bluuebunny/crossref_metadata_2025 huggingface dataset:
Hypoglycemia in the Diabetes Control and Complications Trial
Authors:   The Diabetes Control and Complications Trial Research Group
Published: September 2013

A total of 1,441 patients with IDDM were randomly assigned to receive either intensive (n = 711) or conventional (n = 730) diabetes therapy in the Diabetes Control and Complications Trial (DCCT). The patients were followed for an average of 6.5 years. Subjects were instructed to report all episodes of suspected severe hypoglycemia to their health care team. In addition, at quarterly follow-up visits, each subject was asked about the occurrence of severe hypoglycemia. There were 3,788 episodes of severe hypoglycemia (requiring assistance); 1,027 of these episodes were associated with coma and/or seizure. A total of 65% percent of patients in the intensive group vs. 35% of patients in the conventional group had at least one episode of severe hypoglycemia by the study end; the overall rates of severe hypoglycemia were 61.2 per 100 patient-years vs. 18.7 per 100 patient-years in the intensive and conventional treatment groups, respectively, with a relative risk (RR) of 3.28. The relative risk for coma and/or seizure was 3.02 for intensive therapy. The increased risk with intensive treatment persisted over each of the 9 years of follow-up in the DCCT and over the calendar years 1984–1993 during which the study was conducted. When baseline patient characteristics were examined for effects on the risk of severe hypoglycemia, the relative risk of hypoglycemia for intensive versus conventional treatment was ≥2 for all subgroups. Several subgroups defined by baseline characteristics, including males, adolescents, and subjects with no residual C-peptide or with a prior history of hypoglycemia, had a particularly high risk of severe hypoglycemia in both treatment groups. Analyses of the cumulative incidence of successive episodes indicated that intensive treatment was also associated with an increased risk of multiple episodes within the same patient (e.g., 22% experienced five or more episodes of severe hypoglycemia within the first 5 years of follow-up vs. 4% in the conventional group). Within both treatment groups, patients who experienced severe hypoglycemia were at increased risk of subsequent episodes. Approximately 30% of patients in each group experienced a second episode within the 4 months following the first episode of severe hypoglycemia. Within each treatment group, the number of prior episodes of hypoglycemia was the strongest predictor of the risk of future episodes, followed closely by the current HbA1c value. After adjustment for the current quarterly HbA1c level, intensive treatment was still associated with a significantly increased risk of hypoglycemia, indicating that the increased risk with intensive treatment is not completely explained by differences in HbA1c values.
Classification, Differential Diagnosis, and Staging of Diabetic Peripheral Neuropathy
Authors: Thomas PK
Published: September 2013

The peripheral nerve disorders associated with diabetes are complex and probably involve a variety of causative mechanisms. This may give rise to difficulty in the classification of individual cases. A broad separation into rapidly reversible or more persistent phenomena is helpful. The former, which can be categorized as “hyperglycemic neuropathy,” include minor sensory symptoms, reduced nerve conduction velocity, and resistance to ischemic conduction failure. From analogy with experimental studies in animals, nerve hypoxia is likely to play a significant role in their origin. Of the more persistent phenomena, a distal symmetric polyneuropathy that predominantly affects sensory and autonomic function is the most common manifestation. A distal axonopathy of dying-back type may represent the underlying pathogenetic basis. Other more persistent phenomena consist of focal and multifocal lesions giving rise to cranial, thoraco-abdominal, and limb neuropathies, including proximal lower limb motor neuropathy (diabetic amyotrophy). Some of these may have an ischemic basis. Multifocal proximal lesions can summate to produce an approximately symmetric diffuse distal neuropathy. Focal lesions at sites of entrapment or external compression may reflect an abnormal susceptibility of diabetic nerve to compressive damage. There is also evidence that focal inflammatory, including vasculitic, lesions may be involved in proximal lower limb neuropathies. Finally, superimposed chronic inflammatory demyelinating polyneuropathy may occur. For the evaluation of possible treatment regimens, it is essential that cases should be correctly classified as to type. Thus, the features falling into the category of hyperglycemic neuropathy should not contaminate the assessment of distal symmetric polyneuropathy. For this type, a widely accepted scheme for staging devised by P.J. Dyck is available. Other schemes are also available for the assessment of such cases, with differing degrees of complexity. Evaluation by serial nerve biopsies has also been proposed.
Pathophysiology of Diabetic Gastroparesis
Authors: Enck Paul , Frieling Thomas
Published: September 2013

Our understanding of the nature of diabetic gastroparesis has advanced in the last decade due to new investigational procedures (electrogastrography, visceral evoked potential recording), and transferring these insights into clinical routine will be our task in the future. Meanwhile, the clinical relevance of gastroparesis—whether overt or silent—remains unquestioned: proper gastric emptying is a prerequisite for adequate metabolic control, and its disturbance may result not only in further progression of the chronic complications of the disease, but also in the false assumption that these patients are not compliant with their doctor's management—the patients just may have delayed emptying of their stomach without noticing it.
Essential Fatty Acids in the Management of Impaired Nerve Function in Diabetes
Authors: Horrobin David F
Published: September 2013

