bluuebunny/crossref_metadata_2025 huggingface dataset:
TNF-α-Induced Insulin Resistance In Vivo and Its Prevention by Troglitazone
Authors: Miles Philip D G , Romeo Oreste M , Higo Katsuya , Cohen Aaron , Rafaat Karim , Olefsky Jerrold M
Published: September 2013

Tumor necrosis factor (TNF)-α may play a role in the insulin resistance of obesity and NIDDM. Troglitazone is a new orally active hypoglycemic agent that has been shown to ameliorate insulin resistance and hyperinsu-linemia in both diabetic animal models and NIDDM subjects. To determine whether this drug could prevent the development of TNF-α-induced insulin resistance, glucose turnover was assessed in rats infused with cytokine and pretreated with troglitazone. Normal male Sprague-Dawley rats were fed normal powdered food with or without troglitazone as a food admixture (0.2%). After ∼10 days, rats were infused with TNF-a for 4–5 days, producing a plasma concentration of 632 ± 30 pg/ml. In vivo insulin action was measured by the euglycemic-hyperinsulinemic clamp technique at a sub-maximal (24 μmol · kg−1 · min−1) and maximal insulin infusion rate (240 μmol · kg−1 · min−1). TNF-α infusion resulted in a pronounced reduction in submaximal insulin-stimulated glucose disposal rate (GDR) (97 ± 10 vs. 141 ± 4 μmol · kg−1 · min−1 P < 0.05), maximal GDR (175 ± 8 vs. 267 ± 6 μmol · kg−1 · min−1 P < 0.01), and in insulin receptor-tyrosine kinase activity (IR-TKA) (248 ± 39 vs. 406 ± 32 pmol ATP/pmol IR, P < 0.05). It also led to a marked increase in basal insulin (90 ± 24 vs. 48 ± 6 pmol/1, P < 0.05) and free fatty acid (FFA) concentration (2.56 ± 0.76 vs. 0.87 ± 0.13 mmol/1, p < 0.01). Troglitazone treatment completely prevented the TNF-α-induced decline in submaximal GDR (133 ± 16 vs. 141 ± 4 umol · kg−1 · min−1, NS) and maximal GDR (271 ± 19 vs. 267 ± 6 μmol · kg1 · min1, NS). The hyperlipidemia was partially corrected by troglitazone (1.53 ± 0.28 vs. 0.87 ± 0.13 mmol/1, P < 0.05), while IR-TKA and insulin concentration remained unaffected by the drug. Troglitazone restores insulin action possibly by lowering the FFA concentration of the blood and/or by stimulating glucose uptake at an intracellular point distal to insulin receptor autophosphorylation in muscle. If TNF-α plays a role in the development of the obe-sity/NIDDM syndrome, troglitazone may prove useful in its treatment.
Expression of Nitric Oxide Synthase in Skeletal Muscle: A Novel Role for Nitric Oxide as a Modulator of Insulin Action
Authors: Kapur Sonia , Bédard Sylvain , Marcotte Bruno , Côté Claude H , Marette André
Published: September 2013

Previous studies have shown that nitric oxide synthase (NOS), the enzyme that catalyzes the formation of nitric oxide (NO), is expressed in skeletal muscle. The aim of the present study was to test the hypothesis that NO can modulate glucose metabolism in slow- and fast-twitch skeletal muscles. Calcium-dependent NOS was detected in skeletal muscle, and the enzyme activity was greater in fast-type extensor digitorum longus (EDL) muscles than in slow-type soleus muscles. Both the neuronal-type (nNOS) and endothelial-type (eNOS) enzymes are expressed in resting skeletal muscles. However, nNOS protein was only detected in EDL muscles, whereas eNOS protein contents were comparable in soleus and EDL muscles. NOS expression in muscle cryosections (diaphorase histochemistry) was located in vascular endothelium and in muscle fibers, and the staining was greater in type lib than in type I and Ha fibers. The macrophage-type inducible NOS (iNOS) was not detected in resting muscle, but endotoxin treatment induced its expression, concomitant with elevated NO production. iNOS induction was associated with impaired insulin-stimulated glucose uptake in isolated rat muscles. In vitro, NOS blockade with specific inhibitors did not affect basal or insulin-stimulated glucose transport in EDL or soleus muscles. In contrast, the NO donors GEA 5024 and sodium nitro-prusside induced dose-dependent inhibition (up to 50%) of maximal insulin-stimulated glucose transport in both muscles with minor effects on basal uptake values. GEA 5024 also blunted insulin-stimulated glucose transport and amino acid uptake in cultured L6 muscle cells without affecting insulin binding to its receptor. On the other hand, the permeable cGMP analogue dibutyryl cGMP did not affect muscle glucose transport. These results strongly suggest that NO modulates insulin action in both slow- and fast-type skeletal muscles. This novel autocrine action of NO in muscle appears to be mediated by cGMP-independent pathways.
Prediction of IDDM in the General Population: Strategies Based on Combinations of Autoantibody Markers
Authors: Bingley Polly J , Bonifacio Ezio , Williams Alistair J K , Genovese Stefano , Bottazzo Gian Franco , Gale Edwin A M
Published: September 2013

