bluuebunny/crossref_metadata_2025 huggingface dataset:
Effect of Insulin on Metabolism of Fetal Sheep Hindquarters
Authors: Milley J Ross , Papacostas Julie S
Published: September 2013

Fetal hyperinsulinemia causes fetal arterial hypoxia because fetal O2 use increases, whereas the supply of O2 to the fetus does not. To find which of the fetal tissues accounts for such an increase in fetal O2 use, we examined the effect of plasma hyperinsulinemia on O2, glucose, and lactate use by the hindquarters of 13 fetal sheep. Spinal anesthesia was used for the ewes, and local anesthesia was used for the fetuses during placement of catheters. The ewes then recovered for 5 days. After 18 h of insulin infusion, blood samples were drawn, and microspheres were injected to measure blood flow to the tissues of the hindquarters. Three to five infusions of various insulin concentrations in each fetus were followed by blood sampling and blood-flow measurements. Fetal hyperinsulinemia (≤437 pM) increased blood flow to and O2 use by the hindquarters of the fetal sheep but did not affect the glucose-O2 quotients of these tissues. Consequently, glucose use increased proportionately to the increased O2 use. Lactate production was not affected by insulin. We conclude that increased O2 use by all the nonvisceral fetal tissues accounted for the increased O2 use of the entire fetus reported during fetal hyperinsulinemia and, consequently, for the fetal arterial hypoxemia associated with fetal hyperinsulinemia. If the hyperinsulinemic human fetus (such as the infant of the diabetic mother) also increases O2 use in nonvisceral tissue, such an increase might contribute to the susceptibility of the infants to late intrauterine fetal death, polycythemia, and hyperbilirubinemia, all of which may be consequences of intrauterine arterial hypoxemia.
Relation of Insulin Deficiency to Impaired Insulin Action in NIDDM Adult Rats Given Streptozocin as Neonates
Authors: Blondel Olivier , Bailbé Daniele , Portha Bernard
Published: September 2013

We assessed the impact of chronic insulin deficiency on basal and insulin-stimulated glucose utilization by the whole-body mass in vivo in female albino Wistar rats. This assessment was based on a comparison of results in rats given streptozocin (STZ) on day of birth (n0-STZ), when 2 days old (n2-STZ), or when 5 days old (n5-STZ). At 10 wk of age, the n2-STZ rats exhibited characteristics similar to those obtained in the n0-STZ rats: normal growth, modest elevation of basal plasma glucose (8.23 ± 0.24 mM), glucose intolerance, depleted pancreatic insulin stores (∼50% of normal value), and lack of insulin release in response to glucose in vivo. In contrast, the n5-STZ rats exhibited frank basal hyperglycemia (glucose 11.9 ± 1.1 mM) and glucose intolerance, increased glycosylated hemoglobins, strong reduction of the pancreatic insulin stores (10% of normal value), decreased basal plasma insulin levels (50% of normal value), and lack of insulin release in response to glucose in vivo. Changes in the sensitivity of the neonatal β-cell to STZ and the regeneration capacity of the β-cells during the 1st postnatal wk were liable factors for the contrast. In vivo insulin action was assessed with the euglycemic-hyperinsulinemic clamp technique in 10-wk-old anesthetized animals. In the n2-STZ rats compared with controls 1) endogenous glucose production was significantly higher despite a normal plasma insulin level in the basal state, 2) endogenous glucose production rate was similarly suppressed by hyperinsulinemia, and 3) glucose utilization by the whole-body mass was similarly increased by hyperglycemia. Thus, in terms of basal hepatic glucose production, a discrete alteration of insulin action, with mild insulin deficiency, was detected in this model. In the n5-STZ rats compared with controls 1) basal hepatic glucose production was significantly higher, 2) hepatic glucose production rate was less efficiently suppressed by submaximal or maximal insulin levels, and 3) glucose utilization by the whole-body mass induced by hyperinsulinemia was significantly reduced (by 35% according to submaximal or maximal insulin levels). Thus, insulin resistance was present in vivo at the level of both the peripheral tissues (probably the muscles) and the liver of the n5-STZ rat. These data confirm that when β-cell destruction is the primary insult responsible for the emergence of moderate to severe hyperglycemia in rats, insulin resistance can develop secondarily. However, the assumption that insulin deficiency is invariably followed by an overall peripheral insulin resistance is not tenable, because there was no peripheral insulin resistance in the n0-STZ and n2-STZ models. These data are consistent with the suggestion that a certain degree of insulin deficiency is necessary to induce a clear-cut decrease in insulin action.
Antibodies Against Gastrointestinal Carcinoid Tumors in IDDM
Authors: Miettinen Aaro , Holthöfer Harry , Kontiainen Sirkka , Miettinen Markku , Andersson Leif C
Published: September 2013

In this study, 92% of patients'serums known to contain antibodies against islet cells, including the Juvenile Diabetes Foundation provisional reference serum, had antibodies reacting with gastrointestinal carcinoid tumors. Twelve percent of the control serums from healthy individuals bound to carcinoid cells, and 2% bound to islet cells. Seventy-five percent of the children with newly diagnosed insulin-dependent diabetes mellitus had carcinoid tumor antibodies, and 83% had islet cell antibodies. These findings suggest that antigenic determinants are shared between endocrine cells of islets of Langerhans and neuroendocrine tumors of the same embryological derivation. Carcinoid tumors may not only provide an alternative source for islet cell antibody assays but also supply material for isolation of antigens possibly involved in the immunopathogenesis of diabetes.
A New Phase Of Insulin Secretion: How Will It Contribute to Our Understanding of β-Cell Function?
Authors: Grodsky Gerold M
Published: September 2013