Impaired conversion of linoleic acid to γ-linolenic acid (GLA) has been demonstrated in animal diabetes and inferred from blood fatty acid profiles in human diabetes. This impairment could theoretically lead to defective nerve function because metabolites of GLA are known to be important in nerve membrane structure, nerve blood flow, and nerve conduction. Administration of GLA corrects the impaired nerve function in animal models of diabetes. Two multicenter, randomized, placebo-controlled trials in humans with diabetic neuropathy have shown significant benefits of GLA as compared with placebo in neurophysiological parameters, thermal thresholds, and clinical sensory evaluations. Further work is needed to define the place of this therapeutic approach and its interactions with other treatment modalities.
Cell-Specific Regulation of IRS-1 Gene Expression: Role of E Box and C/EBP Binding Site in HepG2 Cells and CHO Cells
Authors: Matsuda Kohji , Araki Eiichi , Yoshimura Ryohei , Tsuruzoe Kaku , Furukawa Noboru , Kaneko Kengo , Motoshima Hiroyuki , Yoshizato Kazuaki , Hideki Kishikawa , Shichiri Motoaki
Published: September 2013

Insulin receptor substrate 1 (IRS-1) is one of the major substrates of insulin receptor tyrosine kinase and mediates multiple insulin signals downstream. We have previously shown that the levels of IRS-1 mRNA varied in different tissues. To elucidate the molecular mechanisms of the tissue specific regulation of IRS-1, we have studied the cis-acting elements and transacting factors in CHO and HepG2 cells. Using the chloramphenicol acetyltransferase (CAT) assay with the various deletion mutants of the IRS-1 promoter–CAT fusion plasmids, several regions responsible for positive or negative regulation in each cell line were identified. A region from −1645 to −1585 bp, which regulated expression negatively in CHO cells and positively in HepG2 cells, was further analyzed. Within this region sa fragment from −1645 to −1605 bp upregulated the IRS-1 promoter only in HepG2 cells, whereas a fragment from −1605 to −1585 bp downregulated only in CHO cells. In the gel mobility shift assay, several nuclear proteins that bind to these fragments were detected, and among them, two nuclear proteins that bind to a potential E box (nucleotide [nt] −1635 to −1630) and two nuclear proteins that bind to a potential C/EBP binding site (nt −1599 to −1591) were identified in HepG2 and CHO cells, respectively. CAT assays using promoters mutated at the E box or at the C/EBP binding site revealed that these sequences were responsible for cell-specific regulation of the IRS-1 gene. We therefore concluded that the two nuclear proteins that bind to the E box regulate IRS-1 gene expression positively in HepG2 cells and the two nuclear proteins that bind to the C/EBP binding site regulate it negatively in CHO cells.
New Monoclonal Antibody Diagnostic Reagents for Type I Diabetes: Differential Lymphocyte Surface Antigen Expression Related to Disease
Authors: Pinies Jose A , Fu Yineng , Vazquez J A , Faustman Denise
Published: September 2013

New cellular-based reagents are needed to diagnose type I diabetes as well as to monitor the outcomes of clinical trials at early time points. Four new monoclonal antibodies (mAbs) have been shown to demonstrate reduced binding to lymphocytes from identical twins with long-term type I diabetes relative to that observed with lymphocytes from their twin partners without diabetes or from control subjects. Biochemical analysis revealed mAb 3G12EG recognized an unidentified 45-kDa protein, whereas mAb 2E8F1 and 5B6E11 did not appear to precipitate specific proteins as detected by SDS-PAGE. Electrophoresis under reducing and nonreducing conditions and peptide mapping revealed that mAb 8F410 recognizes a novel dimeric form of HLA class I molecule. Predictions from crystallography studies suggested previously this class I dimer as the optimal activation of a single CD8 T-cell. In B-cells from both normal and diabetic individuals, the class I dimer was minimally associated with β2-microglobulin rapidly formed in the endoplasmic reticulum. These new reagents appear to be able to identify new lymphocyte surface phenotypes associated with diabetes expression in both fresh blood samples and Epstein-Barr virus–established cell lines.
Impaired Basal Glucose Effectiveness in NIDDM: Contribution of Defects in Glucose Disappearance and Production, Measured Using an Optimized Minimal Model Independent Protocol
Authors: Basu Ananda , Caumo Andrea , Bettini Francesca , Gelisio Anna , Alzaid Aus , Cobelli Claudio , Rizza Robert A
Published: September 2013

People with NIDDM are resistant to insulin. The present studies sought to determine whether the ability of glucose to regulate its own metabolism in the presence of basal insulin concentrations is impaired. To address this question, basal insulin concentrations were maintained constant with an exogenous insulin infusion, while endogenous hormone secretion was inhibited by somatostatin. The integrated glycemic response above baseline during identical prandial glucose infusions was greater (1,411 ± 94 vs. 938 ± 45 mmol/l per 5 h; P < 0.01) in the diabetic subjects than in the nondiabetic subjects, indicating a decrease in net glucose effectiveness. [6-3H]glucose also was infused to determine whether the decrease in net glucose effectiveness was due to a decrease in the ability of glucose to stimulate its own uptake and/or to suppress its own production. Despite identical rates of tracer infusion, the increment in plasma concentration of [6-3H]glucose was higher (4.50 ± 0.29 vs. 3.16 ± 0.21 × 105 dpm/ml per 5 h; P < 0.05) in the diabetic subjects than in the nondiabetic subjects. This was due to both a decrease (P < 0.05) in the ability of glucose to stimulate its own disappearance via mass action and to a greater (P < 0.01) inhibitory effect of glucose on its own clearance. The increase in glucose concentration resulted in prompt and comparable suppression of endogenous glucose production in both groups. Under these optimized conditions, indexes of glucose effectiveness calculated with both the “cold” and “hot” minimal models also were lower (P < 0.05) in the diabetic subjects than in the nondiabetic subjects and were highly correlated (r = 0.94–0.99; P < 0.001) with the indexes of glucose effectiveness calculated from the increments above baseline of glucose and [6-3H]glucose concentration. We conclude that the ability of glucose to regulate its own metabolism in the presence of basal insulin concentrations is abnormal in people with NIDDM.
Differential Regulation of the p80 Tumor Necrosis Factor Receptor in Human Obesity and Insulin Resistance
Authors: Hotamisligil Gökhan S , Arner Peter , Atkinson Richard L , Spiegelman Bruce M
Published: September 2013

