bluuebunny/crossref_metadata_2025 huggingface dataset:
Pulsatile Insulin Has Greater Hypoglycemic Effect Than Continuous Delivery
Authors: Matthews D R , Naylor B A , Jones R G , Ward G M , Turner R C
Published: September 2013

The relative hypoglycemic effects of pulsatile versus steadily infused insulin have been examined in six normal subjects in whom pancreatic insulin output was suppressed by somatostatin-14. Soluble insulin was infused continuously overnight on one occasion and on another occasion the same quantity was given in pulses of 2-min duration with a gap of 11 min. The mean plasma glucose concentrations were lower when pulsed insulin was given [mean for the last hour: 4.66 ± 0.08 mmol/L (±SEM) versus 5.53 ± 0.06 mmol/L (±SEM) for steady infusion], diverging significantly (P < 0.05 paired t test) 7 h after the start of the study. The specific binding of 125I(A14)mono-iodo-insulin to monocytes was greater after pulsed insulin (2.9% with pulsed versus 2.4% with steadily infused insulin at tracer-only point; P < 0.02 paired t test). Thus, intravenous insulin has greater hypoglycemic effect when pulsed, possibly mediated by greater insulin receptor binding.
Measurement of Insulin Binding by Erythrocyte Ghosts: A Method That Allows Storage Without Loss of Binding Characteristics
Authors: Williams Paul F , Watson Sue K , Yue Dennis K , Turtle John R
Published: September 2013

The insulin binding of red cell ghosts was studied and found to be similar to insulin binding of the original erythrocyte. Red cell ghosts were stable on storage, and serial samples can be measured in one assay, thus eliminating the problem of interassay variation. Erythrocyte ghosts may be preferable to whole red cells for long-term insulin receptor studies.
Retrograde Axonal Transport of Intravenously Administered 125I-Nerve Growth Factor in Rats with Streptozotocin-induced Diabetes
Authors: Schmidt Robert E , Modert Christopher W , Yip Henry K , Johnson Eugene M
Published: September 2013

We have previously demonstrated a reproducible axonopathy, involving but not confined to sympathetic neurons, which involves selectively the mesenteric nerves of chronically diabetic rats. We describe a series of experiments aimed at correlating changes in the retrograde transport of 125I-nerve growth factor (125I-NGF) with development of the lesions.125I-NGF was injected systemically and the time course of accumulation compared in superior cervical ganglion (SCG) and superior mesenteric ganglion (SMG). 125I-NGF accumulated in the SCG with a sharp peak at 12 h, whereas in the SMG, accumulation reached a plateau, remaining at relatively constant amounts for 8 h before decreasing. There was a marked decrease in the accumulation of 125I-NGF (averaging 44% in 11 separate experiments) in the SMG at early times (approximately 12 h). These decreases were seen in animals diabetic for times ranging from 2 days to 10 mo. In contrast, no consistent decrease was observed in the SCG. The time course of 125I-NGF accumulation was unaltered in the SCG of diabetic rats but was clearly retarded in the SMG of diabetic animals. Morphometric examination of the SMG of diabetic animals and controls showed little or no atrophy or neuron loss due to diabetes. We conclude that decreases in the retrograde transport of NGF occur selectively in those ganglia (SMG) serving the alimentary tract in which mesenteric nerves develop distal axonopathy, but not in ganglia whose nerves do not ultimately develop such lesions (SCG). The decreases in retrograde transport precede the development of morphologic lesions in this system. These results are consistent with the suggestion that impairment of axonal transport may play a role in the development of axonopathy.
Receptor Binding Properties of Monoiodotyrosyl Insulin Isomers Purified by High Performance Liquid Chromatography
Authors: Frank Bruce H , Peavy Daniel E , Hooker Carol S , Duckworth William C
Published: September 2013

The four monoiodinated forms of pork insulin were prepared by lactoperoxidase catalyzed iodination followed by separation on reverse-phase high-performance liquid chromatography. The procedure was rapid and extremely reproducible, resulting in total separation of the B16, B26, and A14 monoiodinated insulins from one another and from unreacted insulin. The A19 derivative eluted closely to the unreacted insulin, requiring an additional step on ion exchange chromatography for complete purification of this derivative. The receptor binding properties of the derivatives were examined in isolated rat adipocytes and hepatocytes and in cultured human IM-9 lymphocytes. The relative binding affinities of the derivatives varied in the different cell types. At tracer concentrations, the B26 derivative had the highest apparent affinity in IM-9 lymphocytes and in adipocytes, but the A14 had the greatest affinity in hepatocytes. At equivalent concentration of each tracer, IM-9 lymphocytes bound the B26 isomer 144 ± 9% (P < 0.05) and the B16 isomer 118 ± 3% (P < 0.05) as well as the 125I (A14) insulin. Isolated rat adipocytes also bound the B26 to a greater extent at tracer concentrations (117 ± 6%; P < 0.01) compared with A14, but bound B16 less than A14 (91 ± 2%; P < 0.01). Isolated rat hepatocytes bound more of the A14 derivative than either of the B-chain labels. The B26 bound 72 ± 2% (P < 0.01) and the B16 90 ± 1% (P < 0.01) as well as the A14 derivative to isolated hepatocytes.

In contrast to the results obtained at tracer concentrations of the isomers, higher concentrations of native insulin were required for 50% competitive displacement (ED50) of the B26 derivative as compared with the other two derivatives in all three cell types examined.

These results indicate a higher average affinity for the B26 isomer in the three tissues, and suggest that results obtained from tracer binding alone may not provide an accurate reflection of receptor affinity for the monoiodoinsulin isomers.

Competitive displacement of the 125I-labeled derivatives by 127I (nonradioactive) labeled homologues in the IM-9 lymphocytes also showed the B26 derivative to have the highest affinity.