Strategies for assessing risk of progression to IDDM, based on single and combined autoantibody measurement, were evaluated in 2,855 schoolchildren (median age 11.4 years) and 256 children with newly diagnosed IDDM (median age 10.2 years), recruited to a population-based study in the Oxford region. In 256 children with IDDM, levels of antibodies ≥97.5th centile of the schoolchild population were found in 225 (88%) for islet cell antibodies (ICAs), in 190 (74%) for antibodies to GAD, in 193 (75%) for antibodies to protein tyro-sine phosphatase IA-2 (IA-2), and in 177 (69%) for autoantibodies to insulin (IAAs). Estimates of risk of progression to IDDM within 10 years, derived by comparing the distribution of antibody markers in the two populations (schoolchildren and children with IDDM), were 6.7% (ICAs), 6.6% (GAD antibodies), 5.6% (IA-2 antibodies), and 4.8% (IAAs) for schoolchildren with levels above the 97.5th centile, increasing to 20, 23, 24, and 11%, respectively, for antibody levels >99.5th centile. Most children with IDDM had multiple antibody markers, and 89% of those diagnosed over age 10 years had ≥2 antibodies above the 97.5th centile, as compared against 0.7% of schoolchildren, in whom this combination gave a 27% 10-year estimated risk of IDDM. Risk increased but sensitivity fell as combined antibody thresholds were raised, or the number of antibodies above the threshold was increased. Strategies based on detection of ≥2 antibodies with primary testing for GAD and IA-2 antibodies and second line testing for ICAs and/or IAAs were evaluated. Detection of at least two markers selected from GAD antibodies ≥97.5th centile and/or IA-2 antibodies ≥99.5th centile and/or ICAs ≥97.5th centile identified 0.25% of schoolchildren and 83% of children with newly diagnosed IDDM, with an estimated risk of 71% (95% CI 57–91). Although confirmation from prospective studies is still needed, this analysis suggests that antibody combinations can predict diabetes in the general population.
Relation Between Antioxidant Enzyme Gene Expression and Antioxidative Defense Status of Insulin-Producing Cells
Authors: Tiedge Markus , Lortz Stephan , Drinkgern Jens , Lenzen Sigurd
Published: September 2013

Antioxidant enzyme expression was determined in rat pancreatic islets and RINm5F insulin-producing cells on the level of mRNA, protein, and enzyme activity in comparison with 11 other rat tissues. Although superoxide dismutase expression was in the range of 30% of the liver values, the expression of the hydrogen peroxide-inactivating enzymes catalase and glutathione per-oxidase was extremely low, in the range of 5% of the liver. Pancreatic islets but not RINm5F cells expressed an additional phospholipid hydroperoxide glutathione peroxidase that exerted protective effects against lipid peroxidation of the plasma membrane. Regression analysis for mRNA and protein expression and enzyme activities from 12 rat tissues revealed that the mRNA levels determine the enzyme activities of the tissues. The induction of cellular stress by high glucose, high oxygen, and heat shock treatment did not affect antioxidant enzyme expression in rat pancreatic islets or in RINm5F cells. Thus insulin-producing cells cannot adapt the low antioxidant enzyme activity levels to typical situations of cellular stress by an upregulation of gene expression. Through stable transfection, however, we were able to increase catalase and glutathione peroxidase gene expression in RINm5F cells, resulting in enzyme activities more than 100-fold higher than in nontransfected controls. Catalase-transfected RINm5F cells showed a 10-fold greater resistance toward hydrogen peroxide toxicity, whereas glutathione peroxidase overexpression was much less effective. Thus inactiva-tion of hydrogen peroxide through catalase seems to be a step of critical importance for the removal of reactive oxygen species in insulin-producing cells. Overexpression of catalase may therefore be an effective means of preventing the toxic action of reactive oxygen species.
Inhibition of Food Response to Intracerebroventricular Injection of Leptin Is Attenuated in Rats With Diet-Induced Obesity
Authors: Widdowson P S , Upton R , Buckingham R , Arch J , Williams G
Published: September 2013

The fat-derived hormone, leptin, is thought to regulate adipose tissue mass by acting on the brain to reduce food intake and increase thermogenesis. We have produced obesity in rats more than 8 weeks old by feeding a high-calorie diet and have then examined the inhibitory effect of intracerebroventricularly injected recombi-nant murine leptin on their food intake versus control rats. In control rats, randomized injections of leptin (0.5, 2.0, or 10.0 μg) or sterile saline vehicle into the lateral ventricle produced a dose-dependent reduction in normal laboratory diet consumed 1, 4, and 24 h after the lights were turned off. However, in diet-induced obesity, the dose-dependent inhibition of food intake was observed at 1 h only, and the effect was attenuated. Switching the diet-induced obese rats to a normal laboratory diet 1 week before injections of leptin were commenced resulted in a reduction in the daily food consumption. These data suggest that rats made obese by feeding a high-calorie diet override the normal satiety effects of leptin since when they are returned to a normal laboratory diet, they reduce their calorie intake, possibly as a result of a restoration of the satiety effects of endogenous leptin. However, the fact that the hypophagic response to exogenous leptin is impaired in these rats at this time suggests some residual impairment of the satiety signal, perhaps caused by reduced receptor sensitivity and/or near total occupation of receptors by endogenous leptin molecules, levels of which are raised in plasma.
Elevated Intakes of Supplemental Chromium Improve Glucose and Insulin Variables in Individuals With Type 2 Diabetes
Authors: Anderson Richard A , Cheng Nanzheng , Bryden Noella A , Polansky Marilyn M , Cheng Nanping , Chi Jiaming , Feng Jinguang
Published: September 2013

