bluuebunny/crossref_metadata_2025 huggingface dataset:
Insulin Resistance in the Liver-Specific IGF-1 Gene-Deleted Mouse Is Abrogated by Deletion of the Acid-Labile Subunit of the IGF-Binding Protein-3 Complex
Authors: Haluzik Martin , Yakar Shoshana , Gavrilova Oksana , Setser Jennifer , Boisclair Yves , LeRoith Derek
Published: March 2007

Liver IGF-1 deficient (LID) mice demonstrate a 75% reduction in circulating IGF-1 levels and a corresponding fourfold increase in growth hormone (GH) levels. At 16 weeks of age, LID mice demonstrate, using the hyperinsulinemic-euglycemic clamp, insulin insensitivity in muscle, liver, and fat tissues. In contrast, mice with a gene deletion of the acid-labile subunit (ALSKO) demonstrate a 65% reduction in circulating IGF-1 levels, with normal GH levels and no signs of insulin resistance. To further clarify the relative roles of increased GH and decreased IGF-1 levels in the development of insulin resistance, we crossed the two mouse lines and created a double knockout mouse (LID+ALSKO). LID+ALSKO mice demonstrate a further reduction in circulating IGF-1 levels (85%) and a concomitant 10-fold increase in GH levels. Insulin tolerance tests showed an improvement in insulin responsiveness in the LID+ALSKO mice compared with controls; LID mice were very insulin insensitive. Surprisingly, insulin sensitivity, while improved in white adipose tissue and in muscle, was unchanged in the liver. The lack of improvement in liver insulin sensitivity may reflect the absence of IGF-1 receptors or increased triglyceride levels in the liver. The present study suggests that whereas GH plays a major role in inducing insulin resistance, IGF-1 may have a direct modulatory role.
Nestin-Lineage Cells Contribute to the Microvasculature but Not Endocrine Cells of the Islet
Authors: Treutelaar Mary Kay , Skidmore Jennifer M. , Dias-Leme Claudia L. , Hara Manami , Zhang Lizhi , Simeone Diane , Martin Donna M. , Burant Charles F.
Published: March 2007

To clarify the lineage relationship between cells that express the neural stem cell marker nestin and endocrine cells of the pancreas, we analyzed offspring of a cross between mice carrying a nestin promoter/enhancer-driven cre-recombinase (Nestin-cre) and C57BL/6J-Gtrosa26tm1Sor mice that carry a loxP-disrupted β-galactosidase gene (Rosa26). In nestin-cre+/tg;R26RloxP/+ embryos, cre-recombinase was detected in association with nestin-positive cells in the pancreatic mesenchyme with some of the nestin-positive cells lining vascular channels. In postnatal mice, pancreatic β-galactosidase expression was restricted to vascular endothelial cells of the islet and a subset of cells in the muscularis of arteries in a distribution identical to endogenous nestin expression. Ex vivo explants of mouse pancreatic ducts grew dense cultures that costained for nestin and β-galactosidase, demonstrating recombination in vitro. The cultures could be differentiated into complex stereotypic structures that contain nestin- and insulin-expressing cells. Nestin-cre+/tg;R26RloxP/+-derived duct cultures showed that insulin-positive cells were negative for β-galactosidase. These results indicate that both in vivo and in vitro pancreatic endocrine cells arise independently of nestin-positive precursors. The apparent vascular nature of the nestin-positive cell population and the close association with endocrine cells suggest that nestin-positive cells play an important role in the growth and maintenance of the islet.
Characterization and Isolation of Promoter-Defined Nestin-Positive Cells from the Human Fetal Pancreas
Authors: Humphrey Rohan K. , Bucay Nathan , Beattie Gillian M. , Lopez Ana , Messam Conrad A. , Cirulli Vincenzo , Hayek Alberto
Published: March 2007

Studies using adult human islets and mouse embryonic stem cells have suggested that the neurepithelial precursor cell marker nestin also identifies and can be used to purify β-cell precursors. To determine whether nestin can be used to identify β-cell progenitors in the developing human pancreas, we characterized nestin expression from 12 to 24 gestational weeks, purified nestin+ cells using an enhancer/promoter-driven selection plasmid, and determined whether nestin+ cells can differentiate into β-cells. Nestin was visualized in the platelet endothelial cell adhesion molecule and α smooth muscle actin–positive blood vessels and colocalized with vimentin in the interstitium. Nestin was not observed in pan cytokeratin (pCK)-positive ductal epithelium or insulin cells. Purified nestin+ cells also coexpressed vimentin and lacked pCK immunoreactivity. Purified adult and fetal pancreatic fibroblasts also expressed nestin. The nestin enhancer/promoter used in the selection plasmid was sufficient to drive reporter gene expression, green fluorescent protein, in human fetal pancreatic tissue. Exposure of selected nestin+ cells to nicotinamide, hepatocyte growth factor/scatter factor, betacellulin, activin A, or exendin-4 failed to induce pancreatic and duodenal homeobox gene-1 or insulin message as determined by RT-PCR. Transplantation of nestin+ cells and fetal pancreatic fibroblasts into athymic mice also failed to result in the development of β-cells, whereas nestin− fetal pancreatic epithelial cells gave rise to functional insulin-secreting β-cells. We conclude that nestin is not a specific marker of β-cell precursors in the developing human pancreas.
Sulfonylureas Rapidly Cross Phospholipid Bilayer Membranes by a Free-Diffusion Mechanism
Authors: Kamp Frits , Kizilbash Nadeem , Corkey Barbara E. , Berggren Per-Olof , Hamilton James A.
Published: March 2007

