bluuebunny/crossref_metadata_2025 huggingface dataset:
Glucose Transporters in Human Renal Proximal Tubular Cells Isolated From the Urine of Patients With Non–Insulin-Dependent Diabetes
Authors: Rahmoune Hassan , Thompson Paul W. , Ward Joanna M. , Smith Chari D. , Hong Guizhu , Brown John
Published: March 2007

The bulk of glucose that is filtered by the renal glomerulus is reabsorbed by the glucose transporters of the proximal convoluted tubular epithelium. However, it has been difficult to investigate this in diseases such as type 2 diabetes because of the inability to isolate primary renal cells from patients without a renal biopsy. We report here a method for the immunomagnetic isolation and novel primary culture of human exfoliated proximal tubular epithelial cells (HEPTECs) from fresh urine. The primary isolates are highly enriched and differentiated and express characteristic proximal tubular phenotypic markers. They continue to express the proximal tubular markers CD13/aminopeptidase-N, sodium glucose cotransporter (SGLT) 2, and alkaline phosphatase through up to six subsequent subcultures in a similar way to human proximal cells isolated from renal biopsies. In a hyperglycemic environment, HEPTECs isolated from patients with type 2 diabetes expressed significantly more SGLT2 and the facilitative glucose transporter GLUT2 than cells from healthy individuals. We also demonstrated a markedly increased renal glucose uptake in HEPTECs isolated from patients with type 2 diabetes compared with healthy control subjects. Our findings indicate for the first time in a human cellular model that increased renal glucose transporter expression and activity is associated with type 2 diabetes.
Molecular Characterization of New Selective Peroxisome Proliferator–Activated Receptor γ Modulators With Angiotensin Receptor Blocking Activity
Authors: Schupp Michael , Clemenz Markus , Gineste Romain , Witt Henning , Janke Jürgen , Helleboid Stephane , Hennuyer Nathalie , Ruiz Patricia , Unger Thomas , Staels Bart , Kintscher Ulrich
Published: March 2007

Selective peroxisome proliferator–activated receptor (PPAR) γ modulation is a new pharmacological approach that, based on selective receptor-cofactor interactions and target gene regulation, should result in potent insulin sensitization in the absence of PPARγ-mediated adverse effects. Here, we characterize two angiotensin receptor blockers (ARBs), telmisartan and irbesartan, as new selective PPAR modulators (SPPARMs). Analysis of PPARγ protein conformation using protease protection showed that telmisartan directly interacts with the receptor, producing a distinct conformational change compared with a glitazone. Glutathione S-transferase pull-down and fluorescence resonance energy transfer assays revealed selective cofactor binding by the ARBs compared with glitazones with an attenuated release of the nuclear receptor corepressor and absence of transcriptional intermediary factor 2 recruitment by ARBs. Consistently, selective cofactor binding resulted in differential gene expression profiles in adipocytes (ARB versus glitazone treated) assessed by oligo microarray analysis. Finally, telmisartan improved insulin sensitivity in diet-induced obese mice in the absence of weight gain. The present study identifies two ARBs as new SPPARMs. SPPARM activity by ARBs could retain the metabolic efficacy of PPARγ activation with reduction in adverse effects exerting in parallel AT1 receptor blockade. This may provide a new therapeutic option for better cardiovascular risk management in metabolic diseases and may initiate the development of new classes of drugs combining potent antihypertensive and antidiabetic actions.
Changes in Exercise-Induced Gene Expression in 5′-AMP–Activated Protein Kinase γ3–Null and γ3 R225Q Transgenic Mice
Authors: Barnes Brian R. , Long Yun Chau , Steiler Tatiana L. , Leng Ying , Galuska Dana , Wojtaszewski Jørgen F.P. , Andersson Leif , Zierath Juleen R.
Published: March 2007

5′-AMP–activated protein kinase (AMPK) is important for metabolic sensing. We used AMPKγ3 mutant–overexpressing Tg-Prkag3225Q and AMPKγ3-knockout Prkag3−/− mice to determine the role of the AMPKγ3 isoform in exercise-induced metabolic and gene regulatory responses in skeletal muscle. Mice were studied after 2 h swimming or 2.5 h recovery. Exercise increased basal and insulin-stimulated glucose transport, with similar responses among genotypes. In Tg-Prkag3225Q mice, acetyl-CoA carboxylase (ACC) phosphorylation was increased and triglyceride content was reduced after exercise, suggesting that this mutation promotes greater reliance on lipid oxidation. In contrast, ACC phosphorylation and triglyceride content was similar between wild-type and Prkag3−/− mice. Expression of genes involved in lipid and glucose metabolism was altered by genetic modification of AMPKγ3. Expression of lipoprotein lipase 1, carnitine palmitoyl transferase 1b, and 3-hydroxyacyl–CoA dehydrogenase was increased in Tg-Prkag3225Q mice, with opposing effects in Prkag3−/− mice after exercise. GLUT4, hexokinase II (HKII), and glycogen synthase mRNA expression was increased in Tg-Prkag3225Q mice after exercise. GLUT4 and HKII mRNA expression was increased in wild-type mice and blunted in Prkag3−/− mice after recovery. In conclusion, the Prkag3225Q mutation, rather than presence of a functional AMPKγ3 isoform, directly promotes metabolic and gene regulatory responses along lipid oxidative pathways in skeletal muscle after endurance exercise.
Polymorphisms in igf-Binding Protein 1 Are Associated With Impaired Renal Function in Type 2 Diabetes
Authors: Stephens Richard H. , McElduff Patrick , Heald Adrian H. , New John P. , Worthington Jane , Ollier William E. , Gibson J. Martin
Published: March 2007