Chromium is an essential nutrient involved in normal carbohydrate and lipid metabolism. The chromium requirement is postulated to increase with increased glucose intolerance and diabetes. The objective of this study was to test the hypothesis that the elevated intake of supplemental chromium is involved in the control of type 2 diabetes. Individuals being treated for type 2 diabetes (180 men and women) were divided randomly into three groups and supplemented with: 1) placebo, 2) 1.92 μmol (100 μg) Cr as chromium picolinate two times per day, or 3) 9.6 μmol (500 μg) Cr two times per day. Subjects continued to take their normal medications and were instructed not to change their normal eating and living habits. HbA1c values improved significantly after 2 months in the group receiving 19.2 pmol (1,000 pg) Cr per day and was lower in both chromium groups after 4 months (placebo, 8.5 ± 0.2%; 3.85 pmol Cr, 7.5 ± 0.2%; 19.2 pmol Cr, 6.6 ± 0.1%). Fasting glucose was lower in the 19.2-μmol group after 2 and 4 months (4-month values: placebo, 8.8 ± 0.3 mmol/1; 19.2 μmol Cr, 7.1 ± 0.2 mmol/1). Two-hour glucose values were also significantly lower for the subjects consuming 19.2 μmol supplemental Cr after both 2 and 4 months (4-month values: placebo, 12.3 ± 0.4 mmol/1; 19.2 μmol Cr, 10.5 ± 0.2 mmol/1). Fasting and 2-h insulin values decreased significantly in both groups receiving supplemental chromium after 2 and 4 months. Plasma total cholesterol also decreased after 4 months in the subjects receiving 19.2 μmol/day Cr. These data demonstrate that supplemental chromium had significant beneficial effects on HbA1c, glucose, insulin, and cholesterol variables in subjects with type 2 diabetes. The beneficial effects of chromium in individuals with diabetes were observed at levels higher than the upper limit of the Estimated Safe and Adequate Daily Dietary Intake.
Impairment of Cerebrovascular Reactivity in Long-Term Type 1 Diabetes
Authors: Fülesdi Béla , Limburg Martien , Bereczki Dániel , Michels Robert P J , Neuwirth Gyula , Legemate Dink , Valikovics Attila , Csiba László
Published: September 2013

The early preclinical detection of cerebrovascular complications in individuals with diabetes is one of the goals of care described in the St. Vincent Declaration. In accordance with this goal, the aim of the present work was to investigate whether altered cerebral microvascular function in patients suffering from type 1 diabetes can be detected with a transcranial Doppler probe after the administration of acetazolamide. A total of 72 type 1 diabetic patients and 40 healthy control subjects entered the study. Patients were divided into two groups: those with long-term diabetes (disease duration of >10 years, n = 37) and those with short-term diabetes (disease duration of ≤10 years, n = 35). Mean blood-flow velocity in the middle cerebral artery (MCAV) was measured at rest and at 5, 10, 15, and 20 min after intravenous administration of 1 g acetazolamide with a transcranial Doppler probe and expressed as the percentage change from the pretest measurement. The percentage increase in MCAV (cerebrovascular reactivity) was calculated at each time point and compared between the groups. Cerebrovascular reserve capacity (CRC), expressed as the maximal percentage increase of the MCAV, was compared between the groups. Additionally, a reproducibility study of CRC was performed in 10 patients, using intra-class correlations. Cerebrovascular reactivity in the long-term diabetes group was lower (means ± SD: 5 min, 23.4 ± 15.4%; 10 min, 28.8 ± 17.0%; 15 min, 30.0 ± 15.6%; 20 min, 24.2 ± 17.8%) than that of the control subjects (5 min, 43.5 ± 23.9%; 10 min, 55.3 ± 24.0%; 15 min, 56.7 ± 23.8%; 20 min, 54.8 ± 25.9%) and the short-term diabetic patients (5 min, 43.6 ± 25.9%; 10 min, 52.2 ± 27.7%; 15 min, 55.3 ± 32.2%; 20 min, 45.8 ± 35.8%). CRC was lower in the long-term diabetes group than in the control group or the short-term diabetes group. Impairment of cerebrovascular reactivity was associated with retino- and nephropathy and increased levels of fibrinogen. In contrast, CRC was independent from actual glucose, insulin, glycosylated hemoglobin, von Willebrand factor antigen, and α-2 macroglobulin levels. Transcranial Doppler measurements of the changes in MCAV after stimulation with acetazolamide can detect altered cerebral microvascular function in patients with diabetes. Cerebrovascular reactivity and reserve capacity are reduced in patients with long-term diabetes. Further prospective studies should delineate the clinical significance of our results.
Nitric Oxide Stimulates Skeletal Muscle Glucose Transport Through a Calcium/Contraction– and Phosphatidylinositol-3-Kinase–Independent Pathway
Authors: Etgen Garret J , Fryburg David A , Gibbs E Michael
Published: September 2013

Recently published data have provided evidence that nitric oxide (NO) and cyclic guanosine monophosphate (cGMP) are signaling intermediates in the pathway through which muscle contraction stimulates glucose transport. As exercise promotes both NO production and calcium flux, we examined the relationships between NO-stimulated glucose uptake and calcium-, contraction-, and phosphatidylinositol-3-kinase (PI-3-K)-mediated glucose transport in the isolated incubated rat epitrochlearis muscle preparation. The NO donor sodium nitroprusside (SNP; 10 mmol/1) and dibutyryl cGMP (100 μmol/l) accelerated epitrochlearis glucose transport four- to fivefold above basal levels (P < 0.001) in a manner similar to in vitro contractile activity and the calcium releasing agent N-(6-aminohexyl)-5-chloro-l-naphthalenesulfonamide (W7; 100 μmol/1). In the case of SNP, this effect could be completely attributed to an increase in cell surface GLUT4. The effect of SNP on glucose transport was not inhibitable by either wortmannin (1.5 μmol/1) or dantrolene (12.5 μmol/1). Similarly, neither calcium nor contraction stimulation of glucose transport was affected by the NO synthase inhibitors NG-monomethyl-L-arginine (L-NMMA; 100 μmol/1) or 7-nitroindazole (1 mmol/1). Furthermore, whereas SNP raised epitrochlearis cGMP levels tenfold (P < 0.001), neither in vitro contractile activity nor W7 significantly elevated cGMP. These results indicate that NO/cGMP can markedly stimulate skeletal muscle glucose transport by increasing GLUT4 levels at the cell surface by a mechanism that does not depend on activation of PI-3-K. In addition, since calcium/contraction-stimulated glucose transport is not blocked by NO synthase inhibition and did not elevate cGMP, NO/cGMP may be part of a novel pathway that is distinct from both the insulin- and contraction-activated mechanisms.
Tumor Necrosis Factor-α Induces Apoptosis of Human Adipose Cells
Authors: Prins Johannes B , Niesler Carola U , Winterford Clay M , Bright Nicholas A , Siddle Kenneth , O'Rahilly Stephen , Walker Neal I , Cameron Donald P
Published: September 2013