The present study demonstrates that monoiodoinsulin selectively labeled on each of the tyrosines can be prepared and separated on HPLC and used to examine insulin receptor binding in different cell types. Furthermore, the data demonstrate different apparent affinities of the selectively-labeled monoiodoinsulin isomers in different cell types, suggesting variability in insulin binding properties in different tissues.
Experimental Diabetes Mellitus and Age-simulated Changes in Intact Rat Dermal Collagen
Authors: Yosha S R , Elden H R , Rabinovitch A , Mintz D H , Boucek R J
Published: September 2013

Alterations in the physical properties of dermal collagen by streptozotocin-induced diabetes mellitus were investigated in young adult Lewis rats by mounting standardized rings of tail skin on an Instron Universal Testing Apparatus and measuring the thermally induced isometric contraction and stress at rupture of the tissue. Diabetes significantly increased the maximum tension (Tmax) of the contraction and raised the teature for the Tmax while the stress at rupture (TR) was unaffected when compared with values for controls fed ad libitum and controls fed restricted diets for weight loss equivalence. The diabetes-mediated changes in thermal contractility appeared to be independent of the collagen concentration or negative caloric balance and resembled the reported age-related change in rat skin collagen.
Insulin Binding and Responsiveness in Fat Cells from Patients with Reduced Glucose Tolerance and Type II Diabetes
Authors: Lönnroth Peter , Digirolamo Mario , Krotkiewski Marcin , Smith Ulf
Published: September 2013

Adipose tissue was obtained from 66 individuals including 21 patients with type II diabetes of different severity (16 SU-treated and 5 diet-treated only) as well as 9 obese subjects with reduced glucose tolerance. Adi-pocyte insulin binding, antilipolytic effect of insulin, and glucose incorporation into triglycerides were measured in the diabetic and the obese subjects and the data compared with that of normal controls of similar age and relative weight.

Insulin binding per cell was normal in the diabetic patients and was significantly increased at low insulin concentrations in the obese patients with reduced glucose tolerance, suggesting increased affinity. Furthermore, insulin binding correlated negatively with age but, when age was corrected for, did not correlate significantly with fasting insulin or glucose levels, relative body weight, or fat cell size. Insulin sensitivity, measured as the antilipolytic effect of insulin, was similar in all patient groups. Patients with the most severe type II diabetes (SU-treated group) demonstrated, in contrast to the less severely diabetic patients, a marked reduction in both basal and insulin-stimulated glucose incorporation into triglycerides showing the presence of a pronounced postreceptor defect. The insulin effect on glucose incorporation correlated negatively with the fasting glucose levels, suggesting that the postreceptor defect seen in the adipocyte reflectsperturbations in other organs, like muscle or liver, of greater importance for glUCOSe homeostasis.
Virus-induced Diabetes in Autoimmune New Zealand Mice
Authors: Yoon Ji-Won , Melez Kathleen A , Smathers Patricia A , Archer Juanita A , Steinberg Alfred D
Published: September 2013

Infection of autoimmune New Zealand mice with the D variant of encephalomyocarditis (EMC) virus results in beta-cell damage and clinical diabetes. The induction of diabetes in parental NZB and NZW strains was independent of sex. However, the susceptibility to virus-induced diabetes in their F1 offspring was sex dependent. This susceptibility was significantly higher in male (NZB x NZW) F1 mice as compared with female F1 mice. Castration of male F1 mice significantly reduced the susceptibility to diabetes. These results suggest that parental NZB and NZW strains have recessive genes at different loci which do not allow sex hormones to influence the susceptibility to diabetes. It is concluded that both the genetic background of the host and sex hormones influence the development of virus-induced diabetes in autoimmune New Zealand mice.
Has the Recessive Hypothesis for Susceptibility to Insulin-dependent Diabetes Mellitus Been Firmly and Unequivocally Rejected?
Authors: Williams Robert C
Published: September 2013

An analysis of an implied assumption, that of Hardy-Wei n berg equilibrium, reveals that the single locus, recessive allele hypothesis for susceptibility to insulin-dependent diabetes mellitus has not been firmly and unequivocally rejected by the method of Rotter et al1.
Analysis of Insulin Gene Expression in Human Pancreas
Authors: Giddings Stephen J , Rotwein Peter , Chirgwin John M , Scharp David , Permutt M Alan
Published: September 2013

The purpose of these studies was to determine whether insulin gene expression at the level of proin-sulin mRNA could be studied in human pancreas. RNA was isolated from autopsy specimens and analyzed by RNA-blot hybridization with various 32P-human insulin gene probes spanning either the entire gene or the second intervening sequence. A major band 0.62 kilo-bases (kb) in length accounted for over 95% of the mRNA, consistent in size with presumed mature proin-sulin mRNA. In addition, minor bands of 1.5 and 1.3 kb were seen, consistent with an initial gene transcript containing both intervening sequences and with a processed intermediate. The 1.5-and 1.3-kb RNA were confirmed to be proinsulin mRNA precursors by hybridization specifically with the IVS II probe. Total RNA and polyadenylated RNA from five normal pancreata and two insulinomas revealed the same pattern. This method provides a means of determining whether altered insulin gene expression is one cause of diabetes.
In Vivo Modulation of Gap Junctions and Dye Coupling Between B-Cells of the Intact Pancreatic Islet
Authors: Meda P , Michaels R L , Halban P A , Orci L , Sheridan J D
Published: September 2013