Previous studies have shown that tumor necrosis factor (TNF)-α production from adipose tissue is elevated in rodent and human obesity and plays an important role in insulin resistance in experimental animal models. In this study, we examined the adipose expression of both TNF receptors (TNFR1 and TNFR2) in human obesity and demonstrated that obese female subjects express approximately twofold more TNFR2 mRNA in fat tissue and approximately sixfold more soluble TNFR2 in circulation relative to lean control subjects. In contrast, TNFR1 expression and protein levels were similar in these subjects. TNFR2 expression levels in adipose tissue were strongly correlated with BMI (r = 0.65, P < 0.001) and level of hyperinsulinemia (P < 0.001), an indirect measure of insulin resistance, as well as level of TNF-α mRNA expression in fat tissue (r = 0.56, P < 0.001). These results suggest that TNFR2 might play a role in human obesity by modulating the actions of TNF-α.
Anti-Insulin Activity in Normal Newborn Cord-Blood Serum: Absence of IgG-Mediated Insulin Binding
Authors: Bilbao J Ramón , Calvo Begoña , Urrutia Inés , Linares Alberto , Castaño Luis
Published: September 2013

Insulin autoantibodies (IAAs) are present in ∼60% of type I diabetes patients at onset and are used as predictors for the disease. Although the prevalence of IAAs in the general population has been reported to be <1%, preliminary data have pointed out a higher proportion of IAA positivity in newborn cord-blood serum, and some authors have suggested that they are immunoglobulin G antibodies, resulting from a hypothetical gestational insulitis. To characterize this insulin-binding activity, we analyzed cord-blood sera from 100 healthy newborns, as well as serum from 21 of their mothers at delivery, 179 new-onset type I diabetic patients, and 200 healthy control subjects. IAAs were present in 0.5% of the control subjects and 54% of newonset type I diabetic patients. On the other hand, 96% of the newborn cord-blood sera showed anti-insulin activity, while it was detected in only 14% of their mothers. No significant differences were observed between cord sera and the general population for isletcell or anti-GAD autoantibodies. Anti-insulin activity in cord serum was not bound by protein A or protein G, in contrast with type I diabetes–related IAA activity. We conclude that this insulin-binding activity, present in most newborn cord sera and specific to the child, is not IgG mediated. These data, together with the absence of other pancreatic autoimmunity markers in this population, suggest that it is an isolated phenomenon not related to type I diabetes or other pancreatic autoimmune processes and is due to the presence of a cross-reacting molecule in cord blood that has yet to be identified.
Rejection of Islets Differing by a Single Antigen Is Dependent on Donor MHC
Authors: Vigneau-Hermellin Murielle , Vitali Letizia , Tardivel Isabelle , Rabaud Myriam , Holers Michael V , Carel Jean-Claude
Published: September 2013

To study the effect of expression of a single foreign antigen on the outcome of otherwise compatible mouse islet grafts, we have used transgenic mice expressing the human complement receptor 2 (CR2, CD21, C3d/EBV receptor) on their pancreatic (β-cells (RIPCR2 mice). Donors were RIP-CR2 mice, typed at the major histocompatibility complex (MHC) as H-2k, H-2b, or H-2bxk, and recipients were streptozotocintreated nontransgenic B6 × CBA Fl mice (H-2bxk). H-2b or H-2bxk CR2-expressing islets were not rejected (mean survival time [MST] >100 days) but induced a peri-insulitis and an antibody response to CR2. In contrast, H-2k CR2-expressing islets were rejected in 80% of the cases with a MST of 65 ± 23 days and were massively infiltrated by a destructive insulitis. In both cases, the infiltrate was mainly made of CD4+ cells, with few CD8+ cells. The isotype of IgG antibody response to CR2 was studied: recipients of H-2k grafts had a predominantly IgGl response, while recipients of H-2b grafts had a balanced IgG2a and IgGl response. To further evaluate the mechanism of differential rejection of the two types of grafts, recipients were immunized with CR2-expressing rat insulinoma cells before transplantation. Preimmunization with CR2 did not affect the outcome of H-2b grafts but greatly accelerated the rejection of H-2k grafts. These experiments indicate that expression of a single foreign antigen on β-cells triggers an immune response leading to rejection or to peri-insulitis, depending on the MHC of donor islets.
Binding Properties of Glutamate Receptors in Streptozotocin-Induced Diabetes in Rats
Authors: Joël Gagné , Milot Martin , Gélinas Sylvie , Lahsaïni Ahmed , Trudeau François , Martinoli Maria-Grazia , Massicotte Guy
Published: September 2013

The biochemical mechanisms by which diabetes modulates cognitive function are not well established. Here, we determined the effects of streptozotocin (STZ) administration on the binding properties of α-amino-3-hydroxy-5-methylisoxazolepropionic acid (AMPA) and N-methyl-D-aspartate (NMDA) subtypes of glutamate receptors in rats, using quantitative autoradiographic analysis of 3H-AMPA and [3H]glutamate binding on brain tissue sections. The STZ injection (70 mg/kg intraperitoneally) produced a reduction of 3H-AMPA binding in various brain regions, an effect that is due to a decrease in receptor affinity. The STZ-induced reduction of 3H-AMPA binding varied in different brain structures, being more pronounced in the striatum, cerebral cortex, and hippocampus and almost absent in the cerebellum. Western blots performed on hippocampal membranes revealed that the decrease in 3H-AMPA binding is possibly associated with changes in immunologic properties for one glutamate receptor subunit (GluRl). Finally, the effect of STZ-induced diabetes appeared to be specific to the AMPA subtype of glutamate receptors, as the same treatment did not modify [3H]glutamate binding to NMDA receptors. These changes in AMPA receptor properties may have important implications for understanding the biochemical mechanisms underlying cognitive impairment in diabetes.
New Susceptibility Locus for NIDDM Is Localized to Human Chromosome 20q
Authors: Ji Linong , Malecki Maciek , Warram James H , Yang Yadong , Rich Stephen S , Krolewski Andrzej S
Published: September 2013