Tumor necrosis factor-α (TNF-α) production by adipocytes is elevated in obesity, as shown by increased adipose tissue TNF-α mRNA and protein levels and by increased circulating concentrations of the cytokine. Furthermore, TNF-α has distinct effects on adipose tissue including induction of insulin resistance, induction of leptin production, stimulation of lipolysis, suppression of lipogenesis, induction of adipocyte dedifferentiation, and impairment of preadipocyte differentiation in vitro. Taken together, these effects all tend to decrease adipocyte volume and number and suggest a role for TNF-α in limiting increase in fat mass. The aim of the present study was to determine if TNF-α could induce apoptosis in human adipose cells, hence delineating another mechanism by which the cytokine could act to limit the development of, or extent of, obesity. Cultured human preadipocytes and mature adipocytes in explant cultures were exposed in vitro to human TNF-α at varying concentrations for up to 24 h. Apoptosis was assessed using morphological (histology, nuclear morphology following acridine orange staining, electron microscopy) and biochemical (demonstration of internucleosomal DNA cleavage by gel electrophoresis and of annexin V staining using immunocytochemistry) criteria. In control cultures, apoptotic indexes were between 0 and 2.3% in all experiments. In the experimental systems, TNF-α induced apoptosis in both preadipocytes and adipocytes, with indexes between 5 and 25%. Therefore, TNF-α induces apoptosis of human preadipocytes and adipocytes in vitro. In view of the major metabolic role of TNF-α in human adipose tissue, and the knowledge that adipose tissue is dynamic (with cell acquisition via preadipocyte replication/differentiation and cell loss via apoptosis), these findings describe a further mechanism whereby adipose tissue mass may be modified by TNF-α.
The Relation of Proinsulin, Insulin, and Proinsulin-to-Insulin Ratio to Insulin Sensitivity and Acute Insulin Response in Normoglycemic Subjects
Authors: Mykkänen Leena , Haffner Steven M , Hales C Nick , Rönnemaa Tapani , Laakso Markku
Published: September 2013

Plasma levels of proinsulin and its conversion intermediates are elevated in NIDDM patients. Recent studies have suggested that proinsulin levels are also increased relative to insulin levels in subjects who subsequently develop NIDDM. This may be due to insulin resistance or a defect in proinsulin processing or insulin secretion. If insulin resistance is the trigger, the proinsulinto-insulin ratio would be higher in insulin-resistant subjects than in insulin-sensitive subjects. We examined the association of fasting proinsulin, 32,33 split proinsulin, and the proinsulin-to-insulin ratio with insulin sensitivity (SI), estimated by a frequently sampled intravenous glucose tolerance test and the minimal model in 138 normoglycemic subjects ages 53–61 years. We also investigated the relation of proinsulins and the proinsulin-to-insulin ratio to acute insulin response (AIR). Fasting specific insulin (r = −0.64), intact proinsulin (r = −0.43), and 32,33 split proinsulin (r = −0.54) concentrations were inversely correlated and the proinsulin-to-insulin ratio positively (r = 0.31) correlated with SI (P < 0.001). Fasting specific insulin (r = 0.64), intact proinsulin (r = 0.35), and 32,33 split proinsulin (r = 0.45) concentrations were positively correlated and proinsulin-to-insulin ratio (r = −0.40) inversely correlated with AIR (P < 0.001). The proinsulin-to-insulin ratio increased by increasing levels of SI (quartiles of 5, from low to high: 0.048, 0.078, 0.078, 0.068; P = 0.012) and decreased by increasing SI (quartiles of SI from low to high: 0.088, 0.068, 0.058, 0.058; P = 0.005). These associations were independent of age, sex, BMI, and waist-to-hip ratio. Furthermore, the relation between the proinsulin-to-insulin ratio and AIR was independent of SI In conclusion, in normoglycemic subjects, insulin resistance (low SI was associated with a low rather than a high proinsulin-to-insulin ratio. Subjects who maintained normoglycemia with a low AIR had an increased proinsulin-to-insulin ratio compared with those who needed high to maintain normoglycemia. These results suggest that, in subjects with normal glucose tolerance, insulin resistance does not induce increased proinsulin relative to insulin secretion, but rather is associated with enhanced processing of proinsulin.
Diet-Induced Muscle Insulin Resistance in Rats Is Ameliorated by Acute Dietary Lipid Withdrawal or a Single Bout of Exercise: Parallel Relationship Between Insulin Stimulation of Glucose Uptake and Suppression of Long-Chain Fatty Acyl-CoA
Authors: Oakes Nicholas D , Bell Kim S , Furler Stuart M , Camilleri Souad , Saha Asish K , Ruderman Neil B , Chisholm Donald J , Kraegen Edward W
Published: September 2013