Because sulfonylureas directly activate the exocytotic machinery, we were interested in the extent to which these compounds penetrate the β-cell plasma membrane and the underlying molecular mechanism(s). We now provide evidence that sulfonylureas cross phospholipid bilayer membranes rapidly and effectively by a free-diffusion mechanism. Two sulfonylurea compounds investigated by 1H nuclear magnetic resonance spectroscopy, glibenclamide and tolbutamide, were found to incorporate into phospholipid bilayers, with the ionizable sulfonamide exposed to the aqueous interface and its apparent dissociation constant (pKa) increased to ∼7.0. Diffusion of weak amphiphilic acids across membranes is associated with a measurable change in pH. Thus, by using a fluorescence-based pH assay, we could investigate the diffusion of sulfonylurea compounds across phospholipid bilayer membranes. A fluorescent pH indicator (pyranin or [2′,7′-bis (2-carboxyethyl)-5(6)-carboxyfluorescein] [BCECF]) was trapped in egg phosphatidylcholine vesicles. Addition of glibenclamide decreased internal pH (pHin), and addition of albumin reversed this drop by 50%. With the same amount of tolbutamide, the decrease in pHin was much smaller, primarily because of the lower partitioning of tolbutamide into phospholipid bilayers. Using similar protocols, we also demonstrated diffusion by the same mechanism across the β-cell plasma membrane. Thus, we now provide a molecular mechanism by which sulfonylureas can penetrate the plasma membrane and reach intracellular sites regulating exocytosis.
Neuroprotective Effect of Docosahexaenoic Acid-Enriched Phospholipids in Experimental Diabetic Neuropathy
Authors: Coste Thierry C. , Gerbi Alain , Vague Philippe , Pieroni Gérard , Raccah Denis
Published: March 2007

A deficiency in essential fatty acid metabolism has been widely reported in both human and animal diabetes. Fish oil supplementations (n-3 fatty acids), containing docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA), were less effective on diabetic neuropathy than (n-6) fatty acids. This partial effect of (n-3) fatty acids might be attributed to the presence of EPA, a competitor of arachidonic acid, which enhanced the diabetes-induced decrease of this fatty acid in serum and tissues. For determining whether a supplementation with DHA alone could prevent neuropathy in streptozotocin-induced diabetes, diabetic rats were given daily, by gavage, liposomes containing DHA phospholipids, at a dose of 60 mg/kg. Eight weeks of diabetes induced significant decreases in nerve conduction velocity (NCV), nerve blood flow (NBF), and sciatic nerve and erythrocyte (red blood cells [RBCs]) Na,K-ATPase activities. DHA phospholipids totally prevented the decrease in NCV and NBF observed during diabetes when compared with the nonsupplemented diabetic group. DHA phospholipids also prevented the Na,K-ATPase activity decrease in RBC but not in sciatic nerve. Moreover, DHA level in sciatic nerve membranes was correlated with NCV. These results demonstrate a protective effect of daily doses of DHA on experimental diabetic neuropathy. Thus, treatment with DHA phospholipids could be suitable for evaluation in clinical trials.
Glucose Induces Clonal Selection and Reversible Dinucleotide Repeat Expansion in Mesangial Cells Isolated from Glomerulosclerosis-Prone Mice
Authors: Fornoni Alessia , Lenz Oliver , Striker Liliane J. , Striker Gary E.
Published: March 2007

Clonal selection has been proposed as a pathogenetic mechanism in various chronic diseases, such as scleroderma, hypertension, pulmonary fibrosis, interstitial fibrosis of the kidney, atherosclerosis, and uterine leiomyomatosis. We previously found that mesangial cells from ROP mice prone to develop glomerulosclerosis changed their phenotype in response to high glucose concentrations. Here, we investigate whether clonal selection might contribute to this phenotype change. We found that in ROP mice at least two distinct mesangial cell clones exist. They are characterized by a different length of the d(CA) repeat in the MMP-9 promoter and exhibit a significantly different gene expression profile. Exposure of ROP mesangial cells to 25 mmol/l glucose for 35 days induces both clonal selection and reversible dinucleotide repeat expansion. None of these findings were present in mesangial cells isolated from C57BL/6 mice, which are not sclerosis-prone. We conclude that mesangial cell michrochimerism may be a marker for the susceptibility to glomerulosclerosis, that dinucleotide repeat expansion may be a novel mechanism for glucose-induced changes in gene expression, and that clonal selection may partially explain the change in mesangial cell phenotype in diabetes.
Relationship of Family History of Type 2 Diabetes, Hypoglycemia, and Autoantibodies to Weight Gain and Lipids With Intensive and Conventional Therapy in the Diabetes Control and Complications Trial
Authors: Purnell Jonathan Q. , Dev Raj K. , Steffes Michael W. , Cleary Patricia A. , Palmer Jerry P. , Hirsch Irl B. , Hokanson John E. , Brunzell John D.
Published: March 2007

Intensive therapy for type 1 diabetes results in greater weight gain than conventional therapy. Many factors may predispose to this greater weight gain, including improved glycemic control, genetic susceptibility to obesity, and hypoglycemia. To study this, relationships among family history of type 2 diabetes, frequency of severe hypoglycemia, β-cell autoantibodies, and weight gain were examined in 1,168 subjects aged ≥18 years at baseline randomized to intensive and conventional therapy groups in the Diabetes Control and Complications Trial. With intensive therapy, subjects with a family history of type 2 diabetes had greater central weight gain and dyslipidemia characterized by higher triglyceride levels and greater cholesterol in VLDLs and intermediate-density lipoproteins compared with subjects with no family history. Neither the frequency of severe hypoglycemia nor positivity to GAD65 and insulinoma-associated protein 2 antibodies was associated with increased weight gain with either intensive or conventional therapy. These data support the hypothesis that increased weight gain with intensive therapy might be explained, in part, by genetic traits.
Increased Insulin Sensitivity and Hypoinsulinemia in APS Knockout Mice
Authors: Minami Asako , Iseki Masanori , Kishi Kazuhiro , Wang Miao , Ogura Makoto , Furukawa Noboru , Hayashi Sanae , Yamada Mizuki , Obata Toshiyuki , Takeshita Yukari , Nakaya Yutaka , Bando Yoshimi , Izumi Keisuke , Moodie Shonna A. , Kajiura Fumiko , Matsumoto Mitsuru , Takatsu Kiyoshi , Takaki Satoshi , Ebina Yousuke
Published: March 2007

