferative and suppressor cells, with the relative balance of these cells dependent on the nature of the autoantigen.
Muscarinic Stimulation of Pancreatic Insulin and Glucagon Release Is Abolished in M3 Muscarinic Acetylcholine Receptor–Deficient Mice
Authors: Duttaroy Alokesh , Zimliki Charles L. , Gautam Dinesh , Cui Yinghong , Mears David , Wess Jürgen
Published: March 2007

Pancreatic muscarinic acetylcholine receptors play an important role in stimulating insulin and glucagon secretion from islet cells. To study the potential role of the M3 muscarinic receptor subtype in cholinergic stimulation of insulin release, we initially examined the effect of the muscarinic agonist, oxotremorine-M (Oxo-M), on insulin secretion from isolated pancreatic islets prepared from wild-type (WT) and M3 receptor–deficient mice (M3+/− and M3−/− mice). At a stimulatory glucose level (16.7 mmol/l), Oxo-M strongly potentiated insulin output from islets of WT mice. Strikingly, this effect was completely abolished in islets from M3−/− mice and significantly reduced in islets from M3+/− mice. Additional in vitro studies showed that Oxo-M–mediated glucagon release was also virtually abolished in islets from M3−/− mice. Consistent with the in vitro data, in vivo studies showed that M3−/− mice displayed reduced serum insulin and plasma glucagon levels and a significantly blunted increase in serum insulin after an oral glucose load. Despite the observed impairments in insulin release, M3−/− mice showed significantly reduced blood glucose levels and even improved glucose tolerance, probably due to the reduction in plasma glucagon levels and the fact that M3−/− mice are hypophagic and lean. These findings provide important new insights into the metabolic roles of the M3 muscarinic receptor subtype.
Insulin Feedback Alters Mitochondrial Activity Through an ATP-sensitive K+ Channel–Dependent Pathway in Mouse Islets and β-Cells
Authors: Nunemaker Craig S. , Zhang Min , Satin Leslie S.
Published: March 2007

Recent work suggests that insulin may exert both positive and negative feedback directly on pancreatic β-cells. To investigate the hypothesis that insulin modulates β-cell metabolism, mouse islets and β-cell clusters were loaded with rhodamine 123 to dynamically monitor mitochondrial membrane potential (ΔΨm). Spontaneous oscillations in ΔΨm (period: 218 ± 26 s) were observed in 17 of 30 islets exposed to 11.1 mmol/l glucose. Acute insulin application (100 nmol/l) hyperpolarized ΔΨm, indicating a change in mitochondrial activity. The ATP-sensitive K+ (KATP) channel opener diazoxide or the l-type calcium channel blocker nifedipine mimicked the effect of insulin, suggesting that insulin activates KATP channels to hyperpolarize ΔΨm by inhibiting calcium influx. Treatment with forskolin, which increases endogenous insulin secretion, also mimicked the effect of exogenous insulin, suggesting physiological feedback. Pretreatment with nifedipine or the KATP inhibitor glyburide prevented insulin action, further implicating a KATP channel pathway. Together, these data suggest a feedback mechanism whereby insulin receptor activation opens KATP channels to inhibit further secretion. The resulting reduction in β-cell calcium increases the energy stored in the mitochondrial gradient that drives ATP production. Insulin feedback onto mitochondria may thus help to calibrate the energy needs of the β-cell on a minute-to-minute basis.
Single-Dose Dexamethasone Induces Whole-Body Insulin Resistance and Alters Both Cardiac Fatty Acid and Carbohydrate Metabolism
Authors: Qi Dake , Pulinilkunnil Thomas , An Ding , Ghosh Sanjoy , Abrahani Ashraf , Pospisilik John Andrew , Brownsey Roger , Wambolt Rich , Allard Michael , Rodrigues Brian
Published: March 2007