Chronic high-fat feeding in rats induces profound whole-body insulin resistance, mainly due to effects in oxidative skeletal muscle. The mechanisms of this reaction remain unclear, but local lipid availability has been implicated. The aim of this study was to examine the influence of three short-term physiological manipulations intended to lower muscle lipid availability on insulin sensitivity in high-fat–fed rats. Adult male Wistar rats fed a high-fat diet for 3 weeks were divided into four groups the day before the study: one group was fed the normal daily high-fat meal (FM); another group was fed an isocaloric low-fat high-glucose meal (GM); a third group was fasted overnight (NM); and a fourth group underwent a single bout of exercise (2-h swim), then were fed the normal high-fat meal (EX). In vivo insulin action was assessed using the hyperinsulinemic glucose clamp (plasma insulin 745 pmol/l, glucose 7.2 mmol/l). Prior exercise, a single low-fat meal, or fasting all significantly increased insulin-stimulated glucose utilization, estimated at either the whole-body level (P < 0.01 vs. FM) or in red quadriceps muscle (EX 18.2, GM 28.1, and NM 19.3 vs. FM 12.6 ± 1.1 μmol · 100 g−1 · min−1; P < 0.05), as well as increased insulin suppressibility of muscle total long-chain fatty acyl-CoA (LCCoA), the metabolically available form of fatty acid (EX 24.0, GM 15.5, and NM 30.6 vs. FM 45.4 nmol/g; P < 0.05). There was a strong inverse correlation between glucose uptake and LC-CoA in red quadriceps during the clamp (r = −0.7, P = 0.001). Muscle triglyceride was significantly reduced by short-term dietary lipid withdrawal (GM −22 and NM −24% vs. FM; P < 0.01), but not prior exercise. We concluded that muscle insulin resistance induced by high-fat feeding is readily ameliorated by three independent, short-term physiological manipulations. The data suggest that insulin resistance is an important factor in the elevated muscle lipid availability induced by chronic high-fat feeding.
Cholesteryl Ester Transfer Protein Gene Polymorphism Is a Determinant of HDL Cholesterol and of the Lipoprotein Response to a Lipid-Lowering Diet in Type 1 Diabetes
Authors: Dullaart Robin P F , Hoogenberg Klaas , Riemens Stephen C , Groener Johanna E M , Tol Arie van , Sluiter Wim J , Stulp Ben K
Published: September 2013

The TaqlB cholesteryl ester transfer protein (CETP) gene polymorphism (B1B2) is a determinant of HDL cholesterol in nondiabetic populations. Remarkably, this gene effect appears to be modified by environmental factors. We evaluated the effect of this polymorphism on HDL cholesterol levels and on the lipoprotein response to a linoleic acid-enriched, low-cholesterol diet in patients with type 1 diabetes. In 44 consecutive type 1 diabetic patients (35 men), CETP polymorphism, apolipoprotein (apo) E genotype, serum lipoproteins, serum CETP activity (measured with an exogenous substrate assay, n = 30), clinical variables, and a diet history were documented. The 1-year response to diet was assessed in 14 type 1 diabetic patients, including 6 B1B1 and 6 B1B2 individuals. HDL cholesterol was higher in 10 B2B2 than in 14 B1B1 homozygotes (1.63 ± 0.38 vs. 1.24 ± 0.23 mmol/l, P < 0.01). HDL cholesterol, adjusted for triglycerides and smoking, was 0.19 mmol/l higher for each B2 allele present. CETP activity levels were not significantly different between CETP genotypes. Multiple regression analysis showed that VLDL + LDL cholesterol was associated with dietary polyunsaturated: saturated fatty acids ratio (P < 0.02) and total fat intake (P < 0.05) in the B1B1 homozygotes only and tended to be related to the presence of the apo E4 allele (P < 0.10). In response to diet, VLDL + LDL cholesterol fell (P < 0.05) and HDL cholesterol remained unchanged in 6 B1B1 homozygotes. In contrast, VLDL + LDL cholesterol was unaltered and HDL cholesterol decreased (P < 0.05) in 6 B1B2 heterozygotes (P < 0.05 for difference in change in VLDL + LDL/HDL cholesterol ratio). This difference in response was unrelated to the apo E genotype. Thus, the TaqlB CETP gene polymorphism is a strong determinant of HDL cholesterol in type 1 diabetes. This gene effect is unlikely to be explained by a major influence on the serum level of CETP activity, as an indirect measure of CETP mass. Our preliminary data suggest that this polymorphism may be a marker of the lipoprotein response to dietary intervention.
Pyruvate Improves Deleterious Effects of High Glucose on Activation of Pentose Phosphate Pathway and Glutathione Redox Cycle in Endothelial Cells
Authors: Kashiwagi Atsunori , Nishio Yoshihiko , Asahina Takayuki , Ikebuchi Motoyoshi , Harada Natsuki , Tanaka Yasushi , Takahara Noriko , Taki Hideki , Obata Toshiyuki , Hidaka Hideki , Saeki Yukikazu , Kikkawa Ryuichi
Published: September 2013

In our previous study (Diabetes 44:520–526, 1995), endothelial cells cultured in high glucose condition showed impairment of an oxidant-induced activation of the pentose phosphate pathway (PPP) and a reduced supply of NADPH to the glutathione redox cycle. To gain insight into the mechanisms of this impairment, the protective effect of pyruvate was studied in human umbilical vein endothelial cells cultured in either 5.5 mmol/l glucose (normal glucose [NG] condition) or 33 mmol/l glucose (high glucose [HG] condition). Through pretreatment of cells with 0.2 mmol/l pyruvate for 5–7 days in the HG condition, glucose oxidation through the PPP and total cellular NADPH content in the presence of 0.2 mmol/l H2O2 were increased by 54 (P < 0.05) and 34%, respectively, and glutathione-dependent degradation of H2O2 in HG cells was enhanced by 41% (P < 0.01), when compared with those cells to which pyruvate was not added. The addition of pyruvate significantly reduced the fructose 1,6-bisphosphate (FDP) content and free cytoplasmic NADH/NAD ratio, estimated by increased pyruvate/lactate ratio in NG and HG cells exposed to H2O2. Furthermore, the addition of pyruvate also showed a 46% reduction (P < 0.01) of endothelial cell damage induced by H2O2 in HG cells. These results indicate that abnormalities in PPP activation and glutathione redox cycle activity induced by H2O2 in HG cells are compensated, and that the accentuated reductive stress is improved by an addition of pyruvate. These pyruvate effects are associated with protection against an oxidant-induced endothelial cell injury in the high glucose condition.
Abnormal Regulation of Hepatic Glucose Output in Maturity-Onset Diabetes of the Young Caused by a Specific Mutation of the Glucokinase Gene
Authors: Tappy Luc , Dussoix Philippe , Iynedjian Patrick , Sylvie Henry , Schneiter Philippe , Zahnd Gaston , Jéquier Eric , Jacques Philippe
Published: September 2013