The dysregulation of the IGF system has been implicated in the pathogenesis of obesity, diabetes, and diabetes complications such as nephropathy, but little is known about the genomics of the IGF system in health and disease. We genotyped 13 single nucleotide polymorphisms (SNPs) in IGFBP1 gene in 732 representative type 2 diabetic patients from the Salford Diabetes Register. Of the 13 SNPs, 8 were polymorphic and 7 of those had minor allele frequencies >0.1, one of which was in the gene promoter and one of which was nonsynonymous in exon 4. The minor alleles of these SNPs and two others were associated with a reduced prevalence of diabetic nephropathy. Haplotype analysis revealed that 97% of the genetic variation for IGFBP1 in the population sample could be accounted for using two of the “reno-protective” SNPs, with other SNPs adding little extra information. One of these two SNPs was the nonsynonymous mutation in exon 4, lying close to the integrin-binding RGD motif, which is thought to affect tissue delivery of IGF-I by IGF-binding protein 1 (IGFBP-1), possibly suggesting a “reno-protective” effect via altered IGFBP-1 binding. In conclusion, we have described the first genomic markers to be associated with diabetic microvascular complications within the human IGFBP1 gene.
A Functional Variant in the Human Betacellulin Gene Promoter Is Associated With Type 2 Diabetes
Authors: Nakano Yoshio , Furuta Hiroto , Doi Asako , Matsuno Shohei , Nakagawa Takayuki , Shimomura Hiroko , Sakagashira Setsuya , Horikawa Yukio , Nishi Masahiro , Sasaki Hideyuki , Sanke Tokio , Nanjo Kishio
Published: March 2007

Betacellulin (BTC) plays an important role in differentiation, growth, and antiapoptosis of pancreatic β-cells. We characterized about 2.3 kb of the 5′-flanking region of human BTC gene and identified six polymorphisms (−2159A>G, −1449G>A, −1388C>T, −279C>A, −233G>C, and −226A>G). The G allele in the −226A>G polymorphism was more frequent in type 2 diabetic patients (n = 250) than in nondiabetic subjects (n = 254) (35.6% vs. 27.8%, P = 0.007), and the −2159G, −1449A, and −1388T alleles were in complete linkage disequilibrium with the −226G allele. The frequencies of the −279A and −233C alleles were low (7.0 and 2.0% in diabetic patients), and no significant differences were observed. In the diabetic group, insulin secretion ability, assessed by the serum C-peptide response to intravenous glucagon stimulation, was lower in patients with the −226G allele (G/G, 2.96 ± 0.16 ng/ml; G/A, 3.65 ± 0.18 ng/ml; A/A, 3.99 ± 0.16 ng/ml at 5 min after stimulation; P = 0.008). Furthermore, in vitro functional analyses indicated that both the −226G and the −233C alleles caused an ∼50% decrease in the promoter activity, but no effects of the −2159A>G, −1449G>A, −1388C>T, and −279C>A polymorphisms were observed. These results suggest that the −226A/G polymorphism of the BTC gene may contribute to the development of diabetes.
Retinoic Acid Induces Pdx1-Positive Endoderm in Differentiating Mouse Embryonic Stem Cells
Authors: Micallef Suzanne J. , Janes Mary E. , Knezevic Kathy , Davis Richard P. , Elefanty Andrew G. , Stanley Edouard G.
Published: March 2007

We have generated an embryonic stem (ES) cell line in which sequences encoding green fluorescent protein (GFP) were targeted to the locus of the pancreatic-duodenal homeobox gene (Pdx1). Analysis of chimeric embryos derived from blastocyst injection of Pdx1GFP/w ES cells demonstrated that the pattern of GFP expression was consistent with that reported for the endogenous Pdx1 gene. By monitoring GFP expression during the course of ES cell differentiation, we have shown that retinoic acid (RA) can regulate the commitment of ES cells to form Pdx1+ pancreatic endoderm. RA was most effective at inducing Pdx1 expression when added to cultures at day 4 of ES differentiation, a period corresponding to the end of gastrulation in the embryo. RT-PCR analysis showed that Pdx1-positive cells from day 8 cultures expressed the early endoderm markers Ptf1a, Foxa2, Hnf4α, Hnf1β, and Hnf6, consistent with the notion that they corresponded to the early pancreatic endoderm present in the embryonic day 9.5 mouse embryo. These results demonstrate the utility of Pdx1GFP/w ES cells as a tool for monitoring the effects of factors that influence pancreatic differentiation from ES cells.
Antigen-Specific FoxP3-Transduced T-Cells Can Control Established Type 1 Diabetes
Authors: Jaeckel Elmar , von Boehmer Harald , Manns Michael P.
Published: March 2007

CD4+CD25+ T-cells can be used to interfere with spontaneous autoimmune diseases such as type 1 diabetes. However, their low frequency and often unknown specificity represent major obstacles to their therapeutic use. Here we have explored the fact that ectopic expression of the transcription factor Foxp3 can confer a suppressor phenotype to naïve CD4+ T-cells. We found that retroviral transduction of polyclonal CD4 T-cells with FoxP3 was not effective in interfering with established type 1 diabetes. Thus, more subtle and more organ-specific regulation might be required to prevent type 1 diabetes, as well as to avoid systemic immunosuppression. However, a single injection of 105 FoxP3-transduced T-cells with specificity for islet antigen stabilized and reversed disease in mice with recent-onset diabetes. By comparing FoxP3-transduced T-cells with various antigen specificities, it became clear that the in vivo effect correlated with specific homing to and activation in pancreatic lymph nodes and not with in vitro suppressor activity or cytokine production. Our results complement recent results on in vitro–amplified antigen-specific T-cells in ameliorating type 1 diabetes and suggest that FoxP3 transduction of expanded T-cells might achieve the same goal.
Redox Paradox
Authors: Goldstein Barry J. , Mahadev Kalyankar , Wu Xiangdong
Published: March 2007