The extent of gap junctions and dye coupling between insulin-producing B-cells was analyzed on islets of Langerhans isolated from adult rats treated for one day with glibenclamide, an insulin secretagogue, or diazoxide, a blocker of insulin release, or a combination of the two drugs. Glibenclamide treatment was associated with a marked depletion of the islet insulin content, an effect which was blocked by pretreatment of the rats with diazoxide. Diazoxide alone caused a marked increase in the plasma glucose level, and a decrease in the level of circulating insulin and in the hormone content of the B-cells. Quantitative analysis showed that (1) under control conditions, B-cells are connected by minute gap junctions (as evaluated on freeze-fracture replicas) and show a nonuniform and apparently restricted dye coupling (as determined by microinjection of the low-molecular-weight fluorescent probe Lucifer Yellow CH); (2) each of the three treatments tested significantly increased the relative and absolute gap junction area of the B-cells and the number of detectable, dye coupled B-cells per microinjec-tion. After treatment with glibenclamide alone or with diazoxide plus glibenclamide, a 1.5–1.8-fold increase in gap junction area and a 2.7–3.7-fold increase in the number of dye-coupled B-cells were observed. In contrast, following treatment with diazoxide alone, gap junctions and dye coupling were found increased 1.8 and 8.7 times, respectively, as compared with control values. The results suggest that small territories of coupled B-cells may exist within adult islets of Langerhans, and show that such a pattern can be modulated in vivo via changes in the gap junctions and dye coupling of B-cells. The mechanism(s) regulating these changes is unknown but is clearly operative after both stimulation and inhibition of insulin release.
The Effects of Aging and Diabetes Mellitus on Human Red and White Cell Calmodulin Levels (Chemotaxis/Phagocytosis/Calcium)
Authors: Morley John E , Levine Allen S , Beyer H Steven , Mooradian Arshag D , Kaiser Fran E , Brown David M
Published: September 2013

Both the calcium ion and calmodulin have been proposed to play a role in producing the intracellular effects of insulin. Abnormalities in calmodulin levels have previously been reported in tissues from diabetic animals. Thus, using a sensitive and specific radioimmunoassay, we measured calmodulin levels in red cells and polymorphonuclear leukocytes from humans with diabetes mellitus. Diabetic patients have significantly lower white cell calmodulin levels than agematched controls. There were no significant differences in red cell calmodulin levels in diabetics compared with controls. Red cell calmodulin levels in normal subjects were decreased with advancing age. We conclude that the alteration in calmodulin levels in leukocytes from diabetics may be associated with the decreased neutrophil function that has been observed in diabetics.
Human Proinsulin-Specific Antigenic Determinants Identified by Monoclonal Antibodies
Authors: Madsen Ole D , Frank Bruce H , Steiner Donald F
Published: September 2013

Antigenic determinants recognized by human proinsulin (HPI)-specific monoclonal antibodies (Mabs) and Mabs crossreacting with free human C-peptide (HCP) were mapped by using various forms of purified, partially converted HPI intermediates. Two HPI-specific mouse Mabs (GS-4G9 and GS-9A8) reacted with the same antigenic determinant, GS, which was localized to the site of linkage of the B-chain to the C-peptide (Arg-Arg) at positions 31–32. These antibodies bind with equal efficiency to C65-A1 split proinsulin and to intact HPI. The binding of C32-C33 split proinsulin is markedly reduced.

A rat Mab (GN-VIIB6), which crossreacts with free HCP in addition to HPI, reacted similarly with various HPI intermediates as it had with the corresponding synthetic HCP fragments, as previously reported (see ref. 9). This determinant (GN) is a three-dimensional structure composed of residues located in two separate regions in the C-peptide segment (positions 40–45 and 57–63). Reduced, carboxymethylated HPI retains the GN-determinant, whereas all insulin-like immunoreactivity identified with a conventional guinea pig insulin antiserum is completely lost. The binding of the two GS Mabs to the denatured HPI was reduced by 40–50% compared with intact HPI.

It is concluded that the strong GN-determinant can readily form in the C-peptide segment of HPI, independently of the presence of ordered structure in the insulin moiety. A predicted β-turn at position 47–50 may play an important role in bringing N- and C-terminal regions of the C-peptide segment into close proximity. The spontaneous formation of the GN-determinant could thus be a crucial step in the formation of the correct disulfide bonds in more distant parts of the newly synthesized HPI-polypeptide chain
Evidence for a Direct Action of Insulin to Increase Renal Reabsorption of Calcium and for an Irreversible Defect in Renal Ability to Conserve Calcium Due to Prolonged Absence of Insulin
Authors: Hoskins Beth , Scott Jennifer M
Published: September 2013

Male rats were injected with streptozotocin (STZ), 60 mg/kg, i.p., on 2 successive days. Six hours after the last STZ injection, some STZ-diabetic rats began receiving daily injections of insulin that were insufficient to control blood glucose. Another group of STZ-diabetic rats received insulin injections after 2 wk duration of untreated diabetes. Still other STZ-diabetic rats received no insulin treatment. Under sodium pentobarbital anesthesia, kidneys from each treatment group were isolated and perfused with an artificial“plasma”containing 45Ca. As urine was collected, urine-to-per-fusate ultrafiltrate (U/P) ratios for 45Ca were determined. The results of the studies showed that: STZ diabetes reduced 45Ca reabsorption by the kidney; the increased urinary excretion of calcium was not due to an osmotic effect or to a direct nephrotoxic action of STZ; and insulin therapy instituted early, but insufficient to control blood glucose, reduced the diabetes-induced calcium loss via a direct action on the kidney, whereas insulin therapy instituted late failed to reverse renal loss Of calcium.
β-Hydroxybutyrate Increases the Insulin Sensitivity of Adipocyte Glucose Transport at a Postreceptor Level
Authors: Green Allan , Bustillos Dalia P , Misbin Robert I
Published: September 2013