Although initially described two decades ago, biphasic insulin secretion has gradually been understood to reflect (β-cell rate sensitivity, be important in minimizing overinsulinization in normal individuals, be defective in non-insulin-dependent diabetes mellitus (NIDDM), and be useful as an early predictor in prediabetic individuals. Recently, a third phase of insulin secretion has been observed in fully in vitro islets or pancreatic preparations. This phase is characterized as a spontaneous decline of secretion (desensitization) during 24 h of sustained exposure to glucose or other secretagogues and does not appear to be simply an artifact of in vitro preparations. The impaired secretion is localized to the final release process in that neither glucose-stimulated proinsulin synthesis nor its conversion to insulin is affected. The mechanisms responsible for the third phase of reduced secretion are unknown. Kinetic evidence suggests it is not caused by emptying of a single finite insulin storage compartment but does not exclude the possibility that the decreased release reflects depletion of threshold-sensitive β-cells recruited at a given secretagogue level. Alternatively, the third phase may reflect inhibition of a priming or terminal insulin-release process by metabolic feedback. Because several secretagogues cause similar third-phase impaired release, even in the absence of glucose, desensitization probably occurs at a common fundamental site in the secretory site (e.g., calcium metabolism). Preliminary studies indicate the third phase is not the result of a paracrine effect by other islet hormones or of a change in muscarinic regulation. Whether other neurologic effectors are involved requires further investigation. Experiments examining the effect of glucose-induced third-phase desensitization on other secretagogues emphasize that priming (potentiation) and desensitization of insulin secretion are occurring simultaneously and that interpretations of results can be highly dependent on the design of the experiment. The possibility that the third phase of insulin secretion may involve mechanisms (waning of priming; desensitization) that relate to glucose-induced β-cell desensitization in NIDDM is discussed. Ultimate appreciation of the significance of the third phase of insulin secretion may develop, as was true for biphasic secretion, with our increasing understanding of the underlying processes in human diabetes.
Effect of Carbohydrate Intake on Kidney Function and Structure in SHR/N-cp Rats: A New Model of NIDDM
Authors: Velasquez Manuel T , Kimmel Paul L , Michaelis Otho E , Carswell Nancy , Abraham Andrew , Bosch Juan P
Published: September 2013

The SHR/N corpulent (cp) rat is a genetically obese rat that develops hyperglycemia, hyperinsulinemia, and proteinuria. This study was designed to evaluate the effects of high carbohydrate (CHO) intake on renal function and structure in this animal model and to determine whether the renal effects are related to the type of CHO ingested. Two groups of 5-wk-old obese male SHR/N-cp rats and lean male littermates were fed diets containing 54% CHO in the form of sucrose or starch. After 12 wk, renal function parameters, including creatinine clearance, urinary glucose excretion, and urinary protein excretion, were measured. Renal morphology was evaluated by semiquantitative light and electron microscopy. On either diet, obese rats had significantly higher urinary glucose and protein excretions than their lean littermates. Mean creatinine clearance (ml/min) in obese rats did not differ significantly from values observed in lean rats. When corrected for body weight, creatinine clearance (ml · min−1 · kg−1) tended to be lower in obese than in lean rats, but the difference was significant (P < .02) only for obese and lean sucrose-fed animals. Obese rats fed sucrose compared with their obese counterparts fed starch had higher body weight (+ 8%, P < .05), glucose excretion (+ 63%, P < .02), and protein excretion (+ 242%, P < .005). In obese rats, protein excretion correlated with glucose excretion (r = .71, P < .01). Glomerular lesions consisting of mesangial expansion and intercapillary nodules were found in obese but not in lean rats.Moreover, obese rats fed sucrose had a significantly greater number of involved glomeruli than obese rats fed starch. There was no evidence of glomerular basement membrane thickening in any of the animals studied. We conclude that the SHR/N-cp rat is sensitive to a high-CHO diet and develops glucosuria, proteinuria, renal insufficiency, and glomerular lesions resembling human diabetic nephropathy and that these characteristics are accentuated by high sucrose intake.
Effects of Cyclosporin on Insulin and C-Peptide Secretion in Healthy Beagles
Authors: Alejandro Rodolfo , Feldman Edward C , Bloom Allen D , Kenyon Norma S
Published: September 2013

Plasma glucose, C-peptide, and insulin responses to intravenous glucose (intravenous glucose tolerance test [IVGTT], 0.5 g/kg), glucagon (1 mg i.v.), and oral glucose (oral glucose tolerance test [OGTT], 1 g/kg) were assessed in six normal beagles before, during, and 1 and 4 mo after the administration of cyclosporin A (CsA) in doses previously shown to be required for uniform prevention of canine islet-allograft rejection (20 mg/kg; mean trough radioimmunoassay serum levels ≥500 ng/ml). Insulin secretion in response to intravenous glucose and glucagon was significantly inhibited during the administration of CsA (areas under insulin-response curves, pmol min−1 L−1; IVGTT, pre-CsA, 11,127 ± 1285; during CsA, 5954 ± 1147, P < .05; glucagon tolerance test, pre-CsA, 18,617 ± 2807; during CsA, 4401 ± 486, P < .05 vs. pretreatment levels). These secretory defects persisted 4 mo after CsA was discontinued (IVGTT, 4358 ± 659; glucagon tolerance test, 10,567 ± 2479, P < .05). C-peptide responses paralleled these changes. Plasma glucose disposal in response to these secretagogues, however, returned to normal 1 mo after discontinuation of CsA. In contrast to the findings for IVGTT and glucagon, insulin-response curves to OGTT were not statistically different during CsA administration. We conclude that, although glucose disappearance rates are normal after discontinuation of the CsA administration, CsA causes irreversible impairment in islet secretory responses detectable with IVGTT and glucagon but not with OGTT. These results suggest that short-term CsA in doses required to prevent islet-allograft rejection in dogs can result in permanent loss of functionally competent β-cells.
Cross-Reactivity of Anti-ssDNA Antibodies With Heparan Sulfate in Patients With Type I Diabetes Mellitus
Authors: Triolo Giovanni , Giardina Ennio , Scarantino Giovanna , Seddio Giovanna , Cardella Francesca , Meli Ferdinando , Bompiani Giandomenico
Published: September 2013