Glucocorticoids impair insulin sensitivity. Because insulin resistance is closely linked to increased incidence of cardiovascular diseases and given that metabolic abnormalities have been linked to initiation of heart failure, we examined the acute effects of dexamethasone (DEX) on rat cardiac metabolism. Although injection of DEX for 4 h was not associated with hyperinsulinemia, the euglycemic-hyperinsulinemic clamp showed a decrease in glucose infusion rate. Rates of cardiac glycolysis were unaffected, whereas the rate of glucose oxidation following DEX was significantly decreased and could be associated with augmented expression of PDK4 mRNA and protein. Myocardial glycogen content in DEX hearts increased compared with control. Similar to hypoinsulinemia induced by streptozotocin (STZ), hearts from insulin-resistant DEX animals also demonstrated enlargement of the coronary lipoprotein lipase (LPL) pool. However, unlike STZ, DEX hearts showed greater basal release of LPL and were able to maintain their high heparin-releasable LPL in vitro. This effect could be explained by the enhanced LPL mRNA expression following DEX. Our data provide evidence that in a setting of insulin resistance, an increase in LPL could facilitate increased delivery of fatty acid to the heart, leading to excessive triglyceride storage. It has not been determined whether these acute effects of DEX on cardiac metabolism can be translated into increased cardiovascular risk.
Direct Insulin Signaling of Neurons Reverses Diabetic Neuropathy
Authors: Brussee Valentine , Cunningham F. Alexander , Zochodne Douglas W.
Published: March 2007

Diabetic polyneuropathy is the most common acquired diffuse disorder of the peripheral nervous system. It is generally assumed that insulin benefits human and experimental diabetic neuropathy indirectly by lowering glucose levels. Insulin also provides potent direct support of neurons and axons, and there is a possibility that abnormalities in direct insulin signaling on peripheral neurons relate to the development of this disorder. Here we report that direct neuronal (intrathecal) delivery of low doses of insulin (0.1–0.2 IU daily), insufficient to reduce glycemia or equimolar IGF-I but not intrathecal saline or subcutaneous insulin, improved and reversed slowing of motor and sensory conduction velocity in rats rendered diabetic using streptozotocin. Moreover, insulin and IGF-I similarly reversed atrophy in myelinated sensory axons in the sural nerve. That intrathecal insulin had the capability of signaling sensory neurons was confirmed by observing that fluorescein isothiocyanate-labeled insulin given intrathecally accessed and labeled individual lumbar dorsal root ganglion neurons. Moreover, we confirmed that such neurons express the insulin receptor, as previously suggested by Sugimoto et al. Finally, we sequestered intrathecal insulin in nondiabetic rats using an anti-insulin antibody. Conduction slowing and axonal atrophy resembling the changes in diabetes were generated by anti-insulin but not by an anti-rat albumin antibody infusion. Defective direct signaling of insulin on peripheral neurons through routes that include the cerebrospinal fluid may relate to the development of diabetic peripheral neuropathy.
Remapping the Insulin Gene/IDDM2 Locus in Type 1 Diabetes
Authors: Barratt Bryan J. , Payne Felicity , Lowe Chris E. , Hermann Robert , Healy Barry C. , Harold Denise , Concannon Patrick , Gharani Neda , McCarthy Mark I. , Olavesen Mark G. , McCormack Rose , Guja Cristian , Ionescu-Tîrgovişte Constantin , Undlien Dag E. , Rønningen Kjersti S. , Gillespie Kathleen M. , Tuomilehto-Wolf Eva , Tuomilehto Jaakko , Bennett Simon T. , Clayton David G. , Cordell Heather J. , Todd John A.
Published: March 2007

Type 1 diabetes susceptibility at the IDDM2 locus was previously mapped to a variable number tandem repeat (VNTR) 5′ of the insulin gene (INS). However, the observation of associated markers outside a 4.1-kb interval, previously considered to define the limits of IDDM2 association, raised the possibility that the VNTR association might result from linkage disequilibrium (LD) with an unknown polymorphism. We therefore identified a total of 177 polymorphisms and obtained genotypes for 75 of these in up to 434 pedigrees. We found that, whereas disease susceptibility did map to within the 4.1-kb region, there were two equally likely candidates for the causal variant, −23HphI and +1140A/C, in addition to the VNTR. Further analyses in 2,960 pedigrees did not support the difference in association between VNTR lineages that had previously enabled the exclusion of these two polymorphisms. Therefore, we were unable to rule out −23HphI and +1140A/C having an etiological effect. Our mapping results using robust regression methods show how precisely a variant for a common disease can be mapped, even within a region of strong LD, and specifically that IDDM2 maps to one or more of three common variants in a ∼2-kb region of chromosome 11p15.
Extracellular Matrix Protects Pancreatic β-Cells Against Apoptosis
Authors: Hammar Eva , Parnaud Géraldine , Bosco Domenico , Perriraz Nadja , Maedler Kathrin , Donath Marc , Rouiller Dominique G. , Halban Philippe A.
Published: March 2007