The effect of insulin on glucose transport (2-deoxyglucose uptake) in adipocytes was measured in the absence and in the presence of 10 mM sodium-DL-β-hydroxybutyrate. The ketone body had little or no effect on the basal or the maximally insulin-stimulated rate of transport. However, β-hydroxybutyrate potentiated the effect of submaximal concentrations of insulin, i.e., it resulted in a leftward shift in the dose-response curve. The half-maximally effective concentration of insulin was decreased by approximately 30% from 0.58 ng/ml to 0.40 ng/ml. β-Hydroxybutyrate caused a slight (approximately 10%) increase in 125I-insulin binding to the cells. To determine whether this small increase in insulin binding is responsible for the increased insulin sensitivity in the presence of the ketone, two mimickers of insulin action were used: a serum containing anti-insulin receptor antibodies and hydrogen peroxide. β-Hydroxybutyrate increased the sensitivity of glucose transport to stimulation by the anti-receptor antibody, demonstrating that insulin itself does not have to be present. β-Hydroxybutyrate also potentiated the effect of hydrogen peroxide (which acts at a level distal to the insulin receptor) even in cells that had been depleted of insulin receptors by trypsin treatment. Therefore, β-hydroxybutyrate acts, at least partly, at a post-insulin receptor level.

Adenosine increases the insulin sensitivity of adipocytes. β-Hydroxybutyrate increased the sensitivity of the cells: (1) in the presence of a maximally effective concentration of adenosine (50 μM), (2) in the presence of adenosine deaminase (10 μg/ml), and (3) in the presence of adenosine deaminase plus a maximally effective concentration of the adenosine analogue, N6-phenylisopropyl adenosine (1 μ). Therefore, β-hydroxybutyrate appears to act independently from, and by a different mechanism to adenosine.
Genetic Control of Pathogenesis of Diabetes in C3H Mice: Influence of the Major Histocompatibility Complex
Authors: Leiter Edward H
Published: September 2013

Inbred strains of genetically diabetic (db/db) male mice with H-2b haplotype were heretofore found resistant to the diabetogenic action of the db mutation, whereas C3HeB/FeJ-db/db males with H-2k haplotype were susceptible. To determine whether the major histocompatibility complex was linked to diabetes predisposition, we mated C3H.SW/SnJ females (H-2b haplotype) with C3HeB/FeJ-+/db males, identified the +/db heterozygotes in the F1 generation (all H-2b/H-2k), and intercrossed these to produce F2 db/db male offspring that were classed and studied according to the three segregating H-2 genotypes. We found an accelerated diabetes pathogenesis in terms of early onset of severe hyperglycemia, destruction of pancreatic beta cells, and mortality that was not linked to H-2. Of 9 F2 male diabetics with H-2b/H-2b genotype, 14 with H-2b/ H-2k genotype, and 5 with H-2k/H-2k genotype, all showed a more severe syndrome than did the grandparental-type C3HeB/FeJ-db/db males. We conclude that on the C3H inbred background, the major histocompatibility complex is not a major background modifier of the diabetes syndrome. The complexity of the results suggests residual non-H-2-related genetic variance between the two grandparental C3H stocks, with C3H.SW/SnJ females possessing diabetes susceptibility factors apparently lacking in C3HeB/FeJ (a stock bred to be free of the milk-borne mouse mammary tumor virus). Since C3H.SW/SnJ females transmitted to F2 males unknown diabetogenic factor(s) that did not appear to segregate, inheritance of a virus was suggested. The lack of association between H-2 and severity of the diabetes syndrome in these obese and insulin-resistant db/db mice makes the mouse model comparable to the human type II (non-insulin-dependent) diabetes syndrome that also appears not to be associated with HLA haplotype.
Hypoglycemic Brain Injury in the Rat: Correlation of Density of Brain Damage with the EEG Isoelectric Time: A Quantitative Study
Authors: Auer Roland N , Olsson Yngve , Siesjö Bo K
Published: September 2013

Thirty-eight male Wistar rats were exposed to insulin-induced hypoglycemia resulting in periods of cerebral isoelectricity ranging from 10 to 60 min. Plasma glucose levels during cerebral isoelectricity ranged from 0.12 mM to 1.36 mM. Control rats were injected with insulin, but hypoglycemia was terminated with glucose at the stage of large δ-wave EEG slowing. After recovery, the rats were allowed to wake up and survive for 1 wk.

The number of dying neurons was assessed with acid-fuchsin/cresyl-violet-stained, whole-brain, sub-serial sections using direct visual counting of acido-philic, cytoclastic neurons. Brains from control rats that were not allowed to become isoelectric showed no dying neurons. Ten minutes of cerebral isoelectricity produced very minimal brain damage. The density of neuronal necrosis was positively related to the number of minutes of cerebral isoelectricity up to the maximum examined period of 60 min, but showed no correlation with the blood sugar levels.

The cerebral cortex, hippocampus, caudate nucleus, spinal cord, and, to a lesser extent, cerebellar Purkinje cells were affected. The distribution of neuronal necrosis was not identical with that seen in ischemia, but, rather, suggested a CSF-borne neurotoxin operant in contributing to the pathogenesis of neuronal necrosis in hypoglycemie brain damage.