Anti-single-stranded–DNA antibodies cross-reactive with heparan sulfate were detected in serums of patients with type I (insulin-dependent) diabetes mellitus. The results suggested that heparan sulfate, the major glycosaminoglycan constituent of the glomerular basement membrane, may serve as a target antigen in vivo for cross-reactive anti-DNA antibodies. These polyreactive antibodies, directed toward repeating negatively charged units, may neutralize the heparan sulfate–associated polyanionic sites in the glomerulus, leading to an abnormal permeability of anionic plasma proteins.
Double-Blind Controlled Trial of Azathioprine in Children With Newly Diagnosed Type I Diabetes
Authors: Cook Jennifer J , Hudson Irene , Harrison Leonard C , Dean Brian , Colman Peter G , Werther Georgea , Warne Garry L , Court John M
Published: September 2013

A double-blind controlled trial of azathioprine (2 mg · kg−1 · day−1) was conducted with 49 patients aged 2–20 yr (mean 10.8 yr) who had newly diagnosed type I (insulin-dependent) diabetes. Patients were randomly assigned to receive either azathioprine (n = 24) or placebo (n = 25) for 12 mo, beginning within the 20 day period after diagnosis. Baseline clinical and metabolic characteristics did not differ between the two groups. No patient experienced complete remission, defined as restoration of normal carbohydrate tolerance without other treatment. Partial remission, defined as good metabolic control (hemoglobin A1c ≤7.9%, preprandial blood glucose ≤8 mM with an insulin dose of <0.5 U. kg−1 · day1), occurred in 10 placebo (40%) and 7 azathioprine (29%) patients at 6 mo and in 4 placebo (16%) and 4 azathioprine (17%) patients at 12 mo (differences not significant). Fasting plasma C-peptide was significantly greater in the azathioprine-treated group at 3 and 6 mo, but this difference was not sustained. C-peptide responses to a standard meal and the frequency of islet cell and insulin antibodies did not differ between the two groups over the 12-mo period. Azathioprine caused no significant side effects. We conclude that in the dosage used, and despite early effects on endogenous insulin secretion, azathioprine alone does not influence the remission phase in children with newly diagnosed type I diabetes.
Microvascular Blood Flow, Volume, and Velocity Measured by Laser Doppler Techniques in IDDM
Authors: Rendell Marc , Bergman Tom , O'Donnell Greg , Drobny Ed , Borgos John , Bonner Robert F
Published: September 2013

A laser Doppler device with the capability to simultaneously measure skin blood flow, microvascular volume, and erythrocyte velocity was used to assess blood flow changes in 35 insulin-dependent diabetes mellitus (IDDM) subjects, mean age 33 ± 1 yr, with average duration of diabetes 14 ± 1 yr, and in a nondiabetic control group. Blood flow was determined at 35 and 44°C at several sites on the upper and the lower extremities with a temperature-regulated probe. Blood flow was highest at both temperatures on the pulps of the index finger and the first toe, regions of high density of arteriovenous anastomoses. There was significantly greater blood flow at most locations for the nondiabetic than the diabetic group at 35°C, and the differences between the two groups were substantially larger at 44°C. At 44°C, blood flow in the control group was ∼40% greater in the upper extremity and 50% greater in the lower extremity than it was in the diabetic subjects. The differences were attributed to decreases of both microvascular volume and velocity in the diabetic group. In the upper extremity, volumes in the diabetic patients were 10–15% lower and velocities 10–40% lower than in the nondiabetic subjects. In the lower extremity, volumes were 20–25% lower and velocities 40–50% lower. We conclude that laser Doppler techniques can be used to assess microvascular changes in the skin of diabetic patients. This approach may be useful to evaluate and model diabetic microangiopathy.
Indirect Effect of Catecholamines on Development of Insulin Resistance in Skeletal Muscle From Diabetic Rats
Authors: Boström Mikael , Nie Zetan , Goertz Georg , Henriksson Jan , Wallberg-Henriksson Harriet
Published: September 2013