To test the hypothesis that a gene (or genes) in the “MODY1 region” of the long arm of chromosome 20 contributes to the development of NIDDM, we conducted linkage studies in 29 extended Caucasian families in which many members were affected with NIDDM. A total of 498 individuals, including 159 NIDDM patients with an average age at diagnosis of 47 years, were genotyped for eight highly polymorphic microsatellite markers spanning a 31-cM region on chromosome 20q12–13.1. Using affected sib-pair analysis, we obtained evidence suggesting linkage between NIDDM and markers D20S119, D20S178, and D20S197 (allele sharing identical-by-descent [IBD], 0.56 for all three; P = 0.005, P = 0.009, and P = 0.004, respectively). Multipoint nonparametric linkage (NPL) analysis also showed evidence for linkage of NIDDM with the same three markers. The evidence for linkage was much stronger (allele sharing IBD by affected sibpairs, 0.64 [P < 0.0001]; maximum NPL score, 3.3 [P = 0.009]) in the 14 families whose average age at diagnosis of NIDDM was above the median (47 years) for all families. In these 14 families, one particular allele of the microsatellite D20S197 was transmitted from heterozygous parents to NIDDM offspring more frequently than expected (P < 0.01). This indicates that the marker allele and the disease allele are in linkage disequilibrium, implying that they are in close proximity. Consequently, the recently identified MODY1 gene (hepatocyte nuclear factor 4) is an unlikely candidate gene for NIDDM in our families, since it is located about 8 cM centromeric of D20S197. In conclusion, we have identified a new region on chromosome 20q that contains one or more NIDDM genes distinct from the recently identified MODY1 gene.
Linkage of Genetic Markers on Human Chromosomes 20 and 12 to NIDDM in Caucasian Sib Pairs With a History of Diabetic Nephropathy
Authors: Bowden Donald W , Sale Michèle , Howard Timothy D , Qadri Asif , Spray Beverly J , Rothschild Cynthia B , Akots Gita , Rich Stephen S , Freedman Barry I
Published: September 2013

The potential contribution of maturity-onset diabetes of the young (MODY) genes to NIDDM susceptibility in African-American and Caucasian NIDDM-affected sibling pairs with a history of adult-onset diabetic nephropathy has been evaluated. Evidence for linkage to NIDDM was found with polymorphic loci that map to the long arms of human chromosomes 20 and 12 in regions containing the MODY1 and MODY3 genes. Nonparametric analysis of chromosome 20 inheritance data collected with the MODYl-linked marker D20S197 provides evidence forlinkage to NIDDM with a P value of 0.005 in Caucasian sib pairs using affected sibpair (ASP) analyses. Nonparametric analysis of chromosome 12 inheritance data collected with the MODY3-linked markers D12S349 and D12S86 provides evidence for linkage to NIDDM with P values of 0.04 and 0.006, respectively, in Caucasian sib pairs using similar analyses. No evidence for linkage of MODY1 and MODY3 markers to NIDDM in African-American sib pairs was observed. In addition, no evidence for linkage to MODY2 (glucokinase-associated MODY) was observed with either study population. Results of multipoint maximum logarithm of odds (LOD) score analysis were consistent with the ASP results. A maximum LOD score of 1.48 was calculated for linkage to MODYl-linked loci and 1.45 to MODY3-linked loci in Caucasian sib pairs. Tabulation of allele sharing in affected sib pairs with D20S197 and D12S349 suggests that affected sibling pairs may inherit susceptibility genes simultaneously from chromosome 20 and chromosome 12. The results suggest that genes contributing to NIDDM in the general Caucasian population are located in the regions containing the MODY1 and MODY3 genes.
A Novel Locus, Mody4, Distal to D7Mit189 on Chromosome 7 Determines Early-Onset NIDDM in Nonobese C57BL/6 (Akita) Mutant Mice
Authors: Yoshioka Masato , Kayo Tsuyoshi , Ikeda Tsurumi , Koizuni Akio
Published: September 2013

In this article, we report on a nonobese C57BL/6 (B6) mouse model of NIDDM named Akita mouse, characterized by early age onset and autosomal dominant mode of inheritance. At 7 weeks of age, the mean morning blood glucose levels (mmol/1) under ad libitum feeding conditions were significantly higher (P < 0.01, analysis of variance [ANOVA]) in diabetic mice than in unaffected mice: 27.3 ± 5.3 for diabetic males (n = 50) and 9.3 ± 1.2 for unaffected males (n = 50); 13.6 ± 3.8 for diabetic females (n = 50) and 8.7 ± 1.1 for unaffected females (n = 50), while corresponding immunoreactive insulin levels in plasma were significantly lower in diabetic mice than in unaffected mice. In vitro insulin secretion was also impaired, even at 4 weeks of age. The 50% survival time for male diabetic mice (305 days) was significantly shorter than that of unaffected counterpart mice but not for diabetic females. Obesity did not occur in diabetic mice. Histological examinations of the pancreas in diabetic mice, from 4 to 35 weeks of age, revealed decreases in the numbers of active β-cells without insulitis. Morphometry demonstrated specific decreases in immunologically detectable insulin density in islets in diabetic mice, even at 4 weeks of age, without changes of relative islet areas. By linkage analysis, a single locus was identified on the basis of 178 N2 mice [(B6 × C3H/He)F1 × B6 and (B6 × C3H/He)F1 × C3H/He]. This locus, which we named Mody4, was mapped to chromosome 7 in a region 2–8 cM distal to D7Mitl89 (logarithm of odds [LOD] score = 15.6 and 10.3).
Modulatory Effect of Erythrocytes on the Platelet Reactivity to Collagen in IDDM Patients
Authors: Vallés Juana , Santos M Teresa , Aznar Justo , Velert Mar , Barberá Gloria , Carmena Rafael
Published: September 2013