Neuronal death does not occur in hypoglycemia unless the EEG becomes isoelectric, whatever the blood sugar level. Serious brain damage does not occur until electrocerebral silence has been established for at least several minutes. Neuronal death accelerates after 30 min of EEG isoelectricity in the rat. Most animals recovered well after up to 40 min of cerebral isoelectricity.
Glucose Counterregulation in Patients After Pancreatectomy: Comparison With Other Clinical Forms of Diabetes
Authors: Polonsky Kenneth S , Herold Kevan C , Gilden Janice L , Bergenstal Richard M , Fang Victor S , Moossa A R , Jaspan Jonathan B
Published: September 2013

Glucose and counterregulatory hormone responses to a high-dose (1.7 mU/kg/min) insulin infusion were studied in 6 patients who had undergone total pancreatectomy, and the results were compared with those of normal controls and patients with other clinical forms of diabetes. The maximum increase in the plasma glucagon concentration during hypoglycemia in the pancreatectomized patients (5 ± 5.6 pg/ml) was less than in normals (121 ± 22 pg/ml). Type I diabetic subjects (28 ± 14 pg/ml), and insulin-treated diabetic subjects of recent onset (36 ± 12 pg/ml) also had reduced responses, while responses were normal in type II diabetic subjects (102 ± 26 pg/ml). The epinephrine response to the hypoglycemie stimulus was reduced after pancreatectomy (278 ± 81 pg/ml) and in type I diabetic subjects (628 ± 244 pg/ml), but was not different from control (858 ±126 pg/ml) in type II and recent-onset diabetic patients. There was considerable overlap in counterregulatory hormone responses in individual patients with and without autonomie neuropathy and with normal or undetectable fasting C-peptide concentrations. While the control subjects all experienced symptoms of hypoglycemia within a narrow range of plasma glucose concentrations (35–46 mg/dl), five of the diabetic subjects experienced symptoms of hypoglycemia at plasma glucose levels of > 55 mg/dl, and five had no subjective awareness of hypoglycemia despite plasma glucose levels < 30 mg/dl. Thus, (1) after pancreatectomy both glucagon and epinephrine responses are reduced, resulting in markedly impaired glucose recovery from insulin-induced hypoglycemia; (2) glucagon responses may be abnormal in insulin-treated diabetic subjects within the first year of diagnosis, despite relatively normal fasting C-peptide concentrations and no clinical evidence of autonomie neuropathy; and (3) glucagon and epinephrine responses are normal in type II diabetic subjects even if the disease is of prolonged duration and clinical evidence of autonomie neuropathy is present.
Effects of Chronic Beta Receptor Stimulation on Glucose Metabolism
Authors: Scheidegger Karl , Robbins David C , Danforth Elliot
Published: September 2013

The acute administration of a beta receptor-stimulating agent profoundly affects insulin-mediated glucose metabolism; however, little is known about the impact of chronic beta receptor stimulation on glucose metabolism and insulin sensitivity. We therefore investigated the effect of the chronic administration of a beta-2-agonist, terbutaline sulfate (TS), on glucose metabolism in 7 healthy, normal-weight, male volunteers between the ages of 21 and 30 yr. Studies were performed using the euglycemic, hyperinsulinemic (1.0 mU/min · kg) clamp technique before and after the oral administration of 5 mg of TS three times a day for 1 and 2 wk. Basal endogenous glucose production (EGP) (2.54 ± 0.11 versus 2.64 ± 0.14 mg/min · kg) and basal glucose oxidation (1.87 ± 0.16 versus 2.0 ± 0.2 mg/ min · kg) were unchanged by the chronic administration of TS. However, insulin-stimulated total glucose metabolism increased by 29% (7.0 ± 0.47 versus 9.05 ± 0.67 mg/min · kg; P < 0.02). Insulin-stimulated, nonoxidative glucose disposal increased by 45% (3.62 ± 0.42 versus 5.26 ± 0.48 mg/min · kg; P < 0.01), while insulin-stimulated glucose oxidation did not change significantly (3.38 ± 0.15 versus 3.79 ± 0.22 mg/min · kg). EGP was completely suppressed under both conditions. Mean basal plasma insulin concentration (41 ± 9 versus 49 ± 15 pmol/L) and insulin clearance during the clamp procedure was unchanged (477 ± 45 versus 474 ± 37 ml/min · m2). We conclude that chronic beta receptor stimulation with TS improves insulin-stimulated glucose disposal in man, mostly by improving nonoxidative glucose disposal, i.e., “glucose storage.” Mechanisms explaining these findings have not yet been elucidated. We suggest that an adaptive response of the sympathetic nervous system plays a role in this phenomenon.
Similar Metabolic Effects of Pulsatile Versus Continuous Human Insulin Delivery During Euglycemic, Hyperinsulinemic Glucose Clamp in Normal Man
Authors: Verdin E , Castillo M , Luyckx A S , Lefebvre P J
Published: September 2013

Seven normal volunteers were studied on two different occasions during which 4-h pulsatile (PULS: 0.8 mU · kg−1 · min−1, 7.5 min of 15) and continuous (CONT: 0.4 mU · kg−1 · min1) intravenous (i.v.) infusions of human insulin (Actrapid HM, Novo) were randomly compared. A euglycemic glucose clamp was performed and a 3-3H-glucose infusion was used for determination of endogenous glucose production (EGP) and metabolic clearance rate (MCR) of glucose. Plasma glucose was similar in both conditions; plasma insulin was stable at about 29 mU/L (CONT) and fluctuated between 10 and 45 mU/L (mean: 28, PULS). Exogenous glucose infused was 1.137 ± 0.058 and 1.088 ± 0.099 g · kg−1 · 4h−1 in CONT and PULS, respectively (NS). EGP was totally suppressed in both conditions. Glucose MCR increased similarly to a maximum of 6.71 ± 0.19 (CONT) and 6.79 ± 0.59 (PULS) ml · kg−1 · min−1 during the fourth hour. C-peptide plasma levels remained stable, whereas plasma glucagon, free fatty acids, and 3-hydroxybutyrate were similarly suppressed in both tests. Thus, under these conditions, pulsatile and continuous insulin infusions have similar metabolic effects. These data contrast with those of Matthews et al. (1983) who reported that, at lower plasma concentrations (5–19 mU/L), pulsatile insulin had greater hypoglycemic effect than did continuous delivery. It is concluded that pulsatile insulin shows no greater activity under normoglycemic, moderately hyperinsulinemic conditions in man.
In Vitro Characterization of Biosynthetic Human Proinsulin
Authors: Podlecki D A , Frank B H , Olefsky J M
Published: September 2013

The binding and biologic properties of human proinsulin produced by recombinant DNA technology have been determined. The biosynthetic human proinsulin was iodinated using lactoperoxidase and subsequently purified by HPLC to yield the [(125I)TyrA14]-proinsulin isomer.