Propelled by the identification of a small family of NADPH oxidase (Nox) enzyme homologs that produce superoxide in response to cellular stimulation with various growth factors, renewed interest has been generated in characterizing the signaling effects of reactive oxygen species (ROS) in relation to insulin action. Two key observations made >30 years ago—that oxidants can facilitate or mimic insulin action and that H2O2 is generated in response to insulin stimulation of its target cells—have led to the hypothesis that ROS may serve as second messengers in the insulin action cascade. Specific molecular targets of insulin-induced ROS include enzymes whose signaling activity is modified via oxidative biochemical reactions, leading to enhanced insulin signal transduction. These positive responses to cellular ROS may seem “paradoxical” because chronic exposure to relatively high levels of ROS have also been associated with functional β-cell impairment and the chronic complications of diabetes. The best-characterized molecular targets of ROS are the protein-tyrosine phosphatases (PTPs) because these important signaling enzymes require a reduced form of a critical cysteine residue for catalytic activity. PTPs normally serve as negative regulators of insulin action via the dephosphorylation of the insulin receptor and its tyrosine-phosphorylated cellular substrates. However, ROS can rapidly oxidize the catalytic cysteine of target PTPs, effectively blocking their enzyme activity and reversing their inhibitory effect on insulin signaling. Among the cloned Nox homologs, we have recently provided evidence that Nox4 may mediate the insulin-stimulated generation of cellular ROS and is coupled to insulin action via the oxidative inhibition of PTP1B, a PTP known to be a major regulator of the insulin signaling cascade. Further characterization of the molecular components of this novel signaling cascade, including the mechanism of ROS generated by insulin and the identification of various oxidation-sensitive signaling targets in insulin-sensitive cells, may provide a novel means of facilitating insulin action in states of insulin resistance.
5-Aminoimidazole-4-Carboxamide-1-β-d-Ribofuranoside Causes Acute Hepatic Insulin Resistance In Vivo
Authors: Pencek R. Richard , Shearer Jane , Camacho Raul C. , James Freyja D. , Lacy D. Brooks , Fueger Patrick T. , Donahue E. Patrick , Snead Wanda , Wasserman David H.
Published: March 2007

The infusion of 5-aminoimidazole-4-carboxamide-1-β-d-ribofuranoside (AICAR) causes a rise in tissue concentrations of the AMP analog 5-aminoimidazole-4-carboxamide-1-β-d-ribofuranotide (ZMP), which mimics an elevation of cellular AMP levels. The purpose of this work was to determine the effect of raising hepatic ZMP levels on hepatic insulin action in vivo. Dogs had sampling and infusion catheters as well as flow probes implanted 16 days before an experiment. After an 18-h fast, blood glucose was 82 ± 1 mg/dl and basal net hepatic glucose output 1.5 ± 0.2 mg · kg−1 · min−1. Dogs received portal venous glucose (3.2 mg · kg−1 · min−1), peripheral venous somatostatin, and basal portal venous glucagon infusions from −90 to 60 min. Physiological hyperinsulinemia was established with a portal insulin infusion (1.2 mU · kg−1 · min−1). Peripheral venous glucose infusion was used to clamp arterial blood glucose at 150 mg/dl. Starting at t = 0 min, dogs received portal venous AICAR infusions of 0, 1, or 2 mg · kg−1 · min−1. Net hepatic glucose uptake was 2.4 ± 0.5 mg · kg−1 · min−1 (mean of all groups) before t = 0 min. In the absence of AICAR, net hepatic glucose uptake was 1.9 ± 0.4 mg · kg−1 · min−1 at t = 60 min. The lower-dose AICAR infusion caused a complete suppression of net hepatic glucose uptake (−1.0 ± 1.7 mg · kg−1 · min−1 at t = 60 min). The higher AICAR dose resulted in a profound shift in hepatic glucose balance from net uptake to a marked net output (−6.1 ± 1.9 mg · kg−1 · min−1 at t = 60 min), even in the face of hyperglycemia and hyperinsulinemia. These data show that elevations in hepatic ZMP concentrations, induced by portal venous AICAR infusion, cause acute hepatic insulin resistance. These findings have important implications for the targeting of AMP kinase for the treatment of insulin resistance, using AMP analogs.
The Extracellular Signal–Regulated Kinase Isoform ERK1 Is Specifically Required for In Vitro and In Vivo Adipogenesis
Authors: Bost Frédéric , Aouadi Myriam , Caron Leslie , Even Patrick , Belmonte Nathalie , Prot Matthieu , Dani Christian , Hofman Paul , Pagès Gilles , Pouysségur Jacques , Le Marchand-Brustel Yannick , Binétruy Bernard
Published: March 2007

Hyperplasia of adipose tissue is critical for the development of obesity, but molecular mechanisms governing normal or pathological recruitment of new adipocytes remain unclear. The extracellular signal–regulated kinase (ERK) pathway plays a pivotal role in many essential cellular functions, such as proliferation and differentiation. Using ERK1−/− mice, we investigated the role of this isoform in adipose tissue development. Mice lacking ERK1 have decreased adiposity and fewer adipocytes than wild-type animals. Furthermore, ERK1−/− mice challenged with high-fat diet are resistant to obesity, are protected from insulin resistance, and have a higher postprandial metabolic rate. To get insights into cellular mechanisms implicated in reduced adiposity in ERK1−/− animals, we analyzed adipocyte differentiation in ERK1−/− cells. Compared with wild-type control cells, mouse embryo fibroblasts and cultures of adult preadipocytes isolated from ERK1−/− adult animals exhibit impaired adipogenesis. An inhibitor of the ERK pathway does not affect the residual adipogenesis of the ERK1−/− cells, suggesting that ERK2 is not implicated in adipocyte differentiation. Our results clearly link ERK1 to the regulation of adipocyte differentiation, adiposity, and high-fat diet–induced obesity. This suggests that a therapeutic approach of obesity targeting specifically the ERK1 isoform and not ERK2 would be of particular interest.
Autoimmune Diabetes Is Suppressed by Transfer of Proinsulin-Encoding Gr-1+ Myeloid Progenitor Cells That Differentiate In Vivo Into Resting Dendritic Cells
Authors: Steptoe Raymond J. , Ritchie Janine M. , Jones Lynelle K. , Harrison Leonard C.
Published: March 2007