A tyrosine kinase adaptor protein containing pleckstrin homology and SH2 domains (APS) is rapidly and strongly tyrosine phosphorylated by insulin receptor kinase upon insulin stimulation. The function of APS in insulin signaling has heretofore remained unknown. APS-deficient (APS−/−) mice were used to investigate its function in vivo. The blood glucose-lowering effect of insulin, as assessed by the intraperitoneal insulin tolerance test, was increased in APS−/− mice. Plasma insulin levels during fasting and in the intraperitoneal glucose tolerance test were lower in APS−/− mice. APS−/− mice showed an increase in the whole-body glucose infusion rate as assessed by the hyperinsulinemic-euglycemic clamp test. These findings indicated that APS−/− mice exhibited increased sensitivity to insulin. However, overexpression of wild-type or dominant-negative APS in 3T3L1 adipocytes did not affect insulin receptor numbers, phosphorylations of insulin receptor, insulin receptor substrate-1, or Akt and mitogen-activated protein kinase. The glucose uptake and GLUT4 translocation were not affected by insulin stimulation in these cells. Nevertheless, the insulin-stimulated glucose transport in isolated adipocytes of APS−/− mice was increased over that of APS+/+ mice. APS−/− mice also showed increased serum levels of leptin and adiponectin, which might explain the increased insulin sensitivity of adipocytes.
Target Cell Expression of Suppressor of Cytokine Signaling-1 Prevents Diabetes in the NOD Mouse
Authors: Flodström-Tullberg Malin , Yadav Deepak , Hägerkvist Robert , Tsai Devin , Secrest Patrick , Stotland Alexandr , Sarvetnick Nora
Published: March 2007

Although lymphocyte infiltration and islet destruction are hallmarks of diabetes, the mechanisms of β-cell destruction are not fully understood. One issue that remains unresolved is whether cytokines play a direct role in β-cell death. We investigated whether β-cell cytokine signaling contributes to autoimmune type 1 diabetes. We demonstrated that NOD mice harboring β-cells expressing the suppressor of cytokine signaling-1 (SOCS-1), an inhibitor of Janus kinase/signal transducers and activators of transcription (JAK/STAT) signaling, have a markedly reduced incidence of diabetes. Similar to their non-transgenic (Tg) littermates, SOCS-1-Tg mice develop insulitis and their splenocytes transfer disease to NODscid recipients. Disease protection correlates with suppression of cytokine-induced STAT-1 phosphorylation in SOCS-1–expressing β-cells and with a reduced sensitivity of these cells to destruction by diabetogenic cells in vivo. Interestingly, lymphocytes recruited to the pancreas of SOCS-1-Tg mice transferred diabetes to NODscid recipients with a reduced efficiency, suggesting that the pancreatic environment in SOCS-1-Tg mice does not support the maintenance of functionally differentiated T-cells. These results suggest that cytokines contribute to the development of type 1 diabetes by acting directly on the target β-cell. Importantly, given that the SOCS-1–expressing mouse maintain normal blood glucose levels throughout life, this study also showed that SOCS-1 expression by β-cells can represent a promising strategy to prevent type 1 diabetes.
Identification of Subjects with Insulin Resistance and β-Cell Dysfunction Using Alternative Definitions of the Metabolic Syndrome
Authors: Hanley Anthony J.G. , Wagenknecht Lynne E. , D’Agostino Ralph B. , Zinman Bernard , Haffner Steven M.
Published: March 2007

Recently, the metabolic syndrome (MetS) has attracted much attention as a risk cluster for cardiovascular disease. Although it is believed that individuals with the MetS have insulin resistance (IR), there are few data using direct measures of IR such as glucose clamps or frequently sampled intravenous glucose tolerance tests (FSIGTTs). We examined associations of MetS with FSIGTT-derived measures of insulin sensitivity and secretion among nondiabetic subjects in the Insulin Resistance Atherosclerosis Study. Two sets of MetS criteria were evaluated: those from the 1999 World Health Organization (WHO) and the 2001 National Cholesterol Education Program (NCEP). Both WHO and NCEP MetS definitions were significantly associated with risk of being in the lowest quartile of directly measured insulin sensitivity (P < 0.0001 for all subjects as well as within ethnic subgroups). However, the associations with WHO-MetS were stronger for all subjects combined (WHO: odds ratio [OR] = 10.2; 95% CI 7.5–13.9; NCEP: OR = 4.6; 3.4–6.2) and in separate analyses of non-Hispanic whites, blacks, and Hispanics. WHO and NCEP MetS definitions were also significantly associated with risk of being in the lowest quartile of insulin sensitivity–adjusted acute insulin response (AIR) and disposition index (DI; all P < 0.01), although the associations were generally weaker than those for insulin sensitivity and there was no difference between the two definitions in all subjects combined (low AIR, WHO: OR = 1.7, 1.2–2.4; NCEP: OR = 1.7, 1.2–2.5). There were, however, a number of ethnic differences, including a stronger association of NCEP-MetS with low AIR among blacks. WHO-MetS was significantly more sensitive than NCEP-MetS in detecting low insulin sensitivity (65.4 vs. 45.6%, respectively; P < 0.0001), with no significant differences in specificity between the definitions (84.4 vs. 84.6%; P = 0.91), although WHO-MetS had a larger area under the receiver operating characteristic curve (75% vs. 65%; P < 0.0001). In conclusion, although both the WHO and NCEP MetS criteria identify nondiabetic individuals with low insulin sensitivity, the associations were notably stronger using the WHO definition. The definitions are generally less useful for identifying those with low AIR or DI, although NCEP-MetS seems to differentiate black subjects with insulin secretion defects.
Amelioration of Sensory Nerve Dysfunction by C-Peptide in Patients With Type 1 Diabetes
Authors: Ekberg Karin , Brismar Tom , Johansson Bo-Lennart , Jonsson Björn , Lindström Per , Wahren John
Published: March 2007