The role of an increased sympathetic activation in the development of insulin resistance in diabetic skeletal muscle was investigated. Epitrochlearis muscles from rats with streptozocin-induced diabetes and from controls were incubated in vitro for 0.5–12.0 h. Diabetes decreased maximal insulin-stimulated (20 mU/ml) glucose transport capacity by 60% (P < .001), but this decreased insulin responsiveness returned to normal on in vitro incubation (3.79 ± 0.59 before vs. 8.92 ± 0.64 μmol · ml1 · h−1 after 12 h of incubation). The reversal of decreased insulin responsiveness in diabetic muscles did not require the presence of insulin and was not affected by the presence of 5.0 × 10−8 M of epinephrine. However, it was possible to partially prevent the development of insulin resistance with regard to glucose transport by treating the rats with the β-adrenergic antagonist propranolol (0.5 mg/kg) every 12 h during the entire 72-h period in which the animals were kept diabetic (insulin responsiveness was 3.16 ± 0.40 μmol · ml−1 · h−1 saline-injected group vs. 5.55 ± 0.46 μmol · ml−1 for propranolol-treated group). This effect was not present after a single injection of the drug 2 h before the experiment or when propranolol treatment was withdrawn 12 h before the experiment. The β-adrenergic blockade markedly reduced the plasma concentration of free fatty acids (0.5 ± 0.01 (μmol/ml for propranolol-treated rats vs. 1.1 ± 0.1 μmol/ml for saline-treated rats; P < .001). These results indicate that catecholamines, acting indirectly by another mediator, possibly free fatty acids, are involved in the development of insulin resistance with regard to glucose transport in diabetic skeletal muscle. Furthermore, the results suggest that the effect of catecholamines is of a chronic, rather than acute, nature.
In Vitro and In Vivo Reduction of Erythrocyte Sorbitol by Ascorbic Acid
Authors: Vinson Joe A , Staretz Marianne E , Bose Pratima , Kassm Hassan M , Basalyga Barbara S
Published: September 2013

The in vitro accumulation of sorbitol by human erythrocytes incubated in a physiological glucose medium was found to be strongly reduced by the addition of ascorbic acid (AA). A maximal inhibition of sorbitol in the erythrocytes of 98.3% occurred when the concentration of AA was at its peak in the cells. After incubation, the erythrocyte sorbitol was found to be inversely correlated with the concentration of AA in the erythrocytes. A human supplementation study was conducted with 10 normoglycemic subjects. Each was given 500 mg/day AA alone or in a citrus fruit medium. Each supplementation lasted 2 wk and was followed by a 10-day washout. The citrus fruit medium produced a significantly greater increase in erythrocyte AA compared with AA alone. AA alone and in citrus fruit medium decreased erythrocyte sorbitol 12.6 and 27.2%, respectively, with the latter being significantly more effective. In a study with 4 subjects, 2000 mg/day AA resulted in a reduction in erythrocyte sorbitol of 56.1%. As in the in vitro study, there was an inverse relationship between erythrocyte AA and sorbitol. Two thousand milligrams of AA per day (AA or citrus fruit medium) was given to 8 diabetic subjects in a preliminary 3-wk supplementation trial in which erythrocyte sorbitol levels were decreased by 44.5%. These results suggest that AA supplementation for diabetic subjects may provide a simple means of preventing and ameliorating the complications of diabetes without the use of drugs.
Stable-Label Intravenous Glucose Tolerance Test Minimal Model
Authors: Avogaro Angelo , Bristow James D , Bier Dennis M , Cobelli Claudio , Toffolo Gianna
Published: September 2013

The minimal model approach to estimating insulin sensitivity (SI) and glucose effectiveness in promoting its own disposition at basal insulin (SG) is a powerful tool that has been underutilized given its potential applications. In part, this has been due to its inability to separate insulin and glucose effects on peripheral uptake from their effects on hepatic glucose inflow. Prior enhancements, with radiotracer labeling of the dosage, permit this separation but are unsuitable for use in pregnancy and childhood. In this study, we labeled the intravenous glucose tolerance test (IVGTT) dosage with [6,6-2H2]glucose, [2-2H]glucose, or both stable isotopically labeled glucose tracers and modeled glucose kinetics in six postabsorptive, nonobese adults. As previously found with the radiotracer model, the tracer-estimated derived from the stable-label IVGTT was greater than SI in each case except one, and the tracer-estimated was less than SG in each instance. More importantly, however, the stable-label IVGTT estimated each parameter with an average precision of ± 5% (range 3–9%) compared to average precisions of ±74% (range 7–309%) for SGand ±22% (range 3–72%) for SI. In addition, because of the different metabolic fates of the two deuterated tracers, there were minor differences in basal insulin-derived measures of glucose effectiveness, but these differences were negligible for parameters describing insulin-stimulated processes. In conclusion, the stable-label IVGTT is a simple, highly precise means of assessing insulin sensitivity and glucose effectiveness at basal insulin that can be used to measure these parameters in individuals of all ages, including children and pregnant women.
Neuraminidase Treatment of Isolated Rat Adipocytes and Differential Regulation of Basal and Insulin-Stimulated Glucose Transport
Authors: Ciaraldi Theodore P
Published: September 2013

The role of membrane carbohydrate in the function of insulin receptors and glucose transporters was investigated with neuraminidase to release sialic acid from isolated rat adipocytes. Pretreatment of adipocytes with neuraminidase resulted in an increase in basal glucose transport. At the same time, insulin-stimulated glucose transport was reduced by an average of 30%. The enzyme action on glucose transport was not due to a nonspecific membrane perturbation because neuraminidase caused only a nonsignificant decrease in the uptake of the amino acid analog α-(methylamino)isobutyric acid and had no effect on basal or insulin-stimulated protein synthesis. Insulin binding was slightly increased in neuraminidase-treated cells, yet the shapes of the dose-response curves for insulin stimulation of glucose transport were similar (EC50 = 0.087 ± 0.010 and 0.082 ± 0.008 nM for control and treated cells, respectively). The neuraminidase-induced increase in basal transport was the result of an increase in the affinity of the glucose-transport system (Km = 7.3 ± 1.4 and 3.6 ± 0.7 mM before and after treatment, respectively) with no change in Vmax. Conversely, enzyme treatment decreased the Vmax of insulin-stimulated 3-O-methylglucose transport from 87.8 ± 13.2 to 41.3 ± 4.9 pmol · 2 × 105 cells−1 · S−1 while not altering the Km. These changes in glucose-transport activity resulting from enzyme treatment were not due to alterations in glucose-transporter concentration on the plasma membrane as measured by the D-glucose-inhibitable binding of [3H]cytochalasin B. Thus, sialic acid plays multiple roles in the control of glucose-transport activity. In the absence of insulin, it places constraints on the affinity of the transporter for substrate, and it may also be involved in maintaining the intrinsic activity of insulin-stimulated transporters, possibly acting through a regulatory protein on the cell surface.
Aldose Reductase Inhibition and Glomerular Abnormalities in Diabetic Rats
Authors: Daniels Barbara S , Hostetter Thomas H
Published: September 2013