Using isolated rat adipocytes, biosynthetic human proinsulin was shown to have approximately 11% of the binding potency of native insulin. At 16°C and 37°C, the ED50 values of biosynthetic human proinsulin were 3.7 nM and 15 nM, respectively, which was significantly different from the insulin values of 0.4 nM and 1.7 nM, respectively. Kinetic analysis suggested that the decreased affinity of biosynthetic human proinsulin was due primarily to a decreased association rate rather than an increased dissociation rate. Similar to insulin, biosynthetic human proinsulin exhibited a decreased half-time of dissociation in the presence of insulin (16.7 nM) or proinsulin (111 nM); however, this negative cooperative effect was lost in the presence of high concentrations of proinsulin (11 μM).

Biologic potency, assessed by measuring glucose transport in rat adipocytes, showed that biosynthetic human proinsulin had 10% of the biologic activity of insulin, suggesting close coupling between binding to receptors and membrane generated cellular response.

By extracting cell surface bound proinsulin with acidic buffer, the amount of 125I-proinsulin that internalized following binding to surface receptors was measured. At equilibrium, 55% of the cell-associated radioactivity was internalized at 37°C. When chloroquine-treated (200 μM) cells were incubated with 125I-proinsulin at 37°C, a 1.5-fold increase in the amount of intra-cellular proinsulin was observed at 1 h. This suggests that like insulin, chloroquine treatment augments the intracellular accumulation of proinsulin by inhibiting intracellular processing of internalized proinsulin.

In conclusion, (1) biosynthetic human proinsulin exhibits ∼ 10% of the binding and biologic potency of native insulin in adipocytes, (2) despite its decreased affinity for surface receptors, proinsulin is readily internalized by adipocytes, where it is degraded by a chloroquine-sensitive pathway.
Concanavalin A and Alloxan Interactions on Glucose-induced Insulin Secretion and Biosynthesis from Islets of Langerhans
Authors: Virji Mohamed A , Steffes Michael W , Estensen Richard D
Published: September 2013

The pretreatment of isolated islets of Langerhans with concanavalin A (Con A) completely blocks alloxan from suppressing the insulin release response to glucose. The lectin itself inhibits insulin secretion. This effect is dose dependent and reversible. Con A, however, has no protective action against the inhibition of glucose-induced insulin biosynthesis in islets exposed to alloxan. The protective action of Con A on alloxan toxicity is likely to be at the beta-cell surface at a membrane recognition site for glucose as a stimulus for secretion. The insulin biosynthetic effect of glucose appears to be mediated through a separate mechanism.
Direct Measurement of Polyol Pathway Activity in the Ocular Lens
Authors: González R Gilberto , Barnett Patrick , Aguayo James , Cheng Hong-Ming , Chylack L T
Published: September 2013

A method to measure the polyol pathway metabolic flux in the intact rabbit lens by 13C nuclear magnetic resonance spectroscopy is described. In the lens exposed to 35.5 mM glucose, the polyol pathway accounts for ⅓ of the total glucose turnover. The high metabolic activity of the pathway suggests a significant alteration in the reduced to oxidized pyridine nucleotide ratio in the lens exposed to high glucose.
β-Endorphin-like Immunoreactivity in Extracts of the Fetal Bovine Pancreas: Column Chromatographic Characterizations of a High-Molecular-Weight Immunoreactive Species
Authors: Tung A K , Cockburn E
Published: September 2013

Acid-ethanol extracts of fetal bovine pancreas were examined for the presence of β-endorphin-like immunoreactivity. Gel-filtration analyses revealed the presence of a major large-molecular-weight β-endorphin immunoreactive species of ∼20K∆. This molecular form maintained its size upon resubmission to gel filtration in the presence of 6 M guanidine hydrochloride, separated from the bulk of the glucagon immunoreactivity upon ion-exchange chromatography, showed proportional dilution in the β-endorphin radioimmunoassay, and interacted in a biospecific manner with Concana-valin-A-Sepharose.
Effects of Epicatechin on Rat Islets of Langerhans
Authors: Hii Charles S T , Howell Simon L
Published: September 2013

Plants containing epicatechin (a flavonoid) have been used to treat diabetes mellitus in Indian medicine. The present study reports effects of this compound on isolated islets of Langerhans. The flavonoid (1 mM) was found to increase insulin secretion from isolated rat islets of Langerhans in the presence of either 2 or 20 mM glucose, in static incubations, or in perifusion. The increase in insulin secretion mediated by epicatechin was both ATP- and temperature-dependent. Ultrastructural studies showed no deleterious changes in the structure of the B-cells after 5 days of exposure to the compound. Intraperitoneal injection of 30 mg/kg body wt of epicatechin twice daily for 4 days increased the islet insulin content by 30%. Secretion of insulin from islets isolated from epicatechin-injected rats was significantly increased when exposed to 20 mM glucose in comparison with water-injected controls. Furthermore, islets of adult rats cultured with 5.5 mM glucose for 4 days showed a significant increase in DNA synthesis in the presence of 0.05 mM epicatechin. These results suggest direct effects of epicatechin on various aspects of islet function.
Culture and Phenotype of Activated T-Cells from Patients with Type I Diabetes Mellitus
Authors: Hayward Anthony R , Herberger Mark
Published: September 2013