Platelets participate in the atherothrombotic complications of diabetes. Recent data demonstrate that platelet reactivity can be modulated via cell-cell interactions with erythrocytes and neutrophils. In this study, platelet reactivity was evaluated in 30 IDDM patients. We used an analytical procedure that permits an independent evaluation of platelet activation (granule release, eicosanoid formation) and platelet recruitment (pro-aggregatory activity of cell-free releasates) after platelet stimulation with collagen in the presence or absence of other blood cells. The interaction between platelets and erythrocytes (hematocrit 40%) resulted in a marked enhancement of platelet activation (5HT, βTG, TXA2 release) and recruitment in both patients and control subjects. The erythrocyte enhancement of platelet TXA2 synthesis and recruitment was significantly higher in the patients, while no differences were detected in platelet granule release. The elevated platelet recruitment in the IDDM patients was found to be due to 1) increased susceptibility of diabetic platelets to the prothrombotic effect of erythrocytes and 2) the greater response of diabetic platelets to their own cell-free releasate. Patients with poor metabolic control (elevated HbA1c) or longer evolution time had an even greater platelet recruitment. The presence of microalbuminuria is not related to the platelet recruitment. Since platelet recruitment is an essential step in thrombus growth, its enhancement may favor thrombotic complications in IDDM.
Genetics of NIDDM in France: Studies With 19 Candidate Genes in Affected Sib Pairs
Authors: Vionnet Nathalie , Hani El Habib , Lesage Suzanne , Philippi Anne , Hager Jörg , Varret Mathilde , Stoffel Markus , Tanizawa Yukio , Chiu Ken C , Glaser Benjamin , Permutt M Alan , Passa Philippe , Demenais Florence , Froguel Philippe
Published: September 2013

As part of an ongoing search for susceptibility loci for NIDDM, we tested 19 genes whose products are implicated in insulin secretion or action for linkage with NIDDM. Loci included the G-protein–coupled inwardly rectifying potassium channels expressed in β-cells (KCNJ3 and KCNJ7), glucagon (GCG), glucokinase regulatory protein (GCKR), glucagon-like peptide I receptor (GLP1R), LIM/homeodomain islet-1 (ISL1), caudal-type homeodomain 3 (CDX3), proprotein convertase 2 (PCSK2), cholecystokinin B receptor (CCKBR}, hexokinase 1 (HK1), hexokinase 2 (HK2), mitochondrial FAD-glycerophosphate dehydrogenase (GPD2), liver and muscle forms of pyruvate kinase (PKL, PKM), fatty acid–binding protein 2 (FABP2), hepatic phosphofructokinase (PFKL), protein serine/threonine phosphatase 1 beta (PPP1CB), and lowdensity lipoprotein receptor (LDLR). Additionally, we tested the histidine-rich calcium locus (HRC) on chromosome 19q. All regions were tested for linkage with microsatellite markers in 751 individuals from 172 families with at least two patients with overt NIDDM (according to World Health Organization criteria) in the sibship, using nonparametric methods. These 172 families comprise 352 possible affected sib pairs with overt NIDDM or 621 possible affected sib pairs defined as having a fasting plasma glucose value of >6.1 mmol/1 or a glucose value of >7.8 mmol/1 2 h after oral glucose load. No evidence for linkage was found with any of the 19 candidate genes and NIDDM in our population by nonparametric methods, suggesting that those genes are not major contributors to the pathogenesis of NIDDM. However, some evidence for suggestive linkage was found between a more severe form of NIDDM, defined as overt NIDDM diagnosed before 45 years of age, and the CCKBR locus (11pl5.4; P = 0.004). Analyses of six additional markers spanning 27 cM on chromosome l ip confirmed the suggestive linkage in this region. Whether an NIDDM susceptibility gene lies on chromosome l ip in our population must be determined by further analyses.
Analysis of Candidate Genes for Susceptibility to Type I Diabetes: A Case-Control and Family-Association Study of Genes on Chromosome 2q31–35
Authors: Owerbach David , Naya Francisco J , Tsai Ming-Jer , Allander Susanne V , Powell David R , Gabbay Kenneth H
Published: September 2013

Recent genome searches suggest a putative linkage of many loci to susceptibility to type I diabetes. The chromosome 2q31–35 region is reported to be linked to susceptibility to type I diabetes and is thought to contain several diabetes susceptibility loci. These candidate genes include the HOXD gene cluster, BETA2, CTLA4, CD28, IGFBP2, and IGFBP5. Association studies in populations and families are required to confirm and/or identify the actual susceptibility loci. We hereby report several previously unknown DNA polymorphisms for HOXD8, BETA2, and IGFBP5, which we have used along with previously known polymorphisms of H0XD8 and CTLA4 to test whether these candidate loci are the susceptibility genes on chromosome 2q31–35. Using a casecontrol design with a subsequent family-association approach to confirm associations, we find no evidence that these candidate genes are associated with susceptibility to type I diabetes.
Leptin Receptor of Zucker Fatty Rat Performs Reduced Signal Transduction
Authors: Yamashita Tatsuya , Murakami Takashi , Iida Mitsuru , Kuwajima Masamichi , Shima Kenji
Published: September 2013