The nature of the T-cell response to antigen is governed by the activation state of the antigen-presenting dendritic cell (DC). Immature or resting DCs have been shown to induce T-cell responses that may protect against the development of autoimmune disease. Effectively harnessing this “tolerogenic” effect of resting DCs requires that it be disease-specific and that activation of DCs by manipulation ex vivo is avoided. We reasoned that this could be achieved by transferring in vivo partially differentiated myeloid progenitor cells encoding a disease-specific autoantigen. With the aim of preventing autoimmune diabetes, we transferred myeloid progenitor cells encoding proinsulin into NOD mice. Bone marrow (BM) was cultured in granulocyte macrophage colony-stimulating factor (GM-CSF) and transforming growth factor-β1, a cytokine combination that expands myeloid cells but inhibits terminal DC differentiation, to yield Gr-1+/CD11b+/CD11c− myeloid progenitor cells and a minor population of CD11c+/CD11b+/CD86lo immature DCs. After transfer, Gr-1+ myeloid cells acquired the characteristics of resting DCs (CD11c+/MHC classIIint/CD86lo/CD40lo). Gr-1+ myeloid cells generated from transgenic NOD mice that expressed proinsulin controlled by a major histocompatibility complex (MHC) class II promoter, but not from wild-type NOD mice, transferred into 4-week-old female NOD mice significantly suppressed diabetes development. The transfer of DC progenitors encoding a disease-specific autoantigen is, therefore, an effective immunotherapeutic strategy that could be applied to humans.
Cytokines Downregulate the Sarcoendoplasmic Reticulum Pump Ca2+ ATPase 2b and Deplete Endoplasmic Reticulum Ca2+, Leading to Induction of Endoplasmic Reticulum Stress in Pancreatic β-Cells
Authors: Cardozo Alessandra K. , Ortis Fernanda , Storling Joachim , Feng Ying-Mei , Rasschaert Joanne , Tonnesen Morten , Van Eylen Françoise , Mandrup-Poulsen Thomas , Herchuelz André , Eizirik Décio L.
Published: March 2007

Cytokines and free radicals are mediators of β-cell death in type 1 diabetes. Under in vitro conditions, interleukin-1β (IL-1β) + γ-interferon (IFN-γ) induce nitric oxide (NO) production and apoptosis in rodent and human pancreatic β-cells. We have previously shown, by microarray analysis of primary β-cells, that IL-1β + IFN-γ decrease expression of the mRNA encoding for the sarcoendoplasmic reticulum pump Ca2+ ATPase 2b (SERCA2b) while inducing expression of the endoplasmic reticulum stress–related and proapoptotic gene CHOP (C/EBP [CCAAT/enhancer binding protein] homologous protein). In the present study we show that cytokine-induced apoptosis and necrosis in primary rat β-cells and INS-1E cells largely depends on NO production. IL-1β + IFN-γ, via NO synthesis, markedly decreased SERCA2b protein expression and depleted ER Ca2+ stores. Of note, β-cells showed marked sensitivity to apoptosis induced by SERCA blockers, as compared with fibroblasts. Cytokine-induced ER Ca2+ depletion was paralleled by an NO-dependent induction of CHOP protein and activation of diverse components of the ER stress response, including activation of inositol-requiring ER-to-nucleus signal kinase 1α (IRE1α) and PRK (RNA-dependent protein kinase)-like ER kinase (PERK)/activating transcription factor 4 (ATF4), but not ATF6. In contrast, the ER stress–inducing agent thapsigargin triggered these four pathways in parallel. In conclusion, our results suggest that the IL-1β + IFN-γ–induced decrease in SERCA2b expression, with subsequent depletion of ER Ca2+ and activation of the ER stress pathway, is a potential contributory mechanism to β-cell death.
Alteration of the Malonyl-CoA/Carnitine Palmitoyltransferase I Interaction in the β-Cell Impairs Glucose-Induced Insulin Secretion
Authors: Herrero Laura , Rubí Blanca , Sebastián David , Serra Dolors , Asins Guillermina , Maechler Pierre , Prentki Marc , Hegardt Fausto G.
Published: March 2007

Carnitine palmitoyltransferase I, which is expressed in the pancreas as the liver isoform (LCPTI), catalyzes the rate-limiting step in the transport of fatty acids into the mitochondria for their oxidation. Malonyl-CoA derived from glucose metabolism regulates fatty acid oxidation by inhibiting LCPTI. To examine directly whether the availability of long-chain fatty acyl-CoA (LC-CoA) affects the regulation of insulin secretion in the β-cell and whether malonyl-CoA may act as a metabolic coupling factor in the β-cell, we infected INS(832/13) cells and rat islets with an adenovirus encoding a mutant form of LCPTI (Ad-LCPTI M593S) that is insensitive to malonyl-CoA. In Ad-LCPTI M593S–infected INS(832/13) cells, LCPTI activity increased sixfold. This was associated with enhanced fatty acid oxidation, at any glucose concentration, and a 60% suppression of glucose-stimulated insulin secretion (GSIS). In isolated rat islets in which LCPTI M593S was overexpressed, GSIS decreased 40%. The impairment of GSIS in Ad-LCPTI M593S–infected INS(832/13) cells was not recovered when cells were incubated with 0.25 mmol/l palmitate, indicating the deep metabolic influence of a nonregulated fatty acid oxidation system. At high glucose concentration, overexpression of a malonyl-CoA–insensitive form of LCPTI reduced partitioning of exogenous palmitate into lipid esterification products and decreased protein kinase C activation. Moreover, LCPTI M593S expression impaired KATP channel–independent GSIS in INS(832/13) cells. The LCPTI M593S mutant caused more pronounced alterations in GSIS and lipid partitioning (fat oxidation, esterification, and the level of nonesterified palmitate) than LCPTI wt in INS(832/13) cells that were transduced with these constructs. The results provide direct support for the hypothesis that the malonyl-CoA/CPTI interaction is a component of a metabolic signaling network that controls insulin secretion.
Hypoglycemia Induces Transient Neurogenesis and Subsequent Progenitor Cell Loss in the Rat Hippocampus
Authors: Suh Sang Won , Fan Yang , Hong Shwuhuey M. , Liu Zhengyan , Matsumori Yasuhiko , Weinstein Philip R. , Swanson Raymond A. , Liu Jialing
Published: March 2007