We examined the effects of aldose reductase inhibition (ARI) on glomerular filtration rate (GFR), albuminuria, and kidney histology in partially insulin-treated streptozocin-induced diabetic (STZ-D) rats. After 1 mo of diabetes, GFR was elevated over control values in the STZ-D rats but was not affected by treatment with statil (an aldose reductase inhibitor). In another set of rats maintained for 7 mo, albuminuria was significantly increased in the diabetic rats from 2 mo on but was also not affected by statil treatment. Similarly, histological glomerular damage and diabetes-induced kidney hypertrophy were also greater in diabetic animals but were not altered by statil treatment. The frequency of diabetic cataracts was reduced by statil, and erythrocyte and kidney sorbitol levels were normalized, confirming the efficacy of ARI. Thus, inhibition of the aldose reductase pathway with statil does not ameliorate the hemodynamic, proteinuric, histological, or growth abnormalities in this model of diabetic nephropathy.
Strong Association Between Diabetes and Displacement of Mouse Anti-Rat Insulinoma Cell Monoclonal Antibody by Human Serum In Vitro
Authors: Thomas Nancy M , Ginsberg-Fellner Fredda , McEvoy Robert C
Published: September 2013

In an attempt to identify novel pancreatic β-cell surface antigens, mouse monoclonal antibodies (MoAbs) were raised against rat insulinoma (RIN5F) cells with standard techniques. Several clones were identified whose antibodies bound specifically to RIN5F cells but not to other rat, mouse, and human target cells. Each of these MoAbs was radiolabeled, and the specificity of binding of each MoAb was determined by the ability of excess cold homologous MoAb to displace the labeled MoAb. Six RIN5F cell-specific MoAbs of different epitopic specificities were identified. The relevance of these β-cell epitopes to human insulindependent diabetes (IDDM) was demonstrated by the differential ability of human serums from control and diabetic children to displace the radiolabeled MoAbs from the RIN5F cells. Serums from 333 children without diabetes or a family history of diabetes and from 156 newly diagnosed IODM patients were tested. Only one IgM MoAb was specifically displaced by the IDDM serums, i.e., 146 of 156, compared to serums from control children, i.e., 10 of 333. With immunofluorescence, the serum component responsible for the displacement of the mouse MoAb was identified as IgG. Most of the positive control serums were from children with active autoimmune thyroiditis. Serums from children with other forms of glucose intolerance did not displace MoAb 1A2. There was no correlation between age and the degree of displacement of 1A2. Thus, the displacement of 1A2 is a specific and sensitive marker of diabetes susceptibility easily applicable to mass screening. However, this assay detects autoantibodies against an antigen different from the conventional immunofluorescence assay on sections of human pancreas, because the International Diabetes Workshop standard positive serum did not displace 1A2.
Nonenzymatic Glycosylation of HDL Resulting in Inhibition of High-Affinity Binding to Cultured Human Fibroblasts
Authors: Duell P Barton , Oram John F , Bierman Edwin L
Published: September 2013

Nonenzymatic glycosylation of plasma proteins may contribute to the excess risk of developing atherosclerosis in patients with diabetes mellitus. Because high-density lipoprotein (HDL) is believed to protect against atherosclerosis and is glycosylated at increased levels in diabetic individuals, the effects of nonenzymatic glycosylation of HDL3 on binding of HDL3 to cultured fibroblasts and to the candidate HDL-receptor protein were examined. HDL3 was glycosylated in vitro with glucose alone or in combination with sodium cyanoborohydride. With this catalyst, up to 40–50% of the lysine residues could be glycosylated, resulting in a progressive drop to nearly 60% in high-affinity binding to cultured fibr oblasts at 4°C. Binding to the 110,000-Mr candidate HDL-receptor protein was reduced by almost 75%. At levels of HDL glycosylation equivalent to the 3–5% observed in diabetes, high-affinity binding to fibroblasts at 4°C was diminished by up to 15–20%. Binding kinetic studies paradoxically suggested that glycosylated HDL3 binds with higher affinity to a reduced number of binding sites. The findings in this study suggest that nonenzymatically glycosylated HDL may be functionally abnormal and might contribute to the development of atherosclerosis in patients with diabetes mellitus.
Increased Sensitivity to Insulin-Releasing and Glucoregulatory Effects of Dynorphin A 1–13 and U 50488h in ob/ob Versus Lean Mice
Authors: Khawaja Xavier Z , Green Irene C , Thorpe Julian R , Bailey Clifford J
Published: September 2013