Studies have demonstrated that proinsulin C-peptide stimulates the activities of Na+,K+-ATPase and endothelial nitric oxide synthase, both of which are enzyme systems of importance for nerve function and known to be deficient in type 1 diabetes. The aim of this randomized double-blind placebo-controlled study was to investigate whether C-peptide replacement improves nerve function in patients with type 1 diabetes. Forty-nine patients without symptoms of peripheral neuropathy were randomized to either 3 months of treatment with C-peptide (600 nmol/24 h, four doses s.c.) or placebo. Forty-six patients (15 women and 31 men, aged 29 years, diabetes duration 10 years, and HbA1c 7.0%) completed the study. Neurological and neurophysiological measurements were performed before and after 6 and 12 weeks of treatment. At baseline the patients showed reduced nerve conduction velocities in the sural nerve (sensory nerve conduction velocity [SCV]: 50.9 ± 0.70 vs. 54.2 ± 1.2 m/s, P < 0.05) and peroneal nerve (motor nerve conduction velocity: 45.7 ± 0.55 vs. 53.5 ± 1.1 m/s, P < 0.001) compared with age-, height-, and sex-matched control subjects. In the C-peptide treated group there was a significant improvement in SCV amounting to 2.7 ± 0.85 m/s (P < 0.05 compared with placebo) after 3 months of treatment, representing 80% correction of the initial reduction in SCV. The change in SCV was accompanied by an improvement in vibration perception in the patients receiving C-peptide (P < 0.05 compared with placebo), whereas no significant change was detectable in cold or heat perception. In conclusion, C-peptide administered for 3 months as replacement therapy to patients with early signs of diabetic neuropathy ameliorates nerve dysfunction.
Role of Corticotrophin-Releasing Hormone in the Impairment of Counterregulatory Responses to Hypoglycemia
Authors: Flanagan Daniel E. , Keshavarz Tara , Evans Mark L. , Flanagan Sally , Fan Xiaoning , Jacob Ralph J. , Sherwin Robert S.
Published: March 2007

We have explored the role of individual elements of the hypothalamic pituitary adrenal axis on the pathogenesis of hypoglycemia-associated autonomic failure. Five groups of male Sprague-Dawley rats were used. Control animals had 3 days of sham treatment followed by a hyperinsulinemic/hypoglycemic glucose clamp on day 4. A second group underwent 3 days of antecedent insulin-induced hypoglycemia then a subsequent clamp. Three more groups underwent pretreatment with corticosterone, adrenocorticotrophic hormone (ACTH), or corticotrophin-releasing hormone (CRH) mirroring the glucocorticoid response of the hypoglycemic group. Subsequent counterregulatory responses showed marked differences. CRH- (and insulin-treated) animals showed markedly reduced epinephrine responses (CRH 1,276 ± 404 pg/ml, controls 3,559 ± 563 pg/ml; P < 0.05). In contrast, ACTH pretreatment augmented epinephrine responses (6,681 ± 814 pg/ml; P = 0.007 versus controls); corticosterone pretreatment caused a similar but nonsignificant enhancement. The same pattern was seen for norepinephrine. CRH pretreatment also suppressed glucagon responses to hypoglycemia (control 157 ± 21, CRH 68 ± 10 pg/ml; P = 0.004). The addition of a CRH receptor 1 (CRHr1) antagonist to the antecedent CRH reversed the subsequent suppression of epinephrine. These findings suggest that CRH acting via CRHr1 plays an important role in the sympathoadrenal downregulation seen in this rodent model of antecedent hypoglycemia; this action is not mediated via activation of the hypothalamic-pituitary-adrenal axis.
Metallothionein Prevents Diabetes-Induced Deficits in Cardiomyocytes by Inhibiting Reactive Oxygen Species Production
Authors: Ye Gang , Metreveli Naira S. , Ren Jun , Epstein Paul N.
Published: March 2007

Many individuals with diabetes experience impaired cardiac contractility that cannot be explained by hypertension and atherosclerosis. This cardiomyopathy may be due to either organ-based damage, such as fibrosis, or to direct damage to cardiomyocytes. Reactive oxygen species (ROS) have been proposed to contribute to such damage. To address these hypotheses, we examined contractility, Ca2+ handling, and ROS levels in individual cardiomyocytes isolated from control hearts, diabetic OVE26 hearts, and diabetic hearts overexpressing antioxidant protein metallothionein (MT). Our data showed that diabetic myocytes exhibited significantly reduced peak shortening, prolonged duration of shortening/relengthening, and decreased maximal velocities of shortening/relengthening as well as slowed intracellular Ca2+ decay compared with control myocytes. Overexpressing MT prevented these defects induced by diabetes. In addition, high glucose and angiotensin II promoted significantly increased generation of ROS in diabetic cardiomyocytes. Chronic overexpression of MT or acute in vitro treatment with the flavoprotein inhibitor diphenyleneiodonium or the angiotensin II type I receptor antagonist losartan eliminated excess ROS production in diabetic cardiomyocytes. These data show that diabetes induces damage at the level of individual myocyte. Damage can be attributed to ROS production, and diabetes increases ROS production via angiotensin II and flavoprotein enzyme-dependent pathways.
The Response of Antioxidant Genes to Hyperglycemia Is Abnormal in Patients With Type 1 Diabetes and Diabetic Nephropathy
Authors: Hodgkinson Andrea D. , Bartlett Tracey , Oates Peter J. , Millward Beverley A. , Demaine Andrew G.
Published: March 2007