Neurogenesis after brain injury not only leads to the replacement of damaged cells but might also contribute to functional recovery, suggesting the possibility of endogenous neural repair. We investigated the extent of hippocampal neural regeneration in a rat model of hypoglycemia. Two weeks after 30 min of insulin-induced isoelectric electroencephalogram, extensive neuronal loss was observed in the hippocampus, including area CA1 and dentate gyrus (DG). A transient increase in progenitor cell proliferation in the DG subgranular zone (SGZ) was detected, leading to an increase of immature neuroblasts 1–2 weeks after hypoglycemic insult. Most of the surviving newborn cells assumed a neuronal phenotype within 1 month in DG, a few cells near the site of granule-cell death becoming astroglia or microglia. No neuronal regeneration was observed in the CA1 after hypoglycemia, although dividing cells appeared to be astroglia or microglia in CA1 and dentate hilus. At 4 weeks after hypoglycemia, proliferative activity in the SGZ diminished below baseline in experimental versus control rats, with a subsequent reduction of neuroblasts. Morphological findings (doublecortin staining) suggest permanent progenitor cell loss in some areas of SGZ. Reduced neurogenesis in DG and lack of neuronal regeneration in CA1 may impede cognitive recovery after severe hypoglycemia injury.
Bumetanide Reduces Cerebral Edema Formation in Rats With Diabetic Ketoacidosis
Authors: Lam Tina I. , Anderson Steven E. , Glaser Nicole , O’Donnell Martha E.
Published: March 2007

The mechanisms responsible for cerebral edema formation in diabetic ketoacidosis (DKA) are not well understood, although evidence suggests ischemia as a contributing factor. Previous studies have shown that the Na-K-Cl cotransporter of cerebral microvascular endothelial cells and astrocytes is a major participant in ischemia-induced cerebral edema in stroke. The present study was conducted to test the hypothesis that the Na-K-Cl cotransporter also contributes to cerebral edema in DKA. Sprague-Dawley rats were administered streptozotocin to induce DKA, and then cerebral edema was assessed by determination of apparent diffusion coefficients (ADC) with magnetic resonance diffusion-weighted imaging. Cerebral ADC values in DKA rats were significantly reduced in both cortex and striatum compared with non-DKA control rats, indicating the presence of cerebral edema. Intravenous administration of bumetanide to DKA rats abolished the drop in cortical ADC values, while having no significant effect in the striatum. Insulin and saline treatment had no effect when given after bumetanide but increased both cortical and striatal ADC values when given before bumetanide. Evidence is also presented here that acetoacetate and β-hydroxybutyrate stimulate brain microvascular Na-K-Cl cotransporter activity. These findings suggest that the Na-K-Cl cotransporter contributes to brain edema in DKA.
Paradoxical Effects of Green Tea (Camellia Sinensis) and Antioxidant Vitamins in Diabetic Rats
Authors: Mustata Georgian T. , Rosca Mariana , Biemel Klaus M. , Reihl Oliver , Smith Mark A. , Viswanathan Ashwini , Strauch Christopher , Du Yunpeng , Tang Jie , Kern Timothy S. , Lederer Markus O. , Brownlee Michael , Weiss Miriam F. , Monnier Vincent M.
Published: March 2007

We tested the hypothesis that green tea prevents diabetes-related tissue dysfunctions attributable to oxidation. Diabetic rats were treated daily with tap water, vitamins C and E, or fresh Japanese green tea extract. After 12 months, body weights were decreased, whereas glycated lysine in aorta, tendon, and plasma were increased by diabetes (P < 0.001) but unaffected by treatment. Erythrocyte glutathione and plasma hydroperoxides were improved by the vitamins (P < 0.05) and green tea (P < 0.001). Retinal superoxide production, acellular capillaries, and pericyte ghosts were increased by diabetes (P < 0.001) and improved by green tea and the vitamins (P variable). Lens crystallin fluorescence at 370/440 nm was ameliorated by green tea (P < 0.05) but not the vitamins. Marginal effects on nephropathy parameters were noted. However, suppressed renal mitochondrial NADH-linked ADP-dependent and dinitrophenol-dependent respiration and complex III activity were improved by green tea (P variable). Green tea also suppressed the methylglyoxal hydroimidazolone immunostaining of a 28-kDa mitochondrial protein. Surprising, glycoxidation in tendon, aorta, and plasma was either worsened or not significantly improved by the vitamins and green tea. Glucosepane cross-links were increased by diabetes (P < 0.001), and green tea worsened total cross-linking. In conclusion, green tea and antioxidant vitamins improved several diabetes-related cellular dysfunctions but worsened matrix glycoxidation in selected tissues, suggesting that antioxidant treatment tilts the balance from oxidative to carbonyl stress in the extracellular compartment.
Adiponectin and Future Coronary Heart Disease Events Among Men With Type 2 Diabetes
Authors: Schulze Matthias B. , Shai Iris , Rimm Eric B. , Li Tricia , Rifai Nader , Hu Frank B.
Published: March 2007