The effects of two K-opiate agonists, U 50488h and dynorphin A1–13, on plasma insulin and glucose concentrations in vivo and insulin release in vitro were tested in fasted genetically obese (ob/ob) and lean (+ / +) mice at 12–15 wk of age. Fasting plasma insulin concentrations in ob/ob and lean mice were 1.22 ± 0.10 and 0.23 ± 0.05 nM, and plasma glucose levels were 6.90 ± 0.84 and 4.70 ± 0.29 mM, respectively. Administration of U 50488h (1 mg/kg body wt i.p.) to ob/ob mice dramatically raised plasma insulin by 670 and 790 pM at 15 and 30 min. Plasma glucose was raised from 5 min onward to a maximum increment of 4.2 mM above baseline. These effects were blocked by simultaneous administration of naloxone (10 mg/kg). A higher dose of U 50488h (10 mg/kg body wt i.p.) was required to produce significant increases in lean mouse plasma insulin (81 pM at 15 min) and glucose (0.7,1.1, and 1.7 mM at 5,15, and 30 min, respectively). Dynorphin (1 mg/kg body wt i.p.) raised plasma insulin in ob/ob mice by 380 and 410 pM at 15 and 30 min and raised plasma glucose by 1.6 mM at 15 min. In lean mice, the same dose of dynorphin had no effect on plasma insulin concentrations but induced a small rise in glucose. In ob/ob mice, the agonist-induced rise in glucose did not cause the insulin response, because insulin levels were not elevated by a glucose challenge. Light- and electron-microscopy studies of sections of pancreas and islets from ob/ob and lean mice demonstrated the presence of three times more dynorphin within glucagon than somatostatin cells in ob/ob mice. These observations coincided with increased responsiveness (10- to 100-fold) of isolated islets of ob/ob mice to the insulinotropic effect of dynorphin and U 50488h. We suggest that this insulin-resistant glucose-insensitive mouse model of obesity and non-insulin-dependent diabetes has increased sensitivity to K-agonists, probably acting peripherally and independently to mediate hyperglycemic and hyperinsulinemic responses.
Reduced Insulinotropic Effects of Glucagonlike Peptide I-(7–36)-Amide and Gastric Inhibitory Polypeptide in Isolated Perfused Diabetic Rat Pancreas
Authors: Suzuki Seiji , Kawai Koichi , Ohashi Shinichi , Mukai Hidehito , Murayama Yasuko , Yamashita Kamejiro
Published: September 2013

The pathophysiological role of incretin in diabetes mellitus has not been established. We therefore examined the effects of glucagonlike peptide l-(7–36)-amide (truncated GLP-I) and gastric inhibitory polypeptide (GIP) on insulin and glucagon release from isolated perfused pancreases of diabetic rats (12–14 wk of age, mean ± SE fasting plasma glucose 8.9 ± 0.6 mM, n = 25) after an injection of 90 mg/kg streptozocin on the 2nd day after birth and compared the results with those of nondiabetic control rats. In diabetic rats, the infusion of 1 nM GLP-I or GIP in perfusates with varying glucose concentrations (2.8, 5.6, 8.3,11.1, or 22.2 mM) caused a nearly equal degree of insulin stimulation from a similar basal insulin level. Meanwhile, basal and GLP-I- or GIPstimulated insulin release increased in correlation with the ambient glucose concentration in nondiabetic rats. The degree of stimulation of insulin release at glucose concentrations of 5.6 mM in diabetic rats was −33% that of nondiabetic rats. The stimulation potency was the same between GLP-I and GIP. The insulin treatment for diabetic rats (5 U/kg NPH insulin at 0900 and 2100 for 6 days) brought only a slight improvement in the glucose dependency of GLP-I-stimulated insulin release. The effects of GLP-I and GIP on glucagon release were completely opposite. GLP-I suppressed release; GIP stimulated it. In diabetic rats, the degree of suppression by GLP-I and stimulation by GIP were almost the same with similar basal glucagon levels in the perfusate with varying glucose concentrations. In nondiabetic rats, these parameters were inversely correlated with ambient glucose concentrations. Insulin treatment did not improve the glucose dependency of either the basal glucagon level or the degree of suppression by GLP-I. These data demonstrate that the incretin effect of GLP-I and GIP, glucose-dependent stimulation of insulin release, is reduced in neonatal streptozocin-induced diabetic rats and that conventional insulin treatment does not normalize this abnormality. Also, inhibition of glucagon release by GLP-I is glucose dependent in nondiabetic rats, and this glucose dependency is lost in diabetic rats.
Marked Improvement of Glucose Homeostasis in Diabetic ob/ob Mice Given Oral Vanadate
Authors: Brichard Sonia M , Bailey Clifford J , Henquin Jean-Claude
Published: September 2013

The trace element vanadium exerts insulinlike effects in vitro and decreases hyperglycemia in insulindeficient animals. This study examined whether vanadate can improve glucose homeostasis in genetically obese hyperglycemic insulin-resistant ob/ob mice, which present metabolic abnormalities similar to those of human non-insulin-dependent diabetes. Sodium orthovanadate (0.3 mg/ml) was administered for 7 wk in H2O. Vanadate treatment induced a fall in fed and fasted plasma glucose and insulin levels and improved tolerance to oral glucose; the stimulated glucose area was decreased by 65%, and an early peak of insulin secretion was restored. During an intravenous glucose tolerance test, the glucose disappearance rate was twofold higher in vanadate-treated mice, and the reappearance of a significant insulin response was also observed. Moreover, vanadate produced a twofold increase in hepatic glycogen content and prevented the exhaustion of pancreatic insulin stores. The hypoglycemic response to exogenous insulin was similar in control and treated mice. In vitro experiments showed that basal glucose oxidation by hemidiaphragms was 32% higher in vanadate-treated mice than in controls, although stimulation by insulin was similar in both groups. In conclusion, oral vanadate caused a marked and sustained improvement of glucose homeostasis in diabetic insulin-resistant mice by exerting an insulinlike effect on peripheral tissues and apparently preventing the exhaustion of pancreatic insulin stores.
Short-Term Administration of Captopril and Nifedipine and Exercise-Induced Albuminuria in Normotensive Diabetic Patients With Early-Stage Nephropathy
Authors: Romanelli Giuseppe , Giustina Andrea , Bossoni Simonetta , Caldonazzo Andrea , Cimino Antonino , Cravarezza Primarosa , Giustina Gianni
Published: September 2013