Increased flux of glucose through the polyol pathway may cause generation of excess reactive oxygen species (ROS), leading to tissue damage. Abnormalities in expression of enzymes that protect against oxidant damage may accentuate the oxidative injury. The expression of catalase (CAT), CuZn superoxide-dismutase (CuZnSOD), glutathione peroxidase (GPX), and Mn superoxide-dismutase (MnSOD) mRNA was quantified in peripheral blood mononuclear cells—obtained from 26 patients with type 1 diabetes and nephropathy, 15 with no microvascular complications after 20 years’ duration of diabetes, and 10 normal healthy control subjects—that were exposed in vitro to hyperglycemia (HG) (31 mmol/l d-glucose). Under HG, there was a twofold increase in the expression of CAT, CuZnSOD, and GPX mRNA in the patients without complications and the control subjects versus patients with nephropathy (P < 0.0001), and MnSOD did not change in any of the groups. The aldose reductase inhibitor zopolrestat partially restored the levels of CAT, CuZnSOD, and GPX mRNA in the patients with nephropathy (P < 0.05). There was a highly significant correlation between increased aldose reductase (ALR2) expression, CAT, CuZnSOD, and GPX mRNA levels under HG conditions and polymorphisms of ALR2 in the patients with nephropathy (P < 0.00001). In conclusion, these results suggest that high glucose flux through aldose reductase inhibits the expression of antioxidant enzymes.
Low Glomerular Filtration Rate in Normoalbuminuric Type 1 Diabetic Patients
Authors: Caramori M. Luiza , Fioretto Paola , Mauer Michael
Published: March 2007

Increased urinary albumin excretion rate is widely accepted as the first clinical sign of diabetic nephropathy. However, it is possible that some diabetic patients could first manifest reduced glomerular filtration rate (GFR) or hypertension. Relatively advanced diabetic renal lesions can be present in some diabetic patients with long-standing normoalbuminuria, and this might indicate increased risk of progression to microalbuminuria and then to overt diabetic nephropathy. The aim of this study was to identify a group of normoalbuminuric type 1 diabetic patients with low GFR and compare them with normoalbuminuric patients with normal GFR. Altogether, 105 normoalbuminuric type 1 diabetic patients with at least 10 years of diabetes duration that had a renal biopsy performed for research purposes were studied. Patients were divided according to GFR into groups with normal (≥90 ml · min−1 · 1.73 m−2) or reduced (<90 ml · min−1 · 1.73 m−2) GFR. Clinical and renal structural parameters were compared between these two groups. Glomerular structural parameters were estimated by electron microscopic morphometry. The 23 patients with reduced GFR had more advanced diabetic glomerular lesions. The finding of reduced GFR was much more common among female patients, particularly if retinopathy and/or hypertension were also present. This report confirms that reduced GFR occurs among long-standing normoalbuminuric type 1 diabetic patients and is associated with more advanced diabetic glomerular lesions and, probably, with increased risk of progression. For these reasons, we suggest that regular measurements of GFR be performed in long-standing normoalbuminuric type 1 diabetic female diabetic patients, especially in those with retinopathy or hypertension.
Using Genetic Admixture to Explain Racial Differences in Insulin-Related Phenotypes
Authors: Gower Barbara A. , Fernández José R. , Beasley T. Mark , Shriver Mark D. , Goran Michael I.
Published: March 2007

Documented differences in measures of insulin secretion and action between African Americans and European Americans may be due to either genetic or environmental factors. This study used genetic admixture (ADM), determined from ∼20 ancestry informative markers, and a questionnaire designed to assess socioeconomic status (SES) to examine potential genetic and environmental contributions to minimal model–derived measures of insulin sensitivity (SI), fasting insulin, and the acute insulin response to glucose (AIRg) in 125 children residing in Birmingham, Alabama. The study was longitudinal in design and yielded multiple outcome measures on each subject. Mixed models analysis was used to determine if ADM and SES were independently related to SI, fasting insulin, and AIRg after adjusting for confounding factors (pubertal status, adiposity, age) and for repeated testing of individuals. In this cohort, African ADM ranged from 0% (individuals with no markers reflecting African ancestry) to 100% (individuals with all 20 markers reflecting African ancestry). Results indicated that ADM was independently related to SI (P < 0.001) and fasting insulin (P < 0.01), with individuals having greater African ADM having a lower SI and a higher fasting insulin concentration. Both ADM (P < 0.001) and SES (P < 0.05) were independently related to AIRg; children with greater African ADM or lower SES had a higher AIRg, even after adjusting for SI. These observations suggest that use of ADM can replace assignment of individuals to categorical racial groups; that lower SI and higher fasting insulin among African Americans compared with European Americans may have a genetic basis; and that higher AIRg among African Americans may be due to both genetic factors and to environmental factors that remain to be identified.
Prolonged Islet Allograft Survival in Diabetic NOD Mice by Targeting CD45RB and CD154
Authors: Molano R. Damaris , Pileggi Antonello , Berney Thierry , Poggioli Raffaella , Zahr Elsie , Oliver Robert , Ricordi Camillo , Rothstein David M. , Basadonna Giacomo P. , Inverardi Luca
Published: March 2007