A subtype of maturity-onset diabetes of the young (MODY) is caused by mutations of the glucokinase gene, an enzyme expressed in pancreatic β-cells and the liver. To assess the consequences of a functional alteration of glucokinase at the level of the liver, endogenous (hepatic) glucose production and glucose cycling (an indirect assessment of hepatic glucokinase activity) were measured with 2-2H glucose and 6,6-2H glucose in patients who developed MODY because of the V203A mutation of glucokinase, and in control subjects at similar levels of glycemia. Measurements were performed in the postabsorptive state and after ingestion of 13C-labeled glucose. In the postabsorptive state, MODY patients had normal glucose production (10.9 ± 1.3 vs. 11.3 ± 0.6 μmol · kg−1 · min−1) but decreased glucose cycling (0.6 ± 0.3 vs. 1.5 ± 0.3 μmol · kg−1 · min−1; P < 0.05) when compared with control subjects. However, at plasma glucose and insulin levels similar to those observed in MODY patients, control subjects' glucose production was markedly lower (3.2 ± 1.5 μmol · kg−1 · min−1). After glucose ingestion, endogenous glucose production was reduced by only 29% in MODY patients compared with 80% in control subjects at a similar level of hyperglycemia (P < 0.05). This suggests that the V203A mutation of glucokinase results in decreased activity of glucokinase in liver cells. Thus endogenous glucose production is inadequately inhibited by hyperglycemia in MODY patients, possibly as a result of impaired hepatic glucokinase activity. These alterations contribute to the pathogenesis of hyperglycemia.
Reduced Insulin, GLUT2, and IDX-1 in β-Cells After Partial Pancreatectomy
Authors: Zangen David H , Bonner-Weir Susan , Lee Chung H , Latimer John B , Miller Christopher P , Habener Joel F , Weir Gordon C
Published: September 2013

Reduction of GLUT2 is associated with loss of glucoseinduced insulin secretion in genetic and chemical diabetes and in transplanted islets exposed to chronic hyperglycemia. To examine the mechanisms for this loss of GLUT2 in normal islets exposed to hyperglycemia, we performed studies on Sprague Dawley rats 4 weeks after a 90% partial pancreatectomy (Px), a well-characterized model of hyperglycemia. GLUT2 immunofluorescence in the β-cell of Px rats was greatly reduced. Western blot analysis of homogenates of isolated Px islets also showed a reduction in GLUT2 protein; densitometry measurements were 36 ± 3% of values from islets of sham-operated controls. Insulin protein levels were decreased to a similar extent. Islet GLUT2 and insulin mRNA were measured with quantitative reverse transcriptase–polymerase chain reaction. The level of GLUT2 mRNA from Px islets was 24 ± 4% of that of islets from sham-operated controls; similar results were obtained for insulin. Because both these (β-cell–specific messages were reduced, we analyzed the Px islets for the pancreas-duodenum–specific transcription factor IDX-1(IPF-1, STF-1, PDX-1) protein. It was markedly reduced (∼80%) in islets from the Px rats. These data suggest that 1) the loss of GLUT2 protein associated with hyperglycemia is at least partially explained by reduced levels of the GLUT2 gene transcripts; 2) the reduction of β-cell insulin content during chronic hyperglycemia may not be completely due to degranulation (reduced levels of gene transcripts may play a role); and 3) the reduction in the transcription factor IDX-1 raises the possibility that dysregulation of transcription factors may contribute to the abnormal β-cell function found in states of chronic hyperglycemia.
Reduction of Postprandial Hyperglycemia and Frequency of Hypoglycemia in IDDM Patients on Insulin-Analog Treatment
Authors: Anderson James H , Brunelle Rocco L , Koivisto Veikko A , Pfützner Andreas , Trautmann Michael E , Vignati Louis , DiMarchi Richard
Published: September 2013

Insulin lispro, an insulin analog recently developed particularly for mealtime therapy, has a fast absorption rate and a short duration of action. We compared insulin lispro and regular human insulin in the mealtime treatment of 1,008 patients with IDDM. The study was a 6-month randomized multinational (17 countries) and multicenter (102 investigators) clinical trial performed with an open-label crossover design. Insulin lispro was injected immediately before the meal, and regular human insulin was injected 30–45 min before the meal. Throughout the study, the postprandial rise in serum glucose was significantly lower during insulin lispro therapy. At the endpoint, the postprandial rise in serum glucose was reduced at 1 h by 1.3 mmol/l and at 2 h by 2.0 mmol/l in patients treated with insulin lispro (P < 0.001). The rate of hypoglycemia was 12% less with insulin lispro (6.4 ± 0.2 vs. 7.2 ± 0.3 episodes/30 days, P < 0.001), independent of basal insulin regimen or HbA1c level. The reduction was observed equally in episodes with and without symptoms. When the total number of episodes for each patient was analyzed according to the time of occurrence, the number of hypoglycemic episodes was less with insulin lispro than with regular human insulin therapy during three of four quarters of the day (P < 0.001). The largest relative improvement was observed at night. In conclusion, insulin lispro improves postprandial control, reduces hypoglycemic episodes, and improves patient convenience, compared with regular human insulin, in IDDM patients.
Polymorphism in the 5′-End of the Aldose Reductase Gene Is Strongly Associated With the Development of Diabetic Nephropathy in Type I Diabetes
Authors: Heesom Angela E , Hibberd Martin L , Millward Ann , Demaine Andrew G
Published: September 2013