Recent studies have demonstrated that short-term angiotensin converting enzyme (ACE) inhibition with captopril can reduce urinary albumin excretion rate (UAER) after exercise in normotensive diabetic patients with early-stage nephropathy. The aim of this study was to investigate whether this effect of ACE inhibition was due to a systemic hypotensive action or a specific action at the intrarenal level. Thus, we compared the acute effects of captopril and the Ca2+-channel blocker nifedipine on exercise-induced UAER in normotensive (blood pressure <165/95 mmHg) diabetic patients who were normoalbuminuric or microalbuminuric at rest (stage 2 or 3 of diabetic nephropathy). Twenty-five stage 2 diabetic nephropathy patients, 39 stage 3 diabetic nephropathy patients, and 12 nondiabetic subjects performed five submaximal cycloergometric exercises (90% of theoretical heart rate) on nonconsecutive days. The first two exercises were performed in basal conditions; the next three exercises were performed 24 h after administration of captopril (25 mg twice daily) or nifedipine AR (20 mg twice daily) or placebo (1 tablet twice daily) according to a randomized double-blind crossover trial. After placebo, blood pressure and UAER did not change at rest or 1 h after exercise. After captopril, blood pressure at rest and during exercise was similar to that observed after placebo. UAER at rest was not modified, whereas 1 h after exercise, it was significantly decreased both in stage 2 and stage 3 diabetic nephropathy patients (P < 0.001). After nifedipine, blood pressure decreased significantly at rest and during exercise in respect to placebo and captopril. UAER at rest did not change significantly. UAER 1 h after exercise was significantly decreased both in stage 2 and stage 3 diabetic nephropathy patients (P < 0.01 vs. placebo), but it was significantly higher than that observed after captopril (P < 0.01). Captopril is more effective than nifedipine in reducing exercise-induced UAER in normotensive diabetic patients even though it does not influence blood pressure response to exercise. The effects of captopril on UAER induced by exercise may be due to a specific action at the intrarenal level.
Effect of Bacitracin on Retroendocytosis and Degradation of Insulin in Cultured Kidney Epithelial Cell Line
Authors: Dahl David C , Tsao Tanny , Duckworth William C , Frank Bruce H , Rabkin Ralph
Published: September 2013

In an earlier study, we described the presence of a retroendocytotic pathway for insulin in a cultured kidney epithelial cell line. Derived from the opossum kidney (OK), these cells possess many features of proximal tubule epithelium, which is the major site of kidney insulin metabolism. We studied the interaction between the retroendocytotic and the degradative pathways with bacitracin as a pharmacological probe. Monolayers of OK cells were loaded with 125I-labeled insulin over 30 min, acid washed to remove membrane-bound insulin, then incubated in fresh medium for 60 min while the release of intracellular radioactivity was monitored. In experiments carried out in the presence of bacitracin (2 mM), there was a twothirds increase in intracellular radioactivity at the end of the loading phase. Measurements made during the subsequent release phase showed that bacitracin reduced the release of degradation products. Thus, although controls released 72.1 ± 8.1% of the internalized radioactivity as trichloroacetic acid (TCA)-soluble products, bacitracin-treated cells released 59.2 ± 9.4% (P < 0.02). In contrast, release of TCAprecipitable insulin increased from 15.2 ± 4.6% in controls to 25.8 ± 3.7% in bacitracin-treated cells (P < 0.01). In separate experiments analyzed by gelexclusion chromatography, 6.4 ± 0.6% of radioactivity released from preloaded control cells into medium over 60 min was insulin sized compared to 29.7 ± 1.4% in bacitracin-treated cells. High-performance liquid chromotography revealed that 61.5 ± 3.5% of this insulin-sized material released from control cells preloaded with A14-insulin eluted as intact insulin and the remainder as unidentified intermediate degradation products. In the bacitracin group, 48.0 ± 2.1% of the radioactivity eluted with insulin, and 18.5 ± 0.6% eluted under a series of peaks representing intermediate products. Included in this material was a doublet, which has been identified in previous experiments with OK cells, other cell types, and liver endosomes. Previous characterization of this doublet indicates that it results from cleavage of insulin by insulin protease or a similar enzyme. We conclude that, by inhibiting the initial steps in the degradation of insulin, bacitracin diverts intact insulin and early degradation products from the degradative to the retroendocytotic pathway. These findings suggest that insulin degradation commences in an extralysosomal site, probably recycling endosomes.
Determination of Portal Insulin Absorption From Peritoneum via Novel Nonisotopic Method
Authors: Selam Jean-Louis , Bergman Richard N , Raccah Denis , Jean-Didier Nathalie , Lozano James , Charles M Arthur
Published: September 2013