Zucker fatty (fa/fa) rats exhibit overt obesity, hypercholesterolemia, hyperlipidemia, and hyperglycemia as recessive traits. The fa mutation has been determined to be a missense mutation in the extracellular domain of the leptin receptor. We report herein the construction of CHO cells that stably express the fa-type leptin receptor and the characterization of this receptor using mRNA expression levels of the immediate early genes, c-fos, c-jun, and jun-B, which are induced by leptin as a criterion of signal transduction. The fa-type receptor not only exhibits a slightly reduced leptinbinding affinity, but also performs reduced signal transduction.
Leptin Suppression of Insulin Secretion by the Activation of ATP-Sensitive K+ Channels in Pancreatic β-Cells
Authors: Kieffer Timothy J , Heller R Scott , Leech Colin A , Holz George G , Habener Joel F
Published: September 2013

In the genetic mutant mouse models ob/ob or db/db, leptin deficiency or resistance, respectively, results in severe obesity and the development of a syndrome resembling NIDDM. One of the earliest manifestations in these mutant mice is hyperinsulinemia, suggesting that leptin may normally directly suppress the secretion of insulin. Here, we show that pancreatic islets express a long (signal-transducing) form of leptin-receptor mRNA and that β-cells bind a fluorescent derivative of leptin (Cy3-leptin). The expression of leptin receptors on insulin-secreting β-cells was also visualized utilizing antisera generated against an extracellular epitope of the receptor. A functional role for the β-cell leptin receptor is indicated by our observation that leptin (100 ng/ml) suppressed the secretion of insulin from islets isolated from ob/ob mice. Furthermore, leptin produced a marked lowering of ]Ca2+]i in ob/ob β-cells, which was accompanied by cellular hyperpolarization and increased membrane conductance. Cell-attached patch measurements of ob/ob β-Cells demonstrated that leptin activated ATP-sensitive potassium channels (KATP) by increasing the open channel probability, while exerting no effect on mean open time. These effects were reversed by the sulfonylurea tolbutamide, a specific inhibitor of KATP. Taken together, these observations indicate an important physiological role for leptin as an inhibitor of insulin secretion and lead us to propose that the failure of leptin to inhibit insulin secretion from the β-Cells of ob/ob and db/db mice may explain, in part, the development of hyperinsulinemia, insulin resistance, and the progression to NIDDM.
Mechanisms of the Kinetic Defect in Insulin Action in Obesity and NIDDM
Authors: Nolan John J , Ludvik Bernhard , Baloga Joseph , Reichart Donna , Olefsky Jerrold M
Published: September 2013

To evaluate kinetic defects in insulin action, we performed time-course studies during hyperinsulinemic (120 mU · m−2 · min−1) isoglycemic clamps in seven subjects with NIDDM (194 ± 29 mg/dl) and in seven lean and seven obese nondiabetic subjects. The time course of whole-body glucose disposal rate (GDR), leg glucose uptake (LGU), hepatic glucose output (HGO), and muscle insulin receptor tyrosine kinase (IRTK) activation were measured. The obese and NIDDM subjects had marked delays in activation of GDR (T50 74 ± 14 and 95 ± 15 min, respectively, compared with 33 ± 2 min in lean control subjects), arteriovenous glucose difference (T50 80 ± 12 and 109 ±31 min compared with 30 ± 3 min) and LGU (T50 89 ± 25 and 98 ± 27 min compared with 29 ± 4 min). All three measurements reached normal levels in the NIDDM group after 4–5 h of insulin infusion. Although only a limited number of data points could be obtained from serial muscle biopsies, no delay in the rate of activation of IRTK was apparent in the obese and NIDDM groups. In conclusion, 1) in obese and NIDDM subjects, insulin-mediated GDR and LGU are delayed to a similar degree; 2) mass action normalizes GDR and LGU in NIDDM, but only after several hours of insulin infusion; and 3) The kinetic defect in NIDDM and obesity most likely involves intracellular loci distal to activation of the insulin receptor kinase.
Augmentation of Hepatic Glucose Uptake by a Positive Glucose Gradient Between Hepatoportal and Central Nervous Systems
Authors: Matsuhisa Munehide , Morishima Toyohiko , Nakahara Itsuro , Tomita Tadahiro , Shiba Yuichi , Kubota Minoru , Wada Masahiki , Kanda Tsutomu , Kubota Masashi , Kawamori Ryuzo , Yamasaki Yoshimitsu
Published: September 2013