We have shown previously that culture of β-cells on matrix derived from 804G cells and rich in laminin-5 improves their function. The purpose of this study was to investigate whether this matrix protects β-cells against apoptosis and to elucidate signaling pathways involved. Matrix protected sorted rat β-cells against apoptosis under standard conditions (11.2 mmol/l glucose, 10% serum), after serum deprivation (1% serum), and in response to interleukin-1β (IL-1β; 2 ng/ml), compared with control (poly-l-lysine [pLL]). Caspase-8 activity was reduced in cells cultured on matrix, whereas focal adhesion kinase (FAK), protein kinase B (PKB, or Akt), and extracellular signal–regulated kinase (ERK) phosphorylation was augmented. Treatment (4 h) with an anti-β1 integrin antibody, with the ERK pathway inhibitor PD98059, and/or with the phosphatidylinositol 3-kinase inhibitor LY294002 augmented cell death on 804G matrix but not on pLL. In long-term assays (48 h), PD98059 but not LY294002 drastically augmented cell death on 804G matrix but did so to a lesser extent on pLL. The protein inhibitor of nuclear factor-κB (IκBα) was overexpressed in cells cultured 18 h on matrix with partial blockade by PD98059. In summary, this study provides evidence for activation of signaling pathways and gene expression by extracellular matrix leading to improved β-cell survival.
Partial Gene Deletion of Endothelial Nitric Oxide Synthase Predisposes to Exaggerated High-Fat Diet—Induced Insulin Resistance and Arterial Hypertension
Authors: Cook Stéphane , Hugli Olivier , Egli Marc , Ménard Barbara , Thalmann Sébastien , Sartori Claudio , Perrin Christophe , Nicod Pascal , Thorens Bernard , Vollenweider Peter , Scherrer Urs , Burcelin Rémy
Published: March 2007

Nitric oxide (NO) plays a major role in the regulation of cardiovascular and metabolic homeostasis, as evidenced by insulin resistance and arterial hypertension in endothelial NO synthase (eNOS) null mice. Extrapolation of these findings to humans is difficult, however, because eNOS gene deficiency has not been reported. eNOS gene polymorphism and impaired NO synthesis, however, have been reported in several cardiovascular disease states and could predispose to insulin resistance. High-fat diet induces insulin resistance and arterial hypertension in normal mice. To test whether partial eNOS deficiency facilitates the development of insulin resistance and arterial hypertension during metabolic stress, we examined effects of an 8-week high-fat diet on insulin sensitivity (euglycemic clamp) and arterial pressure in eNOS+/− mice. When fed a normal diet, these mice had normal insulin sensitivity and were normotensive. When fed a high-fat diet, however, eNOS+/− mice developed exaggerated arterial hypertension and had fasting hyperinsulinemia and a 35% lower insulin-stimulated glucose utilization than control mice. The partial deletion of the eNOS gene does not alter insulin sensitivity or blood pressure in mice. When challenged with nutritional stress, however, partial eNOS deficiency facilitates the development of insulin resistance and arterial hypertension, providing further evidence for the importance of this gene in linking metabolic and cardiovascular disease.
Surface Expression of Collagen Receptor Fc Receptor-γ/Glycoprotein VI Is Enhanced on Platelets in Type 2 Diabetes and Mediates Release of CD40 Ligand and Activation of Endothelial Cells
Authors: Cabeza Natalia , Li Zhongyan , Schulz Christian , Kremmer Elisabeth , Massberg Steffen , Bültmann Andreas , Gawaz Meinrad
Published: March 2007