The absorption mechanism of insulin administered in the peritoneal cavity (IP) is of current interest because of the near availability of implantable insulin-infusion devices for the treatment of diabetes. To determine the fraction of insulin absorbed by the portal circulation after IP administration, a novel nonisotopic method is described. Conscious fasting diabetic dogs were studied at normoglycemia via the euglycemic insulinclamp method. Posthepatic appearance of insulin and C-peptide were measured in peripheral blood during IP or intravenous (IV) equimolar infusion of insulin and Cpeptide at two sequential 3-h infusion rates of 3.2 and 12.8 pmol · kg−1 · min−1. Prior studies have shown that 40–60% of portal insulin is extracted at first pass by the liver, whereas C-peptide is not extracted. Thus, the fraction (F) of IP insulin not taken up by liver at first pass and consequently the fraction absorbed by the portal circulation can be derived from insulin (I) and C-peptide (C) plasma concentration values at steady state with a monocompartmental model where F = (IIP/IIV)(CIV/CIP). The mean ± SE value of F was 49.7 ± 8.8%. Glucose disappearance rates were lower with IP than IV infusion but similar when peripheral insulin levels were matched. We conclude that IP insulin is almost entirely absorbed by the portal circulation and induces lower glucose disappearance rates than IV insulin because of lower peripheral circulating insulin levels. Whether these properties make the IP route a more appropriate route for insulin therapy than the subcutaneous or IV routes remains to be established.
Characteristics of Learning and Memory in Streptozocin-Induced Diabetic Mice
Authors: Flood James F , Mooradian Arshag D , Morley John E
Published: September 2013

We demonstrated that mice with streptozocin-induced diabetes mellitus have normal acquisition for relatively simple tasks but show problems in learning more complex tasks such as shuttle box avoidance. Enhanced learning previously reported in simple passive avoidance tasks appears to be due to increased foot shock sensitivity. Diabetic mice show a marked memory retention deficit after learning an active avoidance T-maze task. This retention deficit was reversed by a single injection of insulin, suggesting that it may be related to hyperglycemia per se. Diabetic mice have a shift to the left in the inverted U-shaped dose-response curve for memory retention produced by the acetylcholine agonist arecoline. Based on a preliminary screening, responses to several other pharmacological memory enhancers are probably altered in diabetic mice. These studies suggest that this mouse model of diabetes mellitus demonstrates a deficit in memory retention and retrieval similar to that seen in humans with diabetes mellitus.
Autoxidative Glycosylation and Possible Involvement of Peroxides and Free Radicals in LDL Modification by Glucose
Authors: Hunt James V , Smith Christopher C T , Wolff Simon P
Published: September 2013

It has been postulated that the etiology of the complications of diabetes involves oxidative stress, perhaps as a result of hyperglycemia. Consistent with this hypothesis, it has been shown that glucose, under physiological conditions, produces oxidants that possess reactivity similar to the hydroxyl free radical. These oxidants hydroxylate benzoic acid, fragment protein, and induce peroxidation in phosphatidylcholine liposomes and low-density lipoprotein (LDL) when LDL is incubated with hyperglycemic levels of glucose in vitro. These reactions are accelerated by transition metals and inhibited by a metal-chelating agent. The atherosclerotic potential of LDL in diabetes mellitus is often discussed in terms of protein glycosylation, which may affect cellular interactions. Our studies demonstrate, however, that peroxidative reactions also accompany LDL glycosylation in vitro. Peroxidative modification of LDL has also been implicated in LDL atherogenicity. Our studies indicate that glycosylation and peroxidation occur concomitantly in LDL modified by glucose in vitro and may both contribute to the behavioral changes of this lipoprotein.
Structural Abnormality in LDL From Diabetic Patients as Revealed by Resonance Raman Spectroscopy
Authors: Verma Surendra P , Philippot Jean R , Bonnet Bernard , Rostain-Capaillon Odile , Sainte-Marie Josette , Colette Claude , Monier Louis
Published: September 2013

Resonance Raman spectra of low-density lipoprotein (LDL) isolated from the plasma of diabetic patients (age range 13–77 yr, mean age 37 yr) and age- and sex-matched control subjects were recorded in the 1000- to 1600-cm−1 region as a function of temperature (0–50°C). Both nondiabetic and diabetic LDL yield spectra characterized by two major bands near 1160 and 1530 cm−1 due to the carotenoid component of lipoproteins. The relative intensity of 1530- and 1160-cm−1 bands, assigned to —C=C— and =C—C= stretchings, respectively, i.e., I1530-I1160 ratio, was plotted against temperature. For nondiabetic control subjects, the plots showed an inflection in the temperature range of 30–39°C, which corresponded to the thermal transition of LDL. This transition was abolished in the LDL of diabetic patients (P < 0.001), suggesting an altered lipid structure. The transition (30–39°C) was also abolished in the in vitro glycosylated nondiabetic LDL. Lipid analysis did not show any appreciable change between nondiabetic control subjects and diabetic patients. The change in the thermal transition properties of diabetic LDL has been attributed to the organizational change in the LDL protein.
Multigenic Basis for Type I Diabetes: Association of HRAS1 Polymorphism With HLA-DR3,DQw2/DR4,DQw8
Authors: Owerbach David , Gunn Sheila , Gabbay Kenneth H
Published: September 2013

We analyzed extended haplotypes composed of DNA loci on the short arm of chromosome 11 for segregation with insulin-dependent (type I) diabetes mellitus. The markers for these loci are tyrosine hydroxylase, insulin, and c-Ha-ras-1 proto-oncogene (HRAS1). We report, in a study of 27 families, that a specific haplotype (H), containing a 3-kilobase (kb) HRAS1-Tag I DNA polymorphism, segregated differentially in diabetic and nondiabetic siblings (P = 0.005). A parallel population study showed that the 3-kb HRAS1-Taq I polymorphism is increased in frequency in type I patients having two strong HLAsusceptibility haplotypes compared with other type I patients or healthy control blood donors (P < 0.010 and P < 0.025, respectively). The polymorphic variable, enhancer, and promoter regions flanking the human insulin gene on the H haplotype were not associated with type I diabetes. These results indicate that the HRAS1 locus or genes in linkage disequilibrium with this locus are involved in the pathogenesis of HLADR3/4 type I diabetes mellitus.