Adiponectin, predominantly synthesized in the adipose tissue, seems to have substantial anti-inflammatory properties and to be a major modulator of insulin resistance and dyslipidemia, mechanisms that are associated with an increased atherosclerotic risk in diabetic patients. However, it is unknown whether higher levels of adiponectin are associated with a reduced risk for coronary heart disease (CHD) among diabetic individuals. We investigated the association between plasma adiponectin levels and incidence of CHD among 745 men with confirmed type 2 diabetes in the Health Professionals Follow-up Study. Participants were aged 46–81 years and were free of diagnosed cardiovascular disease at the time of blood draw in 1993/1994. During an average of 5 years of follow-up (3,980 person-years), we identified 89 incident cases of CHD (19 myocardial infarction and 70 coronary artery bypass surgery), confirmed by medical records. Levels of adiponectin were inversely associated with BMI and directly associated with age, alcohol intake, and duration of diabetes (P < 0.05). After adjustment for age, BMI, smoking, alcohol consumption, duration of diabetes, and other lifestyle factors, adiponectin was associated with a decreased risk for CHD events. The multivariate relative risk for CHD for a doubling of adiponectin was 0.71 (95% CI 0.53–0.95). Further adjustment for HDL cholesterol attenuated this association (0.78 [0.57–1.06]). The inverse association between adiponectin and CHD was consistent across strata of aspirin use, family history of myocardial infarction, alcohol consumption, insulin use, duration of diabetes, and levels of HbA1c, triglycerides, C-reactive protein, and HDL cholesterol. Our study suggests that increased adiponectin levels are associated with a moderately decreased CHD risk in diabetic men. This association seems to be mediated in part by effects of adiponectin on HDL cholesterol levels.
Peroxisome Proliferator-Activated Receptor α Gene Variation Influences Age of Onset and Progression of Type 2 Diabetes
Authors: Flavell David M. , Ireland Helen , Stephens Jeffrey W. , Hawe Emma , Acharya Jay , Mather Hugh , Hurel Steven J. , Humphries Steve E.
Published: March 2007

Dysregulation of fatty acid metabolism is important in the pathogenesis of type 2 diabetes. Peroxisome proliferator-activated receptor (PPAR)α is a master regulator of fatty acid catabolism, and PPARα activators delay the onset of type 2 diabetes. We examined association between three PPARα gene polymorphisms (an A→C variant in intron 1, the L162V variant, and the intron 7 G→C variant) and age at diagnosis of type 2 diabetes in 912 Caucasian type 2 diabetic subjects. Individually, PPARα gene variants did not influence age at diagnosis, but in combination, the rare alleles of both the intron 1 A→C (P < 0.001) and intron 7 G→C (P = 0.025) variants synergistically lowered age at diagnosis (interaction P < 0.001). Overall, the PPARα haplotype signficantly influenced age at diagnosis (P = 0.027), with the C-L-C and C-V-C haplotypes (intron 1-L162V-intron 7) accelerating onset of diabetes by 5.9 (P = 0.02) and 10 (P = 0.03) years, respectively, as compared with the common A-L-G haplotype, and was associated with an odds ratio for early-onset diabetes (age at diagnosis ≤45 years) of 3.75 (95% CI 1.65–8.56, P = 0.002). Intron 1 C-allele carriers also progressed more rapidly to insulin monotherapy (AA 9.4 ± 1.5 and AC + CC 5.3 ± 1.1 years, P = 0.002). These data indicate that PPARα gene variation influences the onset and progression of type 2 diabetes.
A Role for Sphingolipids in Producing the Common Features of Type 2 Diabetes, Metabolic Syndrome X, and Cushing’s Syndrome
Authors: Summers Scott A. , Nelson Don H.
Published: March 2007

Metabolic syndrome X and type 2 diabetes share many metabolic and morphological similarities with Cushing’s syndrome, a rare disorder caused by systemic glucocorticoid excess. Pathologies frequently associated with these diseases include insulin resistance, atherosclerosis, susceptibility to infection, poor wound healing, and hypertension. The similarity of the clinical profiles associated with these disorders suggests the influence of a common molecular mechanism for disease onset. Interestingly, numerous studies identify ceramides and other sphingolipids as potential contributors to these sequelae. Herein we review studies demonstrating that aberrant ceramide accumulation contributes to the development of the deleterious clinical manifestations associated with these diseases.
Munc18c Heterozygous Knockout Mice Display Increased Susceptibility for Severe Glucose Intolerance
Authors: Oh Eunjin , Spurlin Beth A. , Pessin Jeffrey E. , Thurmond Debbie C.
Published: March 2007

The disruption of Munc18c binding to syntaxin 4 impairs insulin-stimulated GLUT4 vesicle translocation in 3T3L1 adipocytes. To investigate the physiological function and requirement for Munc18c in the regulation of GLUT4 translocation and glucose homeostasis in vivo, we used homologous recombination to generate Munc18c-knockout (KO) mice. Homozygotic disruption of the Munc18c gene resulted in early embryonic lethality, whereas heterozygous KO mice (Munc18c−/+) had normal viability. Munc18c−/+ mice displayed significantly decreased insulin sensitivity in an insulin tolerance test and a >50% reduction in skeletal muscle insulin-stimulated GLUT4 translocation when compared with wild-type (WT) mice. Furthermore, glucose-stimulated insulin secretion was significantly reduced in islets isolated from Munc18c−/+ mice compared with those from WT mice. Despite the defects in insulin action and secretion, Munc18c−/+ mice demonstrated the ability to clear glucose to the same level as WT mice in a glucose tolerance test when fed a normal diet. However, after consuming a high-fat diet for only 5 weeks, the Munc18c−/+ mice manifested severely impaired glucose tolerance compared with high-fat−fed WT mice. Taken together, these data suggest that the reduction of Munc18c protein in the Munc18c−/+ mice results in impaired insulin sensitivity with a latent increased susceptibility for developing severe glucose intolerance in response to environmental perturbations such as intake of a high-calorie diet rich in fat and carbohydrate.
Long-Term Graft Function of Adult Rat and Human Islets Encapsulated in Novel Alginate-Based Microcapsules After Transplantation in Immunocompetent Diabetic Mice
Authors: Schneider Stephan , Feilen Peter J. , Brunnenmeier Frank , Minnemann Timo , Zimmermann Heiko , Zimmermann Ulrich , Weber Matthias M.
Published: March 2007