Diabetes is associated with an enhanced collagen-mediated platelet activation that contributes significantly to thromboischemic complications. In this study, the platelet collagen receptor glycoprotein VI (GPVI) was studied in 385 patients with type 2 diabetes. Surface expression of the platelet Fc receptor that forms a functional complex with GPVI was significantly increased in patients with diabetes compared with those without diabetes (P = 0.02). Fc receptor expression correlated with GPVI expression and was found to be independently associated with diabetes (r = 0.529, P < 0.001). Stimulation of GPVI through a specific anti-GPVI monoclonal antibody significantly enhanced surface expression of CD40L (P = 0.006). Because CD40L is a potent platelet-derived cytokine that is involved in thrombosis and atherosclerosis, we evaluated the effect of GPVI-mediated release of CD40L on activation of endothelial cells. Coincubation of GPVI-stimulated platelets resulted in substantial enhanced endothelial surface expression of CD62P, αvβ3, and intercellular adhesion molecule 1 (P < 0.05) and secretion of monocyte chemoattractant protein 1 of cultured human umbilical vein endothelial cells (P < 0.01). These results suggest that the function of collagen receptor GPVI is altered in type 2 diabetes and may play an important role in atherothrombotic complications. Inhibition of GPVI may be a promising pharmacological target in the treatment of high-risk diabetic patients.
Variation at the Insulin Gene VNTR (Variable Number Tandem Repeat) Polymorphism and Early Growth
Authors: Bennett Amanda J. , Sovio Ulla , Ruokonen Aimo , Martikainen Hannu , Pouta Anneli , Taponen Saara , Hartikainen Anna-Liisa , King Vanessa J. , Elliott Paul , Järvelin Marjo-Riitta , McCarthy Mark I.
Published: March 2007

Variation at the insulin gene (INS-)VNTR (variable number of tandem repeats) minisatellite polymorphism has been reported to be associated with both early growth and adult metabolic phenotypes. However, the samples studied have been small and the relationship between INS-VNTR variation and parameters of early growth inconsistent, with four previous studies producing conflicting results. We have studied the relationship between INS-VNTR class (measured by genotyping the nearby −23HphI variant with which it is in tight linkage disequilibrium) and early growth in 5,646 members of the Northern Finnish Birth Cohort of 1966. Comparing class III homozygotes with other genotypes using multivariate linear regression analysis, we found no significant associations with any early growth measure (birth weight, birth length, ponderal index, and head circumference at 1 year), even after stratifying subjects by growth trajectory during infancy and/or birth order. For example, among infants with limited postnatal growth realignment (n = 2,470), class III/III infants were no heavier at birth (difference [±SE] in the means [fully adjusted], 58 ± 51 g; P = 0.26) than class I/− infants. No significant associations were detected following reanalysis with an additive model (for example, for birth weight, β = 20 g [95% CI −3 to 44], P = 0.09). Studies of this large population-based cohort have failed to generate convincing evidence that INS-VNTR variation influences early growth.
Severe Persistent Hyperinsulinemic Hypoglycemia due to a De Novo Glucokinase Mutation
Authors: Cuesta-Muñoz Antonio L. , Huopio Hanna , Otonkoski Timo , Gomez-Zumaquero Juan M. , Näntö-Salonen Kirsti , Rahier Jacques , López-Enriquez Soledad , García-Gimeno Maria A. , Sanz Pascual , Soriguer Federico C. , Laakso Markku
Published: March 2007

Glucokinase (GK) is a glycolytic key enzyme that functions as a glucose sensor in the pancreatic β-cell, where it governs glucose-stimulated insulin secretion (GSIS). Heterozygous inactivating mutations in the glucokinase gene (GCK) cause a mild form of diabetes (maturity-onset diabetes of the young [MODY]2), and activating mutations have been associated with a mild form of familial hyperinsulinemic hypoglycemia. We describe the first case of severe persistent hyperinsulinemic hypoglycemia due to a “de novo” mutation in GCK (Y214C). A baby girl presented with hypoglycemic seizures since the first postnatal day as well as with inappropriate hyperinsulinemia. Severe hypoglycemia persisted even after treatment with diazoxide and subtotal pancreatectomy, leading to irreversible brain damage. Pancreatic histology revealed abnormally large and hyperfunctional islets. The mutation is located in the putative allosteric activator domain of the protein. Functional studies of purified recombinant glutathionyl S-transferase fusion protein of GK-Y214C showed a sixfold increase in its affinity for glucose, a lowered cooperativity, and increased kcat. The relative activity index of GK-Y214C was 130, and the threshold for GSIS predicted by mathematical modeling was 0.8 mmol/l, compared with 5 mmol/l in the wild-type enzyme. In conclusion, we have identified a de novo GCK activating mutation that causes hyperinsulinemic hypoglycemia of exceptional severity. These findings demonstrate that the range of the clinical phenotype caused by GCK mutations varies from complete insulin deficiency to extreme hyperinsulinemia.
Transcortical Direct Current Potential Shift Reflects Immediate Signaling of Systemic Insulin to the Human Brain
Authors: Hallschmid Manfred , Schultes Bernd , Marshall Lisa , Mölle Matthias , Kern Werner , Bredthauer Julia , Fehm Horst L. , Born Jan
Published: March 2007

