bluuebunny/crossref_metadata_2025 huggingface dataset:
Effects of Acute Changes in Blood Glucose on HbA1c
Authors: Goldstein David E , Peth Steven B , England Jack D , Hess Randall L , Costa Jose Da
Published: September 2013

Although hemoglobin A1c (HbA1c) is generally considered to be an accurate index of long-term blood glucose regulation, several recent studies suggest that HbA1c may be acutely responsive to changes in blood glucose. We have examined the effects of acute changes in glucose concentration in vivo and in vitro on HbA1c. HbA1c was measured by a high-performance liquid chromatography (HPLC) method. HbA1c and plasma glucose were measured in seven diabetic patients and five control subjects before and 2 h after a standard breakfast. Only diabetic patients showed increases in HbA1c and plasma glucose values 2 h after the test meal (Δ HbA1c = 0.87 ± 0.24% and Δ glucose = 210 ± 0.33 mg/dl). The increment in HbA1c correlated significantly with the increment in plasma glucose (r = 0.73, P < 0.05). to examine the lability of these postmeal increments in HbA1c, erythrocytes from pre- and postmeal blood samples were incubated in 0.9% NaCl for 5 h at 37°C and HbA1c was re-measured. After saline incubations %HbA1c in pre- and postmeal blood samples decreased in both diabetics and controls, and from each subject time 0 and 2-h HbA1c values were nearly identical. HbA1c was then measured before and after incubations of erythrocytes from a larger group of diabetic patients (N = 55) and control subjects (N = 7). In both diabetic and control cells HbAlc decreased after saline incubations. Pre- and post-saline HbA1c values in diabetics were (means ± SEM) 10.07 ± 0.30% and 9.15 ± 0.27%, respectively; values in controls were 5.87 ± 0.11% and 5.46 ± 0.09%, respectively. The mean decrement in HbA1c was significantly greater in cells from diabetics than from controls (P < 0.001). In diabetics the HbA1c decrement correlated with both plasma glucose (r = 0.58, P < 0.001) and the preincubation HbA1c (r = 0.55, P < 0.001). After dialysis of hemolysates for 5 h at 37°C or 48–72 h at 4°C, HbA1c values showed decreases comparable to those after saline incubations of intact erythrocytes. However, decreases in HbA1c after dialysis were accompanied by increases in HbA1a+b, findings that were not observed after saline incubation. The results suggest that HbA1c exists in two chromatographically indistinguishable forms: one that represents the major portion of HbA1c in normals and diabetics, is not altered by short-term changes in plasma glucose, and can be estimated by measuring HbA1c after saline incubation of erythrocytes; and a second form that is responsive to short-term fluctuations of the blood glucose level. These labile and stable fractions may be identical to the labile Schiff-base and stable ketoamine forms of HbA1c, which have been previously described. For HbA1c to be an accurate indicator of long-term glucose control, saline incubation of erythrocytes or perhaps dialysis of hemolysates before HbA1c assay may be necessary. Otherwise the assay results will reflect recent changes in blood glucose levels.
Effect of Age on Proinsulin and Insulin Secretory Patterns in Isolated Rat Islets
Authors: Gold G , Reaven G M , Reaven E P
Published: September 2013

Although isolated islets from older rats store ample insulin, they do not respond as efficiently as do islets from young animals to a glucose or leucine challenge. The current study was conducted to determine if this decline in insulin secretion is associated with abnormalities in islet content or conversion of proinsulin, or in the mobilization of insulin that is newly synthesized. Isolated islets from 2-, 12-, and 18-mo-old rats were assayed for total islet proinsulin and insulin content, and after stimulation with glucose the incubation medium was assayed for secreted proinsulin and insulin. In addition, islets from 2- and 12-mo-old donors were pulse-labeled with 3H-leucine, after which the temporal conversion and pattern of release of newly synthesized hormone was compared. The results show that the ratio of proinsulin to insulin declines progressively with age for both the stored and secreted hormone. Incorporation of 3H-leucine into proinsulin during glucose stimulation also decreases with age. However, the onset, time course, and endpoint of the conversion of this labeled proinsulin to insulin did not change with age. Although the fractional secretion rate of newly synthesized hormone (whether proinsulin or insulin) is similar in 2- and 12-mo-old rats, it exceeds that of the unlabeled, or stored, insulin. When the wave of labeled protein enters the secretory pool it apparently equilibrates with a portion of the insulin pool, which is more rapidly secreted than the total pool of insulin. We considered this the glucose responsive pool, and estimate that this pool is actually smaller in islets from 12- as compared with 2-mo-old rats. The reduction in size of this responsive compartment of insulin in the older rats may, in part, explain the decreased rates of stimulated insulin release that OCCUr with age.
Glucose Transport Across Ocular Barriers of the Streptozotocin-Diabetic Rat
Authors: Dimattio Joseph , Altszuler Norman , Ellis Steven , Zadunaisky José A
Published: September 2013