Clinical islet transplantation is a successful procedure that can improve the quality of life in recipients with diabetes. A drawback of the procedure is the need for chronic administration of immunosuppressive drugs that, among other side effects, are potentially diabetogenic. Definition of immunosuppressive protocols that utilize nondiabetogenic compounds could further improve islet transplantation outcome. We used the NOD mouse to assess the effect of targeting the T-lymphocyte surface receptors CD45RB and CD154 in preventing loss of allogeneic islet grafts as a result of recurrence of autoimmunity and allorejection. Administration of the two antibodies led to significantly prolonged allograft survival, with a percentage of grafts surviving long-term. The therapeutic efficacy of the treatment was paralleled by a shift in CD45RB isoform expression on T-lymphocytes, increased in vitro responsiveness to interleukin-7, and increased in vitro γ-interferon production after anti-CD3 antibody stimulation. Furthermore, graft infiltration by CD8+ T-cells was remarkably reduced. Recipient mice bearing functioning allografts were otherwise immunocompetent, as assessed in vivo and in vitro by numerous tests, including intragraft cytokine production, responsiveness to polyclonal stimulation and alloantigens, and analysis of cell subset phenotype. These data show that nondiabetogenic regimens of immunomodulation can lead to prolonged islet allograft survival in the challenging NOD mouse model.
5-Amino-Imidazole Carboxamide Riboside Increases Glucose Transport and Cell-Surface GLUT4 Content in Skeletal Muscle From Subjects With Type 2 Diabetes
Authors: Koistinen Heikki A. , Galuska Dana , Chibalin Alexander V. , Yang Jing , Zierath Juleen R. , Holman Geoffrey D. , Wallberg-Henriksson Harriet
Published: March 2007

AMP-activated protein kinase (AMPK) activation by AICAR (5-amino-imidazole carboxamide riboside) is correlated with increased glucose transport in rodent skeletal muscle via an insulin-independent pathway. We determined in vitro effects of insulin and/or AICAR exposure on glucose transport and cell-surface GLUT4 content in skeletal muscle from nondiabetic men and men with type 2 diabetes. AICAR increased glucose transport in a dose-dependent manner in healthy subjects. Insulin and AICAR increased glucose transport and cell-surface GLUT4 content to a similar extent in control subjects. In contrast, insulin- and AICAR-stimulated responses on glucose transport and cell-surface GLUT4 content were impaired in subjects with type 2 diabetes. Importantly, exposure of type 2 diabetic skeletal muscle to a combination of insulin and AICAR increased glucose transport and cell-surface GLUT4 content to levels achieved in control subjects. AICAR increased AMPK and acetyl-CoA carboxylase phosphorylation to a similar extent in skeletal muscle from subjects with type 2 diabetes and nondiabetic subjects. Our studies highlight the potential importance of AMPK-dependent pathways in the regulation of GLUT4 and glucose transport activity in insulin-resistant skeletal muscle. Activation of AMPK is an attractive strategy to enhance glucose transport through increased cell surface GLUT4 content in insulin-resistant skeletal muscle.
Regulation of Microsomal Triglyceride Transfer Protein Gene by Insulin in HepG2 Cells
Authors: Au Wo-Shing , Kung Hsiang-fu , Lin Marie C.
Published: March 2007

Microsomal triglyceride transfer protein (MTP) is rate limiting for the assembly and secretion of apolipoprotein B-containing lipoproteins. Elevated hepatic MTP mRNA level, presumably as a result of impaired insulin signaling, has been implicated in the pathophysiology of dyslipidemia associated with insulin resistance/type 2 diabetes. In this study, we showed that insulin decreases MTP mRNA level mainly through transcriptional regulation in HepG2 cells. We further characterized the corresponding signal transduction pathway, using chemical inhibitors and constitutively active and dominant negative forms of regulatory enzymes. We demonstrated that insulin inhibits MTP gene transcription through MAPKerk cascade but not through the PI 3-kinase pathway. Activation of ras through farnesylation is not a prerequisite for the inhibition. In addition, cellular MAPKerk and MAPKp38 activities play a counterbalancing role in regulating the MTP gene transcription. These complex regulations may represent a means to fine-tuning MTP gene transcription in response to a diverse set of environmental stimuli and may have important implications for the onset and development of diabetes-associated dyslipidemia.
Nuclear Factor κB Protects Pancreatic β-Cells From Tumor Necrosis Factor-α-Mediated Apoptosis
Authors: Chang Inik , Kim Sunshin , Kim Ja Young , Cho Namjoo , Kim Yun-Hee , Kim Hun Sik , Lee Moon-Kyu , Kim Kwang-Won , Lee Myung-Shik
Published: March 2007

Recent studies incriminating tumor necrosis factor (TNF)-α as the final effector in pancreatic β-cell death in type 1 diabetes underscore the potential role of TNF-α-dependent NF-κB activation as an important modulator of pancreatic β-cell death in autoimmune diabetes. Although nuclear factor (NF)-κB activation has been implicated in the protection of target cells against apoptosis by a variety of death effectors, its role in pancreatic islet cell death is not clear. We studied the role of NF-κB activation in pancreatic islet cell death by using a γ-interferon (IFN-γ)/TNF-α synergism model we had previously reported. TNF-α induced inhibitor of κB (IκB) degradation and p65 translocation from cytoplasm to nuclei in MIN6N8 insulinoma cells. The NF-κB DNA-binding nuclear complex activated by TNF-α contained both the p65 and p50 subunit. IFN-γ pretreatment did not affect TNF-α-induced NF-κB activation. Treatment with a proteasome inhibitor blocked p65 translocation and induced susceptibility to TNF-α in otherwise resistant insulinoma cells or primary pancreatic islet cells. Specific inhibition of NF-κB activation by adenoviral transduction of IκB “superrepressor” also sensitized insulinoma cells and primary islet β-cells to TNF-α-induced apoptosis. These results suggest the protective role of NF-κB activation against cytokine-mediated pancreatic β-cell death, contrary to previous reports implicating NF-κB as a mediator of pancreatic islet cell death.
Increased Expression of Ornithine Decarboxylase in Distal Tubules of Early Diabetic Rat Kidneys
Authors: Deng Aihua , Munger Karen A. , Valdivielso Jose M. , Satriano Joseph , Lortie Mark , Blantz Roland C. , Thomson Scott C.
Published: March 2007