We describe the results of the first study to show that adult rat and human islets can be protected against xenogenic rejection in immunocompetent diabetic mice by encapsulating them in a novel alginate-based microcapsule system with no additional permselective membrane. Nonencapsulated islets lost function within 4–8 days after being transplanted into diabetic Balb/c mice, whereas transplanted encapsulated adult rat or human islets resulted in normoglycemia for >7 months. When rat islet grafts were removed 10 and 36 weeks after transplantation, the mice became immediately hyperglycemic, thus demonstrating the efficacy of the encapsulated islets. The explanted capsules showed only a mild cellular reaction on their surface and a viability of >85%, and responded to a glucose stimulus with a 10-fold increase in insulin secretion. Furthermore, transplanted mice showed a slight decrease in the glucose clearance rate in response to intraperitoneal glucose tolerance tests 3–16 weeks after transplantation; after 16 weeks, the rate remained stable. Similar results were obtained for encapsulated human islets. Thus we provide the first evidence of successful transplantation of microencapsulated human islets. In conclusion, we have developed a novel microcapsule system that enables survival and function of adult rat and human islets in immunocompetent mice without immunosuppression for >7 months.
Overexpression of Cyclin D1 in Pancreatic β-Cells In Vivo Results in Islet Hyperplasia Without Hypoglycemia
Authors: Zhang Xiaoboo , Gaspard John P. , Mizukami Yusuke , Li Jingnan , Graeme-Cook Fiona , Chung Daniel C.
Published: March 2007

Cyclin D1 can stimulate proliferation by driving cells from the G1 into the S-phase of the mammalian cell cycle. Previous animal studies have implicated the G1-S transition as a key regulatory checkpoint governing the proliferation of pancreatic islet cells. We expressed cyclin D1 in the β-cells of mice and islet hyperplasia developed in a time-dependent manner. The hyperplastic β-cells exhibited higher rates of proliferation. However, blood glucose levels in fasting as well as nonfasting conditions remained normal. Furthermore, glucose tolerance tests demonstrated nearly normal responses, and diabetes did not develop in any of the animals. No islet cell tumors were observed, even among animals >2 years of age. Under our experimental conditions, the proliferative stimulus provided by cyclin D1 is not tumorigenic, does not result in diabetes, and does not result in hypoglycemia. Cyclin D1 may thus be considered a potential candidate to augment the β-cell population ex vivo as a prelude to islet transplantation for diabetes.
TAT-Mediated Neurogenin 3 Protein Transduction Stimulates Pancreatic Endocrine Differentiation In Vitro
Authors: Domínguez-Bendala Juan , Klein Dagmar , Ribeiro Melina , Ricordi Camillo , Inverardi Luca , Pastori Ricardo , Edlund Helena
Published: March 2007

Stem cell technologies hold great potential for the treatment of type 1 diabetes, provided that functional transplantable β-cells can be selectively generated in an efficient manner. Such a process should recapitulate, at least to a certain extent, the embryonic development of β-cells in vitro. However, progress at identifying the transcription factors involved in β-cell development has not been accompanied by a parallel success at unraveling the pattern of their instructive extracellular signals. Here we present proof of principle of a novel approach to circumvent this problem, based on the use of the HIV/TAT protein transduction domain. Neurogenin 3 (ngn3), a factor whose expression is essential for pancreatic endocrine differentiation, was fused to the TAT domain. Administration of TAT/ngn3 to cultured pancreatic explants results in efficient uptake, nuclear translocation, and stimulation of downstream reporter and endogenous genes. Consistent with the predicted activity of the protein, e9.5 and e13.5 mouse pancreatic explants cultured in the presence of TAT/ngn3 show an increased level of endocrine differentiation compared with control samples. Our results raise the possibility of sequentially specifying stem/progenitor cells toward the β-cell lineage, by using the appropriate sequence and combination of TAT-fused transcription factors.
Functional and Molecular Defects of Pancreatic Islets in Human Type 2 Diabetes
Authors: Del Guerra Silvia , Lupi Roberto , Marselli Lorella , Masini Matilde , Bugliani Marco , Sbrana Simone , Torri Scilla , Pollera Maria , Boggi Ugo , Mosca Franco , Del Prato Stefano , Marchetti Piero
Published: March 2007

To shed further light on the primary alterations of insulin secretion in type 2 diabetes and the possible mechanisms involved, we studied several functional and molecular properties of islets isolated from the pancreata of 13 type 2 diabetic and 13 matched nondiabetic cadaveric organ donors. Glucose-stimulated insulin secretion from type 2 diabetic islets was significantly lower than from control islets, whereas arginine- and glibenclamide-stimulated insulin release was less markedly affected. The defects were accompanied by reduced mRNA expression of GLUT1 and -2 and glucokinase and by diminished glucose oxidation. In addition, AMP-activated protein kinase activation was reduced. Furthermore, the expression of insulin was decreased, and that of pancreatic duodenal homeobox-1 (PDX-1) and forkhead box O1 (Foxo-1) was increased. Nitrotyrosine and 8-hydroxy-2′-deoxyguanosine concentrations, markers of oxidative stress, were significantly higher in type 2 diabetic than control islets, and they were correlated with the degree of glucose-stimulated insulin release impairment. Accordingly, 24-h exposure to glutathione significantly improved glucose-stimulated insulin release and decreased nitrotyrosine concentration, with partial recovery of insulin mRNA expression. These results provide direct evidence that the defects of insulin secretion in type 2 diabetic islets are associated with multiple islet cell alterations. Most importantly, the current study shows that the functional impairment of type 2 diabetic islets can be, at least in part, reversible. In this regard, it is suggested that reducing islet cell oxidative stress is a potential target of human type 2 diabetes therapy.
Contribution of Abdominal Visceral Obesity and Insulin Resistance to the Cardiovascular Risk Profile of Postmenopausal Women
Authors: Piché Marie-Ève , Weisnagel S. John , Corneau Louise , Nadeau André , Bergeron Jean , Lemieux Simone
Published: March 2007