Blood T-cells from 28 patients with type I (insulin-dependent) diabetes (IDDM) of variable duration were examined for the Tac antigen by immunofluorescence, and for proliferation in the presence of interleukin 2 (IL 2). The mean percentage of Tac+ cells in patients whose IDDM was of less than 2-yr duration was 6.2% compared with 2% in patients whose IDDM was of 3 or more years' duration, or in healthy controls. The percentage of Tac+ cells in the patients' blood correlated positively with the amount of thymidine uptake in a 24-h culture of blood mononuclear cells and with the percentage of T-cell blasts generated in a 6-day culture. The patients' T-cell blasts stained with OKT 4 or OKT 8, suggesting that each of these subsets is present in the activated T-cell population in the patients' blood. The T-cell blasts did not show specificity for pork insulin in an antigen restimulation assay. There was no correlation between increased Tac+ cells and the presence or absence of islet cell antibodies. If T-cell activation in IDDM occurs as a result of recognition of islet cell antigens, our results suggest that both HLA-DR-restricted (OKT 4+) and A-, B-, and C-restricted (OKT 8+) T-cell subsets contribute.
Insulin and Glucagon Regulate Cytosolic Phosphoenolpyruvate Carboxykinase (GTP) mRNA in Rat Liver
Authors: Beale Elmus , Andreone Teresa , Koch Stephen , Granner Mark , Granner Daryl
Published: September 2013

Insulin causes a 7–10-fold decrease of both the mRNA that codes for rat hepatic phosphoenolpyruvate carboxykinase (mRNAPEPCK) and of PEPCK synthesis, provided the animals are made diabetic and fed chow. mRNAPEPCK, measured either by in vitro translation or cDNA hybridization, decreases with a half-time of 30–60 min after insulin treatment. This coordinant decrease, which approximates the half-life of mRNAPEPCK measured in a variety of situations, suggests that insulin acts by decreasing mRNAPEPCK production, and that the hormone does not alter the activity of a fixed amount of this RNA, or enhance its degradation.

Glucagon results in a ninefold induction of mRNAPEPCK. Half-maximal induction occurs with doses between 20–75 μg/100 g body wt and occurs within 30–45 min. Maximal induction requires 150 μg/100 g body wt and occurs about 80 min after a single glucagon injection. N6,O2' -dibutyryl cAMP and a cAMP analogue that is not metabolized, 8-(4-chlorophenylthio)cAMP, induce mRNAPEPCK as effectively as glucagon and with similar kinetics. Since sodium butyrate, adenosine, and dibutyryl cGMP are ineffective inducers, cAMP appears to be the active agent in the hepatocyte.
Influence of Thyroid Hormone Level on Insulin Action in Human Adipose Tissue
Authors: Arner Peter , Bolinder Jan , Wennlund Anders , Östman Jan
Published: September 2013

The relationship between the levels of circulating thyroid hormones and the action of insulin on adipose tissue was investigated in 6 hypothyroid patients and 6 hyperthyroid patients, all untreated, and 8 healthy control subjects. All were matched for age, body weight, and fat cell size. Gluteal s.c. adipose tissue was used. The insulin receptor number in isolated adipocytes was increased by 70% in hypothyroldism and decreased by 40% in hyperthyroidism. The sensitivities of the effects of insulin on lipolysis and glucose oxidation were increased fourfold in hypothyroidism and decreased fivefold in hyperthyroidism. The maximum insulin-induced glucose oxidation (insulin responsiveness) was inhibited by 60% in hypothyroidism and enhanced by 180% in hyperthyroidism. The thyroid hormone concentration was significantly correlated with insulin receptor number (r = −0.72), insulin responsiveness (r = 0.71), and insulin sensitivity (r = −0.75). It is suggested that thyroid hormones regulate the effect of insulin on adipose tissue, which occurs at the receptor and postreceptor levels of insulin action.
Amelioration of Diabetes Mellitus in Partially Depancreatized Rats by Poly(ADP-ribose) Synthetase Inhibitors: Evidence of Islet B-Cell Regeneration
Authors: Yonemura Yutaka , Takashima Toru , Miwa Koichi , Miyazaki Itsuo , Yamamoto Hiroshi , Okamoto Hiroshi
Published: September 2013

Ninety percent depancreatized rats received daily intra-peritoneal injection of 0.5 g/kg nicotinamide and 0.05 g/kg 3-aminobenzamide, potent inhibitors of islet poly(ADP-ribose) synthetase. One to three months after the partial pancreatectomy, urinary and plasma glucose levels in nicotinamide- and 3-aminobenza-mide-treated rats were markedly lower than those in saline-treated control rats. Morphologic examination of the remaining pancreata revealed that islets in the poly(ADP-ribose) synthetase inhibitor-treated rats were markedly enlarged and consisted largely of B-cells. These results suggest that poly(ADP-ribose) synthetase inhibitors induce islet B-cell regeneration, thereby preventing or improving diabetes mellitus in partially depancreatized rats.
Glucose-induced Changes in Histochemically Determined Ca2+ in B-cell Granules, 45Ca Uptake, and Total Ca2+ of Rat Pancreatic Islets
Authors: Wolters G H J , Pasma A , Wiegman J B , Konijnendijk W
Published: September 2013

Rat pancreatic islets contain an ionized or readily ionizable calcium fraction that can be determined by the metallochromic indicator glyoxal-bis-(2-hydroxyanil) (GBHA). This calcium fraction is mainly localized in the secretory granules. The relationship between the effects of glucose on 45Ca uptake and on this ionized calcium fraction was investigated. In addition, the effects of glucose on total islet calcium content were also studied. Stimulation of isolated islets for 30 min with 15 mM glucose in the presence of 2.5 mM CaCl2 increased the 45Ca uptake but decreased the GBHA-Ca content, while the total calcium content was not affected. Deletion of CaCl2 caused, at 2.5 mM glucose, an abrupt decrease of GBHA-Ca, which did not occur at 15 mM glucose. Total islet calcium content decreased slowly at 2.5 mM glucose, but this was not significantly affected by glucose stimulation. Islet GBHA-Ca can be reduced by 70% and total islet calcium by 30% by means of washing with calcium-free buffer. Reintroduction of calcium at 2.5 mM glucose partly restored, but glucose 15 mM completely restored, the GBHA-Ca level within 5 min. The total calcium content was restored within 15 min independent of the glucose concentration. The increase of the islet calcium content equalled the 45Ca uptake at 2.5 mM glucose. The 45Ca uptake at 15 mM glucose was higher than the increase of the islet calcium content.