To determine the role of the glucose gradient between the hepatoportal system (HPS) and the central nervous system (CNS) in regulating hepatic glucose uptake, experiments were conducted with seven conscious dogs using a hepatic venous catheterization technique. With the infusion of somatostatin (0.8 µg · kg−1 · min−1), glucagon (0.65 ng · kg−1 · min−1), and insulin (27 pmol · kg−1 · min−1), arterial glucose levels could be maintained at 8 mmol/l by adjusting the intravenous glucose infusion (Ginf) according to the following three periods: 1) peripheral glucose infusion period (PE), Ginf alone; 2) portal glucose infusion period (PO), Ginf plus constant glucose infusion into the portal vein (GIRPV, 55.6 μmol · kg−1 · min−1); 3) portal and brain glucose infusion period (PO+CNS), Ginf and GIRPV plus additional glucose infusion into the unilateral carotid and vertebral arteries to abolish the positive glucose gradient between HPS and CNS. Arterial plasma glucose levels were clamped during the three periods (8.1 ± 0.1, PE; 8.2 ± 0.1, PO; 8.2 ± 0.1 mmol/l, PO+CNS). During PO, when a positive glucose gradient was promoted between HPS and CNS, the net hepatic glucose balance (NHGB) determined by the difference between hepatic glucose inflow and outflow was significantly lower than that of PE (−41.5 ± 5.3, PO vs. −7.5 ± 3.4 µmol · kg−1 · min−1, PE; P < 0.01). However, this decrease in the NHGB significantly increased during PO+CNS, when the glucose gradient between HPS and CNS was minimized, compared with PO (−21.7 ± 3.2 µmol · kg−1 · min−1 , P < 0.05). We conclude that a positive glucose gradient between HPS and CNS is an important regulatory factor of hepatic glucose uptake, but other factors also play important roles because minimizing the glucose gradient between HPS and CNS diminished the net hepatic glucose uptake by 50%.
Incorporation of [3-3H]Glucose and 2-[1-14C]Deoxyglucose Into Glycogen in Heart and Skeletal Muscle In Vivo: Implications for the Quantitation of Tissue Glucose Uptake
Authors: Virkamäki Antti , Rissanen Eeva , Hämäläinen Sari , Utriainen Tapio , Yki-Järvinen Hannele
Published: September 2013

2-deoxyglucose has been widely used to quantitate tissue glucose uptake in vivo, assuming that 2-deoxyglucose is transported and phosphorylated but not further metabolized. We examined the validity of this assumption by infusing [3-3H]glucose and 2-[1-14C]deoxyglucose in a similar primed continuous fashion to chronically catheterized, freely moving rats during normoglycemic hyperinsulinemic conditions. The rates of 2-deoxyglucose uptake were determined from the accumulation of 2-[1-14C]deoxyglucose-6-phosphate and 2-[1-14C]deoxyglucose-6-phosphate combined with the rate of the incorporation of 2-[1-14C]deoxyglucose into glycogen in rectus abdominis muscle and the heart. When the rates of glycogen synthesis during the 2-h hyperinsulinemic period from the two tracers were compared in rectus abdominis muscle, the rate of glycogen synthesis was twofold higher when measured with [3-3H]glucose (337 ± 14 µmol · kg−1 · min−1) than when measured with 2-[1-14C]deoxyglucose (166 ± 10 µmol · kg−1 · min−1, P < 0.001). In the heart, the rate of glycogen synthesis was twofold higher when measured with 2-[1-14C]deoxyglucose (141 ± 20 µmol · kg−1 · min−1) than when measured with [3-3H]glucose (72 ± 15 µmol · kg−1 · min−1, P < 0.001). The rate of 2-deoxyglucose uptake was 29% underestimated in rectus abdominis muscle, when counts found in glycogen were not included in glucose uptake calculations (398 ± 25 vs. 564 ± 25 µmol · kg−1 · min−1, P < 0.001). In the heart, glucose uptake was underestimated by 7% if glycogen counts were not taken into account (1,786 ± 278 vs. 1,926 ± 291 µmol · kg−1 dry · min−1, P < 0.05). The fraction of [3-3H]glucose incorporated into glycogen of total glucose metabolism (calculated from 2-deoxyglucose conversion to 2-deoxyglucose-6-phosphate and glycogen) was 0.6 (337/564) in rectus abdominis muscle and 0.037 (72/1,926) in the heart. We conclude that 2-deoxyglucose is incorporated into glycogen in the heart and in skeletal muscle in vivo under normoglycemic hyperinsulinemic conditions in the rat. Failure to consider the incorporation of 2-deoxyglucose into glycogen will underestimate the rate of tissue glucose uptake. To avoid such problems, the amount of 2-deoxyglucose incorporated into glycogen should be quantitated in subsequent studies.
Voluntary Wheel Running Decreases Adipose Tissue Mass and Expression of Leptin mRNA in Osborne-Mendel Rats
Authors: Zachwieja Jeffrey J , Hendry Stephen L , Smith Steven R , Harris Ruth B S
Published: September 2013

The purpose of this study was to assess the effects of voluntary wheel running on the expression of leptin mRNA in rats that are either sensitive (OM) or resistant (S5B/P1) to diet-induced obesity. Male OM and S5B/P1 rats had ad libitum access to standard rodent diet and water. At 3–5 weeks of age, animals of both strains were randomly assigned to either an exercise or sedentary control group. The exercise groups had 24-h access to a running wheel, and they trained for 7 weeks. During weeks 1–4, animals in both OM and S5B/P1 exercise groups progressively increased their running. During weeks 5–7, S5B/P1 exercisers tended to run more than did OM (∼60 vs. 45 km/week), but by the end of the study both groups had an equally greater heart weight (mg/g body weight) and planteris citrate synthase activity than their sedentary controls. Oral glucose tolerance tests performed during the last week of training revealed that compared with their appropriate controls, insulin sensitivity was enhanced (P < 0.05) in OM but not in the S5B/P1 wheel-running groups. Inguinal, epididymal, and retroperitoneal fat pads weighed less in the running than in the nonrunning groups of both strains (P < 0.01). Additionally, exercised animals had an increased percentage of smaller cells (40–60 µm; P < 0.05) and a decreased percentage of larger cells (120–160 µm; P < 0.05) in the epididymal fat depot. Epididymal leptin mRNA measured by Northern blot analysis was reduced in the exercise-trained rats of both strains (P < 0.05). Furthermore, serum leptin was reduced in exercise-trained compared with the control animals of both strains. In comparison to S5B/P1, control OM animals exhibited both a higher expression and higher circulating levels of leptin (P < 0.05). While serum leptin levels were decreased and food intake was increased in the exercise-trained animals of both strains (P < 0.05), the exact relationship between exercise, leptin, and food intake in this rat model of dietary obesity remains to be determined. Nonetheless, these results suggest that the expression and secretion of leptin can be influenced by exercise training and that these changes (i.e., reduced expression and secretion of protein) can occur independently of changes in whole-body insulin sensitivity and susceptibility to diet-induced obesity.
Neural Tube Defects in Embryos of Diabetic Mice: Role of the Pax-3 Gene and Apoptosis
Authors: Phelan Shelley A , Ito Moriko , Loeken Mary R
Published: September 2013