The aim of this study was to determine the respective contribution of abdominal visceral adipose tissue (AT) accumulation and insulin resistance (IR) to the determination of a comprehensive cardiovascular metabolic risk profile in 108 postmenopausal women not receiving hormone therapy. Insulin sensitivity (M/I) was determined by a hyperinsulinemic-euglycemic clamp, and visceral AT area was measured by computed tomography. Median values of visceral AT (133.9 cm2) and insulin sensitivity (0.010189 mg · kg−1 · min−1 · pmol−1) were used to form four subgroups: 1) low visceral AT-low IR (n = 35), 2) low visceral AT-high IR (n = 19), 3) high visceral AT-low IR (n = 19), and 4) high visceral AT-high IR (n = 35). Women with isolated IR (low visceral AT and high IR) were characterized by significantly higher fasting and 2-h glycemia and higher fibrinogen, triglyceride, and VLDL-apolipoprotein (apo)B concentrations than women with low visceral AT and low IR (P < 0.05). The plasma lipid-lipoprotein profile and inflammatory markers were not significantly different between women with high visceral AT and low IR and women with low visceral AT and low IR. Women with high visceral AT and high IR had higher fasting and 2-h glycemia, triglyceride, and VLDL-apoB levels; lower apoAI and HDL2 cholesterol levels; as well as higher C-reactive protein and interleukin-6 concentrations than women with low visceral AT and low IR (P < 0.05). In addition, 15 of the 35 women (42.9%) in the high visceral AT and high IR group were newly diagnosed with type 2 diabetes, whereas no women were diagnosed with type 2 diabetes in the group of women with low visceral AT and low IR. These results show that although the presence of high IR in its isolated form is associated with some metabolic alterations, it is the combination of both high visceral AT and high IR that is the most detrimental for the metabolic health in postmenopausal women.
Nonviral Gene Transfer of Human Hepatocyte Growth Factor Improves Streptozotocin-Induced Diabetic Neuropathy in Rats
Authors: Kato Naoki , Nemoto Koichi , Nakanishi Kuniaki , Morishita Ryuichi , Kaneda Yasufumi , Uenoyama Maki , Ikeda Tomosumi , Fujikawa Kyosuke
Published: March 2007

Peripheral neuropathy is common and ultimately accounts for significant morbidity in diabetes. Recently, several neurotrophic factors have been used to prevent progression of diabetic neuropathy. In this study, we gave repeated intramuscular injections of the human hepatocyte growth factor (HGF) gene percutaneously, using liposomes containing the hemagglutinating virus of Japan (HVJ), to examine therapeutic efficacy of nonviral gene transfer of HGF for experimental diabetic sensorimotor neuropathy in rats. Experimental diabetes induced by intraperitoneal injection of streptozotocin resulted in a marked tactile allodynia (but not in a thermal hyperalgesia), in a reduction of both the conduction velocity and the amplitude, and in a decreased laser Doppler flux of the nerve and the muscle at 6 weeks after the induction. All these changes were significantly reversed by repeated gene transfer of HGF. Furthermore, we analyzed the density of endoneurial capillaries and morphometrical changes of the nerve. The density of endoneurial capillaries, disclosing marked reduction in diabetic rats, was also reversed significantly by repeated gene transfer of HGF; however, no considerable differences were observed morphometrically in either myelinated or unmyelinated axons. These results suggest that nonviral HVJ liposome-mediated gene transfer of human HGF has potential for the safe effective treatment of diabetic sensorimotor neuropathy.
Metabolic Dysregulation With Atypical Antipsychotics Occurs in the Absence of Underlying Disease A Placebo-Controlled Study of Olanzapine and Risperidone in Dogs
Authors: Ader Marilyn , Kim Stella P. , Catalano Karyn J. , Ionut Viorica , Hucking Katrin , Richey Joyce M. , Kabir Morvarid , Bergman Richard N.
Published: March 2007

Atypical antipsychotics have been linked to weight gain, hyperglycemia, and diabetes. We examined the effects of atypical antipsychotics olanzapine (OLZ) and risperidone (RIS) versus placebo on adiposity, insulin sensitivity (SI), and pancreatic β-cell compensation. Dogs were fed ad libitum and given OLZ (15 mg/day; n = 10), RIS (5 mg/day; n = 10), or gelatin capsules (n = 6) for 4–6 weeks. OLZ resulted in substantial increases in adiposity: increased total body fat (+91 ± 20%; P = 0.000001) reflecting marked increases in subcutaneous (+106 ± 24%; P = 0.0001) and visceral (+84 ± 22%; P = 0.000001) adipose stores. Changes in adiposity with RIS were not different from that observed in the placebo group (P > 0.33). Only OLZ resulted in marked hepatic insulin resistance (hepatic SI [pre- versus postdrug]: 6.05 ± 0.98 vs. 1.53 ± 0.93 dl · min−1 · kg−1/[μU/ml], respectively; P = 0.009). β-Cell sensitivity failed to upregulate during OLZ (pre-drug: 1.24 ± 0.15, post-drug: 1.07 ± 0.25 μU · ml−1/[mg/dl]; P = 0.6). OLZ-induced β-cell dysfunction was further demonstrated when β-cell compensation was compared with a group of animals with adiposity and insulin resistance induced by moderate fat feeding alone (+8% of calories from fat; n = 6). These results may explain the diabetogenic effects of atypical antipsychotics and suggest that β-cell compensation is under neural control.