Circulating insulin is thought to provide a major feedback signal for the hypothalamic regulation of energy homeostasis and food intake, although this signaling appears to be slowed by a time-consuming blood-to-brain transport. Here we show, by recording direct current potentials, a rapid onset of the effects of circulating insulin on human brain activity. Recordings were obtained from 27 men who were intravenously injected with insulin (0.1 mU/kg body wt as bolus) and placebo. In a euglycemic condition, hypoglycemia was prevented, while in the hypoglycemic condition, plasma glucose reached a postinjection nadir of 43 mg/dl. Insulin injection induced a marked negative direct current (DC) potential shift starting within 7 min in all subjects. With euglycemic conditions, the DC potential at 10–60 min postinsulin injection averaged −621.3 μV (compared with preinjection baseline). Hypoglycemia reduced this potential to an average of −331.2 μV. While insulin per se did not affect oscillatory electroencephalographic activity, hypoglycemia peaking 25 min after insulin injection was accompanied by an immediate increase in θ activity. The rapid emergence of the DC potential shift, reflecting gross ionic changes in brain tissues, indicates that systemic insulin can serve as an immediate feedback signal in the control of hypothalamic and higher brain functions.
Increased Serum Leptin Protects From Adiposity Despite the Increased Glucose Uptake in White Adipose Tissue in Mice Lacking p85α Phosphoinositide 3-Kinase
Authors: Terauchi Yasuo , Matsui Junji , Kamon Junji , Yamauchi Toshimasa , Kubota Naoto , Komeda Kajuro , Aizawa Shinichi , Akanuma Yasuo , Tomita Motowo , Kadowaki Takashi
Published: March 2007

Mice lacking the p85α regulatory subunit of phosphoinositide (PI) 3-kinase (Pik3r1−/−) showed increased glucose uptake in white adipose tissue (WAT) and skeletal muscle due to increased phosphatidylinositol (3,4,5)-triphosphate [PtdIns(3,4,5)P3] production and on a normal diet had a body weight and fat mass similar to wild-type mice. After 3 months on a high-fat diet, Pik3r1−/− mice still had increased insulin sensitivity and better glucose tolerance than wild-type mice, but showed markedly greater increases in body weight and WAT mass than wild-type mice. On the normal diet, serum leptin levels of Pik3r1−/− mice were significantly higher than in wild-type mice as a result of increased leptin secretion from adipocytes, presumably due to the increased PtdIns(3,4,5)P3 production in adipocytes. Leptin (5 μg/g body wt per day) caused a reduction in food intake and decrease in body weight by the wild-type mice as well as Pik3r1−/− mice, suggesting Pik3r1−/− mice having leptin sensitivity similar to wild-type mice. The slightly increased serum leptin compensated for the increased glucose uptake by adipocytes in Pik3r1−/− mice, thereby preventing adiposity on the normal diet. On the high-fat diet, leptin (5 μg/g body wt per day) failed to decrease food intake or body weight in either genotype, indicating that both genotypes had indeed become severely leptin resistant. Consequently, leptin secretion was unable to sufficiently compensate for the severe leptin resistance caused by the high-fat diet, thereby failing to prevent obesity in Pik3r1−/− mice. Our findings suggest that primary increase in serum leptin on the normal diet play a role in the protection from adiposity in Pik3r1−/− mice.
Expression of Angiogenic Factors During Acute Coronary Syndromes in Human Type 2 Diabetes
Authors: Marfella Raffaele , Esposito Katherine , Nappo Francesco , Siniscalchi Mario , Sasso Ferdinando Carlo , Portoghese Michele , Pia Di Marino Maria , Baldi Alfonso , Cuzzocrea Salvatore , Di Filippo Clara , Barboso Guglielmo , Baldi Feliciano , Rossi Francesco , D’Amico Michele , Giugliano Dario
Published: March 2007