The results indicate that the intragranular Ca2+ pool, as measured by GBHA, is rapidly and dramatically altered by glucose stimulation. The glucose-induced changes of this Ca2+ fraction are not directly related to changes of the 45Ca uptake or the total islet calcium content. It is suggested that intracellular factors, particularly cAMP (production of which is Ca2+ dependent) and Na+ concentration, alter the amount of granular Ca2+.
Development of Early Lesions of Microangiopathy in Chronically Diabetic Monkeys
Authors: Yasuda H , Harano Y , Kosugi K , Nakano T , Suzuki M , Tsuruoka Y , Taniguchi Y , Nishimori T , Kikkawa R , Shigeta Y
Published: September 2013

A chronic diabetic state was produced in Macaca fuscatus, and these diabetic monkeys were kept without insulin treatment for up to 25 mo. The metabolic derangements were characterized by hyperglycemia, insulinopenia, hyperglucagonemia, ketonemia, and hyperlipidemia. Significant thickening of the capillary basement membrane of the gastrocnemius muscle was observed in the chronically diabetic monkeys, and became obvious in the course of diabetic state; 732 ± 35 Å in controls, 750± 58 Å in diabetic monkeys with duration of 4 mo, and 1165 ± 112 Å in those with duration of more than 11 mo. In addition to duration of the diabetic state, severity of hyperglycemia is also thought to play an important role in the capillary basement membrane thickening judging from the fact that diabetic monkeys with constant hyperglycemia showed a greater membrane thickening. Ultrastructural alterations, such as significant thickening of glomerular basement membrane and increase of mesangial matrix, were observed in kidney as well.

These results indicate that diabetic microangiopathy has been produced by metabolic derangements characterized by chronic hyperglycemia, insulinopenia, and hyperlipidemia.
Downregulation Occurs Normally in Cultured Epstein-Barr Virus-transformed Lymphocytes from Patients with Extreme Insulin Resistance: Discrepancy Between Downregulation In Vivo and In Vitro
Authors: Wagman Lawrence D , Lambert Scott S , McElduff Aidan , Roth Jesse , Gorden Phillip , Taylor Simeon I
Published: September 2013

Levels of fasting plasma insulin are generally inversely correlated with125I-insulin binding to circulating blood cells. In disease states associated with hyperinsulinemia (e.g., obesity and non-insulin-dependent diabetes mellitus), 125I-insulin binding is usually low. In contrast, 125I-insulin binding to circulating cells may be normal in patients with certain forms of extreme insulin resistance despite marked hyperinsulinemia. To explain this paradox, it has been proposed that postbinding defects in insulin action may give rise to defects in downregulation. We have employed cultured Epstein-Barr virus (EBV)-transformed lymphocytes from eight patients with extreme insulin resistance to address the question of whether there is a defect in the downregulation process in vitro.

In this cell type, insulin leads to a decrease in the number of insulin receptors on the cell surface by accelerating the rate of degradation of insulin receptors. We could not detect any abnormality in in vitro downregulation with cultured EBV-transformed lymphocytes from insulin-resistant patients. The apparent discrepancy between the in vivo and in vitro studies raises the possibility that some factor in the patient's internal milieu may prevent insulin-induced downregulation. An alternative possible explanation might be that the mechanism of downregulation in vitro differs from the mechanism whereby receptor number is regulated in vivo in insulin's target cells.
Insulin Regulation of Na/K Pump Activity in Rat Hepatoma Cells
Authors: Gelehrter Thomas D , Shreve Paul D , Dilworth Vickie M
Published: September 2013

Insulin rapidly increases Na/K pump activity in HTC rat hepatoma cells in tissue culture, as measured by the ouabain-sensitive influx of the potassium analogue 86Rb+. Increased influx is observed within minutes and is maximal (70% above control) within 1–2 h. The effect appears to be mediated by the insulin receptas: (1) the concentration dependence on insulin is identical to that for insulin induction of tyrosine aminotransferase and stimulation of 2-aminoisobutyric acid transport, (2) proinsulin is 6% as potent as insulin, and (3) the effect is blocked by anti-receptor antibodies. The early stimulation of potassium influx is not blocked by cycloheximide and is not associated with an increased number of pump sites as measured by 3H-ouabain binding. The insulin effect is blocked by amiloride, which blocks sodium influx, and is mimicked by the sodium ionophore monensin, which increases sodium influx and intracellular accumulation. Insulin also rapidly increases the initial rate of 22Na+ influx, suggesting that insulin may enhance Na/K pump activity, in part, by increasing intracellular sodium concentration. Incubation of HTC cells with insulin for 24 h causes complete unresponsiveness to the insulin induction of transaminase and stimulation of amino acid transport, a phenomenon mediated by postbinding mechanisms. In contrast, similar incubation with insulin does not cause unresponsiveness to the insulin stimulation of Na/K pump activity. Therefore, the site of regulation of responsiveness to insulin must be distal to, or separate from, those events causing imulation of ion fluxes.