Recent studies suggest that the gene encoding aldose reductase (ALR2), the enzyme that converts glucose to sorbitol, may confer susceptibility to microvascular disease. DNA from 275 British Caucasian patients with type I diabetes and 102 normal healthy control patients were typed for a (CA)n dinucleotide repeat polymorphic marker in the 5′-region of the ALR2 gene using polymorase chain reaction (PGR). A highly significant decrease in the frequency of the Z+2 allele was found in patients with nephropathy (nephropathy group) compared with those with no complications after a 20-year duration of diabetes (uncomplicated group) (12.7 vs. 38.2%, respectively, Χ2 = 18.6, P < 0.00001); this was accompanied by an increase in the Z–2 allele in the nephropathy group (32.0 vs. 12.7% in the uncomplicated group). The nephropathy group also had a significant decrease in the Z/Z+2 genotype compared with the uncomplicated patients (10.7 vs. 44.7%, Χ2 = 16.0, P < 0.0001) and an increased frequency of the Z/Z-2 genotype. There was no significant association with diabetic retinopathy. These results demonstrate that the ALR2 gene may play a role in susceptibility to diabetic nephropathy; individuals with the Z+2 allele are more than seven times less likely to develop diabetic renal disease than those without this marker. This marker may prove valuable in screening for patients with diabetic nephropathy at diagnosis of diabetes.
How to Protect the Kidney in Diabetic Patients: With Special Reference to IDDM
Authors: Mogensen CE
Published: September 2013

During the development to overt nephropathy, diabetic patients go through several characteristic stages of renal disease, moving from normo- to micro- to macroalbuminuria. Microalbuminuria is defined as a urinary albumin excretion between 20 and 200 μg/min; values <20 μg/min are designated as normoalbuminuria, and values >200 μg/min are designated as macroalbuminuria. Only with macroalbuminuria does the glomerular filtration rate (GFR) fall consistently. The decisive intermediary endpoints are postponement or prevention of micro/macroalbuminuria and reduction or prevention of the fall in GFR (stronger endpoint), with postponement of end-stage renal disease as a final end-point. Good metabolic control can prevent or postpone the development of microalbuminuria, the earliest sign of diabetic renal disease. The ideal realistic therapeutic window may be an HbA1c value between 7 and 8.5% (mean reference value 5.5%). Thus, efforts should aimed at implementing the best possible control before the onset of microalbuminuria, with the other important aim of minimizing hypoglycemic side effects. In patients with microalbuminuria, blood pressure gradually increases, and early antihypertensive treatment becomes crucial. Good glycemic control (with the same glycemic goal as above) may be difficult to achieve in some of these patients, but it is still important. With overt nephropathy, defined as clinical proteinuria, a relentless decline in GFR is inflicted, unless patients are carefully treated with antihypertensive agents, often in combination therapy. Good metabolic control is still strongly warranted because patients with high HbA1c progress much more rapidly. The natural history of the rate of fall in GFR may be reduced from 12 to 3 ml · min−1 · year−1, but genetic factors may be involved; the ACE-genotype DD seems to progress more rapidly during treatment. Protein restriction is also of some interest. Early screening is recommended in all guidelines, with emphasis on testing for albuminuria, including microalbuminuria, along with careful control of glycemia and blood pressure.
The Roles of Oxidative Stress and Antioxidant Treatment in Experimental Diabetic Neuropathy
Authors: Low Phillip A , Nickander Kim K , Tritschler Hans J
Published: September 2013

Oxidative stress is present in the diabetic state. Our work has focused on its presence in peripheral nerves. Antioxidant enzymes are reduced in peripheral nerves and are further reduced in diabetic nerves. That lipid peroxidation will cause neuropathy is supported by evidence of the development of neuropathy de novo when normal nerves are rendered α-tocopherol deficient and by the augmentation of the conduction deficit in diabetic nerves subjected to this insult. Oxidative stress appears to be primarily due to the processes of nerve ischemia and hyperglycemia auto-oxidation. The indexes of oxidative stress include an increase in nerve, dorsal root, and sympathetic ganglia lipid hydroperoxides and conjugated dienes. The most reliable and sensitive index, however, is a reduction in reduced glutathione. Experimental diabetic neuropathy results in myelinopathy of dorsal roots and a vacuolar neuropathy of dorsal root ganglion. The vacuoles are mitochondrial; we posit that lipid peroxidation causes mitochondrial DNA mutations that increase reduced oxygen species, causing further damage to mitochondrial respiratory chain and function and resulting in a sensory neuropathy, α-lipoic acid is a potent antioxidant that prevents lipid peroxidation in vitro and in vivo. We evaluated the efficacy of the drug in doses of 20, 50, and 100 mg/kg administered intraperitoneally in preventing the biochemical, electrophysiological, and nerve blood flow deficits in the peripheral nerves of experimental diabetic neuropathy, α-lipoic acid dose- and time-dependently prevented the deficits in nerve conduction and nerve blood flow and biochemical abnormalities (reductions in reduced glutathione and lipid peroxidation). The nerve blood flow deficit was 50% (P < 0.001). Supplementation dose-dependently prevented the deficit; at the highest concentration, nerve blood flow was not different from that of control nerves. Digital nerve conduction underwent a dose-dependent improvement at 1 month (P < 0.05). By 3 months, all treated groups had lost their deficit. The antioxidant drug is potentially efficacious for human diabetic sensory neuropathy.
The Pathology of Human Diabetic Neuropathy
Authors: Malik RA
Published: September 2013