Polyamines are small biogenic molecules that are essential for cell cycle entry and progression and proliferation. They can also contribute to hypertrophy. The activity of ornithine decarboxylase (ODC), the rate-limiting enzyme in polyamine biosynthesis, increases in the early diabetic kidney to enable renal hypertrophy. Inhibition of ODC in early diabetes attenuates diabetic renal hypertrophy and glomerular hyperfiltration. The current studies examine the temporal profile of renal ODC protein expression and localization, intrarenal polyamine levels, and sites of proliferation in kidneys of rats during the first 7 days of streptozotocin diabetes. ODC mRNA and protein content were increased in diabetic kidneys. High-performance liquid chromatography analysis showed increased intrarenal polyamine concentrations peaking after 24 h of diabetes. A subsequent increase in the number of proliferating proximal tubular cells was detected by in vivo 5-bromodeoxyuridine (BrdU) incorporation on day 3. Surprisingly, immunohistochemical studies revealed that increased ODC protein was apparent only in distal nephrons, whereas the main site of diabetic kidney hypertrophy is the proximal tubule. These findings raise the possibility that polyamines produced in the distal nephron may mediate the early diabetic kidney growth of the proximal tubules via a paracrine mechanism.
High Glucose-Induced Expression of Proinflammatory Cytokine and Chemokine Genes in Monocytic Cells
Authors: Shanmugam Narkunaraja , Reddy Marpadga A. , Guha Mausumee , Natarajan Rama
Published: March 2007

Monocyte activation and adhesion to the endothelium play important roles in inflammatory and cardiovascular diseases. These processes are further aggravated by hyperglycemia, leading to cardiovascular complications in diabetes. We have previously shown that high glucose (HG) treatment activates monocytes and induces the expression of tumor necrosis factor (TNF)-α via oxidant stress and nuclear factor-kB transcription factor. To determine the effects of HG on the expression of other inflammatory genes, in the present study, HG-induced gene profiling was performed in THP-1 monocytes using cytokine gene arrays containing 375 known genes. HG treatment upregulated the expression of 41 genes and downregulated 15 genes that included chemokines, cytokines, chemokines receptors, adhesion molecules, and integrins. RT-PCR analysis further confirmed that HG significantly increased the expression of monocyte chemoattractant protein-1 (MCP-1), TNF-α, β2-integrin, interleukin-1β, and others. HG treatment increased transcription of the MCP-1 gene, MCP-1 protein levels, and adhesion of THP-1 cells to endothelial cells. HG-induced MCP-1 mRNA expression and monocyte adhesion were blocked by specific inhibitors of oxidant stress, protein kinase C, ERK1/2, and p38 mitogen-activated protein kinases. These results show for the first time that multiple inflammatory cytokines and chemokines relevant to the pathogenesis of diabetes complications are induced by HG via key signaling pathways.
Genetic Variation in the Human Winged Helix/Forkhead Transcription Factor Gene FOXC2 in Pima Indians
Authors: Kovacs Peter , Lehn-Stefan Angela , Stumvoll Michael , Bogardus Clifton , Baier Leslie J.
Published: March 2007

FOXC2 is a winged helix gene that has been shown to counteract obesity, hypertriglyceridemia, and diet-induced insulin resistance in rodents. Therefore, FOXC2 was analyzed as a candidate gene for susceptibility to type 2 diabetes in Pima Indians. Four variants were identified by sequencing the coding region, as well as 638 bp of the 5′ region and 300 bp of the 3′ region of the gene. Two single nucleotide polymorphisms (SNPs) were found in the putative promoter region, a C-512T transition and a G-350T. In addition, two SNPs were found in the 3′ region, a C1548T and a C1702T. The G-350T and the C1702T variants were in complete linkage disequilibrium, and the C1548T variant was relatively rare; therefore, only the C-512T and G-350T variants were additionally genotyped in 937 full-blooded Pima Indians. Neither of these polymorphisms were associated with type 2 diabetes; however, the C-512T variant was associated with BMI (P = 0.03) and percentage of body fat (P = 0.02) in male and female Pima subjects, as well as with basal glucose turnover and fasting plasma triglycerides in women. Our data indicate that variation in FOXC2 may have a minor role in body weight control and seems to be involved in the regulation of basal glucose turnover and plasma triglyceride levels in women, but this gene does not significantly contribute to the etiology of type 2 diabetes in Pima Indians.
Fetal Programming of Perivenous Glucose Uptake Reveals a Regulatory Mechanism Governing Hepatic Glucose Output During Refeeding
Authors: Murphy Helena C. , Regan Gemma , Bogdarina Irina G. , Clark Adrian J.L. , Iles Richard A. , Cohen Robert D. , Hitman Graham A. , Berry Colin L. , Coade Zoe , Petry Clive J. , Burns Shamus P.
Published: March 2007