The transport kinetics across the plasma-aqueous and plasma-vitreous barriers were studied in normal and long-term streptozotocin-diabetic rats, using trace amounts of [14C]-L-glucose and [3H]-3-O-methyl-D-glucose. The former is passively transported while the latter uses the same transport-facilitating system as D-glucose. Transport rates of L-glucose were significantly higher in the diabetic rats, with ocular entry rates from the plasma being increased by 69% across both barriers. Thus, the data indicate that in experimental diabetes the passive permeability of the bloodocular barriers is significantly increased.

By contrast, calculated transport rate constants for 3-O-methyl-O-glucose, when adjusted for the hyperglycemia and the increased passive glucose movement, are not altered in the diabetic animal. Nevertheless, there is actually more mass D-glucose movement due to the prevailing hyperglycemia. The present study suggests that although streptozotocin diabetes alters plasma-ocular glucose transport, there is no direct impairment of glucose carrier function.

Alterations in transport occurred at both ocular barriers, suggesting that involvement is general and that both the retinal pigment epithelium and the ciliary epithelium may be affected by the diabetes. It is unknown whether the increase in passive movement is related to the prevailing hyperglycemia or to insulin deficiency or other unknown factors.
Altered Ability of the Liver to Produce Glucose Following a Period of Glucagon Deficiency
Authors: Deri John J , Williams Phillip E , Steiner Kurt E , Cherrington Alan D
Published: September 2013

It is known that the effectiveness of a physiologic increment in glucagon to stimulate glucose production wanes with time even when counterregulatory insulin secretion is prevented. The aim of the present study was to establish whether the converse is true: does glucagon become a more effective stimulus of glucose production following a period of acute hypoglu-cagonemia?

To determine this, somatostatin was infused along with basal replacement amounts of glucagon (0.65 ng/kg - min) and insulin (228 μU/kg · min) into five postabsorptive conscious dogs. After 1.5 h of basal hormone replacement, the glucagon infusion was terminated and a selective fall in the plasma glucagon level (75 ± 6 to 30 ± 4 pg/ml) occurred. This resulted in a drop in tracer (3H-glucose)-determined glucose production from 3.0 ± 0.4 to 1.5 ± 0.3 mg/kg min. The plasma insulin level remained unchanged at 10 ± 1 μU/ml throughout the experiment. Euglycemia (110 ± 4 mg/dl) was maintained by exogenous glucose infusion. After 3 h of glucagon lack, restoration of the glucagon infusion returned the IRG level to control values (78 ± 6 pg/ml) but restored glucose output only partially (42 ± 9%), necessitating continued glucose infusion to preserve euglycemia. Repetition of the experiment in dogs whose pancreatic glucoregulatory feedback loops were broken surgically (two-stage pancreatectomy) rather than pharmacologically resulted in similar findings. It is concluded that glucagon deficiency of 3-h duration leads to a decrease, rather than an increase, in hepatic sensitivity to glucagon.
Insulin and Multiplication Stimulating Activity (an Insulin-like Growth Factor) Stimulate Islet β-Cell Replication in Neonatal Rat Pancreatic Monolayer Cultures
Authors: Rabinovitch Alexander , Quigley Carol , Russell Thomas , Patel Yogesh , Mintz Daniel H
Published: September 2013

A possible role for insulin in stimulating islet β-cell replication was examined in neonatal rat pancreatic monolayer cultures. Addition of insulin to serum-free medium increased the mitotic index and stimulated dose-dependent increases in [3H]-thymidine incorporation in nuclei of islet β-cells in aldehyde-thionine-stained autoradiographs. The effects of insulin were not associated with any significant changes in glucagon or somatostatin levels in the culture media. Multiplication stimulating activity (MSA), an insulin-like growth factor, was about 100-fold more potent than insulin: 3 ng/ml MSA stimulated a half-maximal increase in thymidine labeling of β-cell (+63%, P < 0.005), whereas 300 ng/ml insulin was required for a similar effect. The maximal effects of insulin and MSA were similar, and the combination of maximal stimulatory concentrations of MSA (30 ng/ml) and insulin (3000 ng/ml) was not more effective than either substance added alone, suggesting that both peptides act on the same mechanism(s) regulating β-cell replication. Furthermore, an antibody to the insulin receptor did not prevent the stimulatory effects of either insulin or MSA on thymidine labeling of β-cells. These results demonstrate that insulin can stimulate islet β-cell replication directly, possibly through a receptor for MSA or another insulin-like growth factor.
A Minimal-Model-Based Glucose Clamp Yielding Insulin Sensitivity Independent of Glycemia
Authors: Pacini Giovanni , Finegood Diane T , Bergman Richard N
Published: September 2013