Quantitative morphological data are presented from a series of studies assessing both nerve fiber and capillary pathology in 30 diabetic patients with varying stages and symptoms of neuropathy. There is a significant relationship between clinical measures of neuropathic severity and myelinated fiber loss. However, unmyelinated fibers continue to regenerate even in patients with established neuropathy. Microvascular abnormalities, particularly basement membrane thickening and endothelial cell hyperplasia, are an early feature of diabetic microangiopathy and relate to neuropathic severity. There are no neurophysiological or morphological differences between patients with type 1 and type 2 diabetes nor between diabetic patients with and without painful neuropathy.
Aldose Reductase Inhibitors: The End of an Era or the Need for Different Trial Designs?
Authors: Pfeifer Michael A , Schumer Mary P , Gelber David A
Published: September 2013

Despite numerous attempts over 16 years, the results of aldose reductase inhibitor (ARI) trials for the treatment of diabetic neuropathy have not proven efficacy. This paper reviews each of the ARI trials, examines confounding factors, and proposes a future course. The confounding factors considered are pharmacokinetics (ARI penetration of human nerve), length of trial (in terms of the natural history of diabetic neuropathy), trial endpoints (reversibility or slowing of progression), reproducibility of clinical measurements (in terms of power calculations), standardization and quality control of endpoints, and clinically meaningful differences in endpoints. We conclude that ARIs are most likely to have a beneficial effect in the management of diabetic distal symmetrical polyneuropathy and autonomic neuropathy but that the clinical role of ARIs is to slow the progression of diabetic neuropathy rather than to reverse it. Future trials should be designed with adequate statistical power, with consideration of the variability of the endpoint measurements for long enough duration, and with rigorous quality control to definitively confirm the utility of ARIs in the treatment of diabetic distal symmetrical polyneuropathy and autonomic neuropathy.
Pathogenesis of Diabetic Vascular Disease: Evidence for the Role of Reduced Heparan Sulfate Proteoglycan
Authors: Jensen Tonny
Published: September 2013

Insulin-dependent diabetic patients with increased urinary albumin excretion are characterized by elevated blood pressure and declining kidney function. In addition, such patients have a high risk of atherosclerotic vascular disease, proliferative retinopathy, and cardiomyopathy, suggesting that albuminuria is a marker of widespread vascular dysfunction. Increased transport of macromolecules across the vascular wall, elevated plasma levels of von Willebrand factor, and impaired fibrinolytic capacity have been demonstrated in albuminuric patients. The cause of this vascular vulnerability in susceptible patients is unknown, but increasing evidence has suggested that loss of the proteoglycan heparan sulfate in the vasculature may explain the widespread nature of the disease. Heparan sulfate is important for the glomerular endothelial cell and basement membrane charge densities, the anticoagulant properties of the vessel wall, and the growth regulation of intimal smooth muscle cells. Recent studies have shown that heparin increases the biosynthesis of heparan sulfate in endothelial cell cultures and prevents the characteristic glomerular basement membrane thickening when given to diabetic rats. Moreover, heparin has been shown to reduce albuminuria in patients with incipient diabetic nephropathy. Although increasing evidence supports the hypothesis that loss of heparan sulfate may play a pathophysiological role in the development of diabetic vascular complications, there are still many unresolved problems. What are the mechanisms of action of glycosaminoglycans at the molecular biology level, and how can we select compounds without anticoagulant activity suitable for long-term use in the prevention and treatment of late diabetic complications?
Long-Chain Fatty Acids Inhibit Acetyl-CoA Carboxylase Gene Expression in the Pancreatic β-Cell Line INS-1
Authors: Brun Thierry , Assimacopoulos-Jeannet Françhise , Corkey Barbara E , Prentki Marc
Published: September 2013

The mechanism whereby long-term exposure of the β-cell to fatty acids alters the β-cell response to glucose is not known. We hypothesized that fatty acids may alter β-cell function by changing the expression level of metabolic enzymes implicated in the regulation of insulin secretion, in particular acetyl-CoA carboxylase (ACC). This enzyme catalyzes the formation of malonyl-CoA, a key regulator of fatty acid oxidation. Using the β-cell line INS-1 as a model, the results show that the polyunsaturated fatty acid linoleate (C18:2) inhibited both basal and glucose-stimulated ACC mRNA induction. The inhibition was detected by 4–6 h, and a maximal 60% effect occurred at 12 h after cell exposure to the fatty acid. Linoleate, as glucose, did not modify the half-life of the ACC transcript. Prolonged exposure of INS-1 cells to linoleate also inhibited ACC protein accumulation at low and high glucose. The saturated fatty acids myristate (C14:0), palmitate (C16:0), and stearate (C18:0) were also effective as well as the monounsaturated oleate (C18:1) and the short-chain fatty acids butyrate (C4:0) and caproate (C6:0); longchain ω3 fatty acids were ineffective. The threshold concentration for long-chain fatty acids was 0.05 mmol/l, and maximal inhibition occurred at 0.3 mmol/l. 2-bromopalmitate, a nonmetabolizable analog, had no effect, suggesting that fatty acids must be metabolized to change ACC gene expression. Prolonged exposure of INS-1 cells to palmitate, oleate, and linoleate markedly altered the glucose-induced insulin response, resulting in high basal insulin release and a suppression of glucose-induced insulin secretion. This was associated with an exaggerated (twofold to threefold) rate of fatty acid oxidation at all tested glucose concentrations. The data provide a possible mechanism to at least partially explain how fatty acids cause β-cell insensitivity to glucose, i.e., by downregulating ACC with a resulting exaggerated fatty acid oxidation.