Inadequate angiogenic response to ischemia in diabetic myocardium could result in poor collateral formation. Because hypoxia-inducible factor (HIF)-1α is a transcriptional activator of vascular endothelial growth factor (VEGF) and is critical for initiating angiogenic responses to hypoxia, we investigated the expression of HIF-1α and VEGF in specimens of human heart tissue to elucidate the molecular responses to myocardial ischemia in diabetic patients during unstable angina. Moreover, accumulation of a marker of protein nitration nitrotyrosine, as well as the superoxide anion (O2−) levels and inducible nitric oxide synthase (iNOS), were evaluated. Ventricular biopsy specimens from 15 type 2 diabetic and 14 nondiabetic patients presenting with unstable angina (ischemic group) and from 20 patients (11 type 2 diabetic and 9 nondiabetic patients) who underwent coronary bypass surgery without angina within the preceding 10 days (control group) were collected during coronary bypass surgery. Nondiabetic patients had higher HIF-1α and VEGF expressions compared with diabetic patients (P < 0.001). As compared with nondiabetic specimens, diabetic specimens showed higher levels of both iNOS mRNA and protein levels (P < 0.001) associated with the highest tissue levels of nitrotyrosine and O2− (P < 0.001). Diabetes is associated with increased myocardial tissue levels of iNOS, O2−, and nitrotyrosine and reduced expression of myocardial angiogenesis factors during ischemia.
Development of Albuminuria and Glomerular Lesions in Normoglycemic B6 Recipients of db/db Mice Bone Marrow
Authors: Zheng Feng , Cornacchia Flavia , Schulman Ivonne , Banerjee Anita , Cheng Qing-li , Potier Mylene , Plati Anna Rita , Berho Mariana , Elliot Sharon J. , Li Jie , Fornoni Alessia , Zang Yun-Juan , Zisman Ariel , Striker Liliane J. , Striker Gary E.
Published: March 2007

The pathologic hallmarks of diabetic nephropathy are excess mesangial extracellular matrix (ECM) and mesangial cell proliferation. We previously showed that mesangial cell phenotypic changes play an important role in the pathogenesis of diabetic nephropathy. We concluded that phenotypic changes were present in bone marrow (BM)-derived mesangial cell progenitors, as transplantation of BM from db/db mice, a model of type 2 diabetic nephropathy, transferred the db genotype and a nephropathy phenotype to naive B6 mice recipients. The recipients did not develop diabetes; however, they did develop albuminuria and glomerular lesions mirroring those in the donors (i.e., glomerular hypertrophy, increased ECM, and increased cell number with cell proliferation). We found that matrix metalloproteinase 2 (MMP-2) facilitated invasion of the mesangial cells into ECM and proliferation in vitro. Thus, increased MMP-2 activity in db/db mesangial cell progenitors may partially explain increased mesangial cell repopulation and proliferation in B6 recipients of db/db BM. In summary, BM-derived mesangial cell progenitors may play a crucial role in the development and progression of ECM accumulation and mesangial cell proliferation in this model of diabetic nephropathy in type 2 diabetes.
Changes in IGF Activities in Human Diabetic Vitreous
Authors: Guidry Clyde , Feist Richard , Morris Robert , Hardwick Charles W.
Published: March 2007

Müller cells, the principal glia of the retina, generate tractional forces in response to IGF-I and platelet-derived growth factor and are present in diabetic fibro-vascular scar tissues causing traction retinal detachment. While diabetes-associated increases in vitreous IGFs have been reported, paradoxically high concentrations of these same growth factors in normal vitreous suggest the presence of more complex mechanisms regulating growth factor bioavailability. To define diabetes-associated changes in vitreous biological activity, the stimulatory effects of 68 samples were evaluated using Müller cell tractional force generation as a target bioassay. Dose-response profiles were used to calculate vitreous specific activity (per unit protein) and total vitreous activity (per unit volume). Vitreous samples from patients lacking diabetes or other retinal pathology had undetectable or low activities, whereas diabetic retinopathy was associated with 6.9- and 8.7-fold increases in vitreous specific and total activities, respectively. Secondary analyses revealed no activity differences associated with patient sex, age, or the presence of vitreous hemorrhage. However, compared with diabetes alone, the presence of proliferative diabetic retinopathy was associated with additional 2.3-fold increases in vitreous specific and total activities. Vitreous dose-response assays performed with and without growth factor–neutralizing antibodies enable attribution of vitreous activity to IGFs (53.9%) and, to a lesser extent, platelet-derived growth factors (14.5%). Because the observed increases in vitreous growth factor activity grossly exceed the reported increases in growth factor concentration, these data indicate that diabetes-associated changes in vitreous biological activity involve more complex biochemical changes that ultimately yield increased growth factor bioavailability and/or Müller cell responsiveness.
Association Analysis of the Lymphocyte-Specific Protein Tyrosine Kinase (LCK) Gene in Type 1 Diabetes
Authors: Hulme John S. , Barratt Bryan J. , Twells Rebecca C.J. , Cooper Jason D. , Lowe Chris E. , Howson Joanna M.M. , Lam Alex C. , Smink Luc J. , Savage David A. , Undlien Dag E. , Guja Cristian , Ionescu-Tîırgovişte Constantin , Tuomilehto-Wolf Eva , Tuomilehto Jaakko , Todd John A.
Published: March 2007