A new technique is introduced for automatic control of the blood sugar (“glucose clamp”) at basal (euglycemic) or elevated (hyperglycemic) levels during variable insulin infusion. A minimal mathematical model of glucose kinetics is implemented on a laboratory minicomputer during clamp experiments. From the measured time course of plasma glucose concentration, the computer estimates the fractional disappearance rate of glucose (X) and calculates the rate of exogenous infusion required to match the desired concentration (M). Eight clamp experiments were performed on three conscious dogs. Euglycemic (N = 4): insulin was infused at 8, 40, and 150 mU/min for sequential 2.5-h periods (I, II, and III); hyperglycemic (N = 4): following the establishment of 140 mg/dl hyperglycemia (period I), insulin was infused at 30 and 120 mU/min for 2.5 h each (periods II and III). Plasma insulin levels [range: 21 ± 9 (basal) to 1729 ± 209 microunits/ml] were matched in comparable periods in the two types of experiments [P > 0.8 (I); P > 0.5 (II), P > 0.2 (III)]. Glucose was successfully clamped for all periods at euglycemia (99.6 ± 0.7% of desired value) and hyperglycemia (100.0 ± 1.1% desired 140-mg/dl value). Glucose infusion rates necessary to achieve stable glycemia were greater at elevated than basal glucose (P < 0.001) despite comparable insulinemia during the three insulin infusion periods. Thus, the glucose infusion rate versus insulin level (M/I) was glycemia-dependent, and therefore not a direct measure of insulin sensitivity. In contrast, the insulin-induced increases in fractional glucose disappearance (ΔX), also provided by our clamp technique during experiments, were highly correlated within insulin (r = 0.08 to r = 0.99, P < 0.01) and independent of glycemia (P > 0.7). The ratio ΔX/ΔI (increase in glucose fractional disappearance rate/incremental insulin level) was a measure of insulin sensitivity independent of glycemia, and could successfully differentiate insulin-resistant from normal animals (P < 0.001; ΔX/ΔI in three animals: 6.9 × 10−4, 5.5 × 10−4, and 1.4 × 10−4 min−1 μ/ml). Thus, this model-dependent glucose clamp technique provides a direct measurement of insulin sensitivity independent of glycemia.
Calcium–Calmodulin-Dependent Myosin Phosphorylation by Pancreatic Islets
Authors: MacDonald Michael J , Kowluru Anjaneyulu
Published: September 2013

Pancreatic islets contain enzyme activity which catalyzes the phosphorylation by MgATP of cardiac, skeletal, or smooth muscle myosin light chains. The enzyme is activated by calcium (Ka = 10 μM) and calmodulin (Ka = 2 μM) and inhibited by trifluoperazine (Ki = 10 μM), a known inhibitor of calmodulin and of insulin secretion. The enzyme binds to a calmodulin affinity column when Ca2+ is present and is eluted when Ca2+ is omitted. These are the properties of myosin light chain kinase. Since phosphorylation of smooth muscle myosin is necessary for its activation by actin, the kinase may have a key role in coupling stimuli that increase intracellular calcium to the contractile processes involved in insulin secretion.
Red Blood Cell Age, Pyruvate Kinase Activity, and Insulin Receptors: Evidence that Monocytes and RBCs May Behave Differently
Authors: Camagna Antonio , Pirro Roberto De , Tardella Laura , Rossetti Luciano , Lauro Renato , Caprari Patrizia , Samoggia Paola , Salvo Giuseppe
Published: September 2013

Data emerging from insulin receptor studies performed on red blood cells (RBCs) and monocytes from the same subject are not always in agreement; dichotomy might occur since variations in mean RBC age are not taken into account or because insulin receptors on the two cell types behave differently.