Increased hepatic gluconeogenesis maintains glycemia during fasting and has been considered responsible for elevated hepatic glucose output in type 2 diabetes. Glucose derived periportally via gluconeogenesis is partially taken up perivenously in perfused liver but not in adult rats whose mothers were protein-restricted during gestation (MLP rats)—an environmental model of fetal programming of adult glucose intolerance exhibiting diminished perivenous glucokinase (GK) activity. We now show that perivenous glucose uptake rises with increasing glucose concentration (0–8 mmol/l) in control but not MLP liver, indicating that GK is flux-generating. The data demonstrate that acute control of hepatic glucose output is principally achieved by increasing perivenous glucose uptake, with rising glucose concentration during refeeding, rather than by downregulation of gluconeogenesis, which occurs in different hepatocytes. Consistent with these observations, glycogen synthesis in vivo commenced in the perivenous cells during refeeding, MLP livers accumulating less glycogen than controls. GK gene transcription was unchanged in MLP liver, the data supporting a recently proposed posttranscriptional model of GK regulation involving nuclear-cytoplasmic transport. The results are pertinent to impaired regulation of hepatic glucose output in type 2 diabetes, which could arise from diminished GK-mediated glucose uptake rather than increased gluconeogenesis.
Insulin Resistance and Lipodystrophy in Mice Lacking Ribosomal S6 Kinase 2
Authors: El-Haschimi Karim , Dufresne Scott D. , Hirshman Michael F. , Flier Jeffrey S. , Goodyear Laurie J. , Bjørbæk Christian
Published: March 2007

The p90 ribosomal S6 kinase 2 (RSK2) is a serine/threonine kinase with high expression levels in adipose tissue. Numerous in vitro studies show that RSK2 is activated by a broad number of cellular stimuli and suggest that RSK2 is involved in the regulation of a variety of cellular processes. However, the physiological role of RSK2 still remains elusive. We therefore generated rsk2 knockout (KO) mice to better understand the function of RSK2 in vivo. Birth weights of RSK2 KO mice are normal, but the body weight is reduced with age, as compared with wild-type littermates. We found that the difference in body weight was largely caused by a specific loss of white adipose tissue that is accompanied by reduced serum levels of the adipocyte-derived peptide, leptin. KO mice also have impaired glucose tolerance and elevated fasting insulin and glucose levels that are restored following administration of low amounts of leptin, which do not affect food intake. We conclude that RSK2 plays a novel and an important role in regulation of adipose mass in mice and speculate that the reduction in fat tissue may negatively affect insulin sensitivity, as observed in human lipodystrophy, through reduced levels of adipocyte-derived factors, such as leptin.
Renal Compensation for Impaired Hepatic Glucose Release During Hypoglycemia in Type 2 Diabetes
Authors: Woerle Hans J. , Meyer Christian , Popa Emilia M. , Cryer Philip E. , Gerich John E.
Published: March 2007

During liver transplantation and after both meal ingestion and prolonged fasting, renal glucose release (RGR) increases while hepatic glucose release (HGR) decreases. These and other observations have led to the concept of hepatorenal reciprocity. According to this concept, reciprocal changes in hepatic and renal glucose release may occur to minimize deviations from normal glucose homeostasis. We further assessed this concept by testing the hypothesis that during counterregulation of hypoglycemia in patients with type 2 diabetes, who would be expected to have reduced HGR, RGR would be increased. Accordingly, we performed hypoglycemic hyperinsulinemic clamp experiments (∼3.1 mmol/l) in 12 type 2 diabetic and in 10 age-weight-matched nondiabetic volunteers and measured total endogenous glucose release (TEGR) and RGR using a combined isotopic net balance approach. HGR was calculated as the difference between TEGR and RGR since only these organs are capable of releasing glucose. We found that during comparable hypoglycemia and hyperinsulinemia, TEGR was reduced in type 2 diabetes (6.6 ± 0.6 vs. 10.2 ± 1.1 μmol · kg−1 · min−1 in nondiabetic volunteers, P = 0.01) due to reduced HGR (3.9 ± 0.5 vs. 8.6 ± 1.0 μmol · kg−1 · min−1 in nondiabetic volunteers, P = 0.0015). In contrast, RGR was increased approximately twofold in type 2 diabetes (3.3 ± 0.5 vs. 1.6 ± 0.3 μmol · kg−1 · min−1 in nondiabetic volunteers, P = 0.015). Plasma epinephrine, lactate, and free fatty acid concentrations, which would promote RGR, were also greater in type 2 diabetes (all P < 0.01). Our results provide further support for hepatorenal reciprocity and may explain at least in part the relatively low occurrence of severe hypoglycemia in type 2 diabetes compared with type 1 diabetes where both HGR and RGR counterregulatory responses are reduced.
The Presence of a Catalytically Inactive Form of Hormone-Sensitive Lipase Is Associated With Decreased Lipolysis in Abdominal Subcutaneous Adipose Tissue of Obese Subjects
Authors: Ray Hind , Beylot Michel , Arner Peter , Larrouy Dominique , Langin Dominique , Holm Cecilia , Large Valérie
Published: March 2007

Hormone-sensitive lipase (HSL)-L is a key enzyme in the mobilization of fatty acids from triglyceride stores in adipocytes. A shorter variant of HSL (HSL-S) was detected in humans. This one is generated through in-frame skipping of exon 6 during the processing of HSL mRNA and results in a protein devoid of lipase activity. The role of HSL-S is unknown. The aims of this study were to identify both HSL variants in adipose tissue biopsies and to determine if the presence of HSL-S is correlated to the lipolytic capacity of adipocytes. The study was performed in human abdominal subcutaneous adipocytes from two groups of seven obese subjects. In the group of subjects with both HSL proteins (L+S) group, two immunoreactive bands (80 and 88 kDa) were detected, whereas only the 88-kDa protein was detected in the group with only the wild-type HSL-protein (L group). In the L+S group, the HSL activity was 20% lower (P < 0.05) and the (S/S+) HSL mRNA ratio was twofold higher than in the L group (P < 0.05). The maximally lipolytic capacities measured from isolated adipocytes incubated with norepinephrine or other lipolytic agents were 40% lower in the L+S group (P < 0.05). These results suggest that the presence of the truncated HSL protein is associated with an impaired adipocyte lipolysis.