Prior data associating the expression of lymphocyte-specific protein tyrosine kinase (LCK) with type 1 diabetes, its critical function in lymphocytes, and the linkage of the region to diabetes in the nonobese diabetic (NOD) mouse model make LCK a premier candidate for a susceptibility gene. Resequencing of LCK in 32 individuals detected seven single nucleotide polymorphisms (SNPs) with allele frequencies >3%, including four common SNPs previously reported. These and six other SNPs from dbSNP were genotyped in a two-stage strategy using 2,430 families and were all shown not to be significantly associated with type 1 diabetes. We conclude that a major role for the common LCK polymorphisms in type 1 diabetes is unlikely. However, we cannot rule out the possibility of there being a causal variant outside the exonic, intronic, and untranslated regions studied.
Identification of a Functionally Important Negatively Charged Residue Within the Second Catalytic Site of the SUR1 Nucleotide-Binding Domains
Authors: Campbell Jeff D. , Proks Peter , Lippiat Jonathan D. , Sansom Mark S.P. , Ashcroft Frances M.
Published: March 2007

The ATP-sensitive K+ channel (KATP channel) couples glucose metabolism to insulin secretion in pancreatic β-cells. It is comprised of sulfonylurea receptor (SUR)-1 and Kir6.2 proteins. Binding of Mg nucleotides to the nucleotide-binding domains (NBDs) of SUR1 stimulates channel opening and leads to membrane hyperpolarization and inhibition of insulin secretion. To elucidate the structural basis of this regulation, we constructed a molecular model of the NBDs of SUR1, based on the crystal structures of mammalian proteins that belong to the same family of ATP-binding cassette transporter proteins. This model is a dimer in which there are two nucleotide-binding sites, each of which contains residues from NBD1 as well as from NBD2. It makes the novel prediction that residue D860 in NBD1 helps coordinate Mg nucleotides at site 2. We tested this prediction experimentally and found that, unlike wild-type channels, channels containing the SUR1-D860A mutation were not activated by MgADP in either the presence or absence of MgATP. Our model should be useful for designing experiments aimed at elucidating the relationship between the structure and function of the KATP channel.
Kinetics-Effect Relations of Insulin-Releasing Drugs in Patients With Type 2 Diabetes
Authors: Melander Arne
Published: March 2007

Sulfonylureas and glinides have similar mechanisms of action but differ in receptor affinity and binding sites and in absorption and elimination rates. This promotes differences in potency, rate of onset, and duration of action. While prominent in single-dose studies, these differences have less importance during long-term sulfonylurea treatment: at ordinary dosages, rapid- and short-acting (glipizide) and slow- and long-acting (glyburide) sulfonylureas maintained continuously effective plasma levels and similar 24-h glucose control. Moreover, there was no difference in patient outcome between the first-generation sulfonylurea chlorpropamide and the second-generation glyburide in the U.K. Prospective Diabetes Study. However, the risk of long-lasting and hence dangerous hypoglycemia is higher with these two long-acting sulfonylureas. Conversely, this risk should be low with the short-acting glinides, but seemingly at the expense of less effective glucose control. The most important kinetics-effect relations are that hyperglycemia delays sulfonylurea absorption and that the sulfonylurea dose-response curve is bell shaped; continuous sulfonylurea exposure over a certain level (e.g., 10 mg glipizide) impairs rather than improves insulin and glucose responses to sulfonylurea (downregulation). Accordingly, a vicious circle may be established: unrelenting hyperglycemia may promote sulfonylurea dose increase, which increases hyperglycemia, promoting further dose increase and eventually therapeutic failure.