In the present investigation RBCs from normal male subjects were separated into five populations of different mean age by means of centrifugation of RBCs on a discontinuous gradient of buffered Percoli for 10 min at 1000 x g. Insulin binding varied significantly depending upon the RBC population tested and was closely correlated to the activity of pyruvate kinase (r2 = 0.86), a well-known marker of RBC age.

These data suggested that pyruvate kinase assay might be helpful in studies of RBCs. To confirm this hypothesis, RBCs from 10 normal male subjects and 13 male patients with hemolytic anemia were studied; insulin binding was correlated to pyruvate kinase activity. By adjusting insulin binding to 2 x 109 RBCs/ml the range of data was abnormally high, but it became acceptable after adjusting insulin binding to pyruvate kinase activity (0.75 U/2 x 109 RBCs). The overalr2 = 0.82) but only slightly to reticulocyte number (r2 = 0.56) since not only reticulocytes but also erythrocytes lose receptors during maturation.

Pyruvate kinase activity was measured in RBCs from normal men and from normally menstruating women at the seventh and twenty-fourth days of the cycle; results demonstrated that adjustment of data, according to mean RBC age, broadens dichotomy of monocyte and RBC data.

In conclusion, this paper demonstrates that (1) insulin binding experiments may be carried out on RBCs separated into populations of different mean age; (2) by assaying the pyruvate kinase activity, it is possible to study RBC samples of different mean age and to reveal insulin binding variations provoked by changes in mean RBC age; (3) RBCs lose insulin receptors physiologically and continuously with age; (4) RBC insulin receptors of patients with hemolytic anemia are normal; and (5) monocyte and RBC insulin receptors may behave differently.
Carbohydrate Oxidation and Storage in Obese Non-insulin-dependent Diabetic Patients: Effects of Improving Glycemic Control
Authors: Boden Guenther , Ray Tarun K , Smith Robert H , Owen Oliver E
Published: September 2013

We have determined total body carbohydrate and lipid oxidation rates in response to a standard breakfast in nine obese patients with non-insulin-dependent diabetes mellitus (NIDDM) and in seven age-and weight-matched controls. The patients with NIDDM were studied twice, once while in poor glycemie control (fasting blood glucose concentration 267 ± 24 mg/dl, urinary glucose excretion 28.9 ± 6.3 g/24 h) and again after modest glycemie improvement following 2 mo of fiber treatment (fasting blood glucose 227 ±19 mg/dl, urinary glucose excretion 10.7 ±1.9 g/24 h). Basal carbohydrate (CHO) oxidation rates were normal in patients with NIDDM before and after fiber treatment. However, in patients before fiber treatment the rise in CHO oxidation rates, the reciprocal fall in lipid oxidation rates, and the rise in serum insulin and C-peptide concentrations after the breakfast were all severely blunted. In addition, storage of ingested CHO was significantly reduced (from 55% to 32%, P < 0.05). After fiber treatment, postbreakfast CHO oxidation rates had improved and were no longer significantly lower than control values. In contrast, CHO storage remained suppressed. We conclude that (1) basal CHO oxidation remained normal but that postbreakfast CHO oxidation was impaired in our obese patients with NIDDM. This impairment, however, appeared to be a relatively late event, occurring only during severely uncontrolled NIDDM. (2) Inability to dispose of CHO by storage appeared to be an earlier defect with a greater impact on glucose tolerance than the impairment of CHO oxidation.
Mannose Phosphorylation by Glucokinase from Liver and Transplantable Insulinoma: Cooperativity and Discrimination of Anomers
Authors: Meglasson Martin D , Schinco Miren , Matschinsky Franz M
Published: September 2013

Glucokinase from rat liver or transplantable, radiation-induced insulinomas was partially purified by ion exchange chromatography using DEAE-Cibacron Blue F3GA agarose. Phosphorylation of α,β-D-mannose by glucokinase occurred with cooperative rate dependence on mannose concentration (nH: 1.50). Half-maximal phosphorylation rate occurred at 14 mM α,β-D-mannose. The α- and β-anomers of mannose were phosphorylated with sigmoidal kinetics (nH: 1.57 and 1.42, respectively). The affinity of glucokinase for α-D-mannose is higher than for β-D-mannose (S05: 12 mM versus 19 mM). The maximum phosphorylation rate is slightly higher, about 10%, with β-D-mannose than with α-D-mannose. Islet glucokinase has previously been shown to be chromatographically and kinetically identical to glucokinase from insulinoma and liver; therefore, evidence that glucokinase from these two tissues phosphorylates mannose with cooperative rate dependence and differentiates mannose anomers supports the glucokinase-glucose sensor hypothesis.
Diminished Production of Thromboxane B2 and Prostaglandin E by Stimulated Polymorphonuclear Leukocytes from Insulin-treated Diabetic Subjects
Authors: Qvist Rajes , Larkins Richard G
Published: September 2013