Neural tube defects are among the most common of the malformations associated with diabetic embryopathy. To study the molecular mechanisms by which neural tube defects occur during diabetic pregnancy, we have developed a new experimental system using pregnant diabetic mice. In this system, the rate of neural tube defects is about three times higher in embryos of diabetic mice than in embryos of nondiabetic mice. Most of the defects affected presumptive midbrain and hindbrain structures and included open defects (i.e., exencephaly) and gross maldevelopment. By semiquantitative reverse transcription–polymerase chain reaction and in situ hybridization, we found that expression of Pax-3, a gene required for neural tube closure in the area of the midbrain and hindbrain, is significantly reduced in the embryos of diabetic mice. The same regions of the neural tube where Pax-3 had been underexpressed were found subsequently to contain high concentrations of cells undergoing apoptosis. Reduced expression of Pax-3 appears to be responsible for this apoptosis because apoptotic cells were also found at sites of neural tube defects in embryos carrying null mutation of the Pax-3 gene. Finally, mouse strains that carry null mutations in Pax-3 develop neural tube defects that resemble the malformations that occur in embryos of diabetic mice. These results suggest that Pax-3 is an important developmental control gene, expression of which is disturbed in embryos of diabetic mice, and that as a consequence, apoptosis of the neural tube occurs. This pathway may be responsible for many of the neural tube defects resulting from diabetic pregnancy.
CL-316,243, a β3-Specific Adrenoceptor Agonist, Enhances Insulin-Stimulated Glucose Disposal in Nonobese Rats
Authors: de Souza Christopher J , Hirshman Michael F , Horton Edward S
Published: September 2013

Administration of the murine-selective β3 adrenoceptor agonist CL-316,243 corrects obesity and elevated blood glucose in diabetic rodents. This antiobesity effect is attributed to an increase in the thermogenic activity of brown adipose tissue (BAT). The antidiabetic effect is unknown, but has been attributed to the decline in body weight and plasma free fatty acids (FFAs). This study using the euglycemic-hyperinsulinemic clamp method was performed in nonobese, nondiabetic Sprague-Dawley rats fed normal rodent chow to determine if the β3 agonist could improve insulin sensitivity and/or responsiveness in the absence of weight loss or lowering of circulating FFAs. Subcutaneous miniosmotic pumps delivered either saline to control or 1 mg · kg−1 · day−1 of CL-316,243 for 10–12 days. Fed plasma glucose, insulin, and FFA levels were similar between the groups. Significant increases in food consumption, resting metabolic rates, and body core temperatures occurred, but only after 7 days of treatment. A 14% decrease in the respiratory quotient was also observed. Plasma glucose and insulin excursions in response to an oral glucose load (2 g/kg) on day 11 were unaltered. CL-316,243 treatment resulted in a decrease in abdominal and epididymal white fat pad weights, while interscapular brown adipose tissue (IBAT) weight doubled. Basal and insulin-stimulated whole-body glucose disposal rates were increased, while hepatic glucose output was suppressed to a greater extent in the CL-316,243 animals after 10 days of uninterrupted treatment. Chronic treatment with CL-316,243 resulted in an increase in basal and insulin-stimulated [3H]2-deoxy-glucose (2-DG) uptake by the retroperitoneal and epididymal white tissue and IBAT, but skeletal muscle 2-DG uptake under the same conditions was unaltered. These studies demonstrate that treatment with CL-316,243 improves basal and insulin-stimulated glucose disposal, and these effects occurred in the absence of a decrease in body weights and FFA concentrations. A particularly interesting observation was that the tissues responsible for this effect were white and brown adipose tissue, while skeletal muscle remained unaffected.
β-Cell Function in Normal Rats Made Chronically Hyperleptinemic by Adenovirus-Leptin Gene Therapy
Authors: Koyama Kazunori , Chen Guoxun , Wang May-Yun , Lee Young , Shimabukuro Michio , Newgard Christopher B , Unger Roger H
Published: September 2013

Leptin was overexpressed in the liver of normal Wistar rats by infusing recombinant adenovirus containing the cDNA encoding leptin. Plasma leptin levels rose to 12–24 ng/ml (vs. <2 ng/ml in control rats), and food intake and body weight fell. Visible fat disappeared within 7 days. Plasma insulin fell to <50% of normal in association with hypoglycemia, suggesting enhanced insulin sensitivity. Although β-cells appeared histologically normal, the pancreases were unresponsive to perfusion with stimulatory levels of glucose and arginine. Since islet triglyceride content was 0, compared with 14 ng/islet in pair-fed control rats, we coperfused a 2:1 oleate:palmitate mixture (0.5 mmol/l). This restored insulin responses to supranormal levels. When normal islets were cultured with 20 ng/ml of leptin, they too became triglyceride-depleted and failed to respond when perifused with glucose or arginine. Perifusion of fatty acids restored both responses. We conclude that in normal rats, hyperleptinemia for 2 weeks causes reversible β-cell dysfunction by depleting tissue lipids, thereby depriving β-cells of a lipid-derived signal required for the insulin response to other fuels.