As an initial step to investigate the possibility that abnormal polymorphonuclear leukocyte (PMNL) function in diabetes might be related to abnormalities of arachidonic acid metabolism, products of the cyclooxygenase pathway were assayed in PMNL from 27 insulintreated diabetic subjects and 27 age- and sex-matched nondiabetic subjects. It was found that the major prostanoid products formed were thromboxane B2 (TxB2) and prostaglandin E (PGE). Production of both these substances was greatly enhanced in PMNL from control and diabetic subjects by exposure to a killed preparation of Staphylococcus aureus (S. aureus) or to zymosan.

There was a marked reduction in the production of TxB2 by PMNL from diabetic subjects in response to stimulation by both S. aureus [670 ± 98 (SE) versus 1010 ± 76 pg/106 PMNL/90 min, P < 0.01] and zymosan (583 ± 53 versus 1034 ± 46 pg/106 PMNL/90 min, P < 0.001). Similarly, production of PGE was significantly reduced in diabetics in response to both S. aureus (145 ± 29 versus 232 ± 16 pg/106 PMNL/90 min, P < 0.05) and zymosan (181 ± 21 versus 271 ± 27 pg/ 106 PMNL/90 min, P < 0.01). There was no relation between the plasma glucose at the time of the test and the production of either prostanoid.

Diminished production of cyclooxygenase products of arachidonic acid metabolism should be added to the known abnormalities of PMNL in diabetes. In view of the demonstrated or inferred effects of cyclooxygenase products on aspects of PMNL function, this observation may be important in understanding the pathogenesis of PMNL dysfunction in diabetes.
Structure of the Insulin Receptor of Rat Adipocytes: The Three Interconvertible Redox Forms
Authors: Yip C C , Moule M L
Published: September 2013

Isolated intact rat adipocytes were photoaffinity labeled with radioactive and photoreactive NαB1-monoazi-dobenzoyl insulin (B1-MABI) or NεB29-monoazidobenzoyl insulin (B29-MABI). Polyacrylamide gel electrophoresis of the labeled plasma membranes solubilized in sodium dodecyl sulfate revealed the specific labeling of three receptor species of 380 kDa, 300 kDa, and 230 kDa. Reduction of each species individually produced the subunits of 130 kDa, 90 kDa, and 40 kDa. Exposure of the adipocytes or plasma membranes after photola-beling to sulfhydryl alkylating agents such as N-ethyl-maleimide or p-chloromercuriphenylsulfonate resulted in the appearance of the receptor quantitatively in the 380-kDa form. The effect of the sulfhydryl reagent was concentration dependent and in the case of p-chloromercuriphenylsulfonate the three receptor species reappeared when high concentrations of the reagent were used. Incubation of the adipocytes with low concentrations of dithiothreitol before photolabeling reduced these receptors to discrete lower-molecular-weight forms. In addition, an 85-kDa subunit was now photolabeled by B1-MABI. This subunit was demonstrated to be different from the 90-kDa subunit normally labeled by B29-MABI. We conclude that on the cell surface of the adipocyte, there is ope molecular-weight form of insulin receptor of 380 kDa composed of one 130-kDa, one 90-kDa, one 85-kDa, and two 40-kDa subunits. The 300 kDa and 230 kDa are partially reduced forms of the 380-kDa species. We further postulate that a membrane factor or factors sensitive to sulfhydryl alkylating reagents may be involved in the partial reduction and oxidation of these three redox receptor species. The distribution of these redox receptor species may be related to the cellular or tissue sensitivity to insulin.
Effects of Long-Term Optimization and Short-Term Deterioration of Glycemic Control on Glucose Counterregulation in Type I Diabetes Mellitus
Authors: Bolli Geremia , Feo Pierpaolo De , Cosmo Salvatore De , Perriello Gabriele , Angeletti Gabriella , Ventura Maria Rosa , Santeusanio Fausto , Brunetti Paolo , Gerich John E
Published: September 2013

To assess the effects of glycemie control on glucose counterregulation, rates of plasma glucose recovery from hypoglycemia and counterregulatory hormonal responses were studied in 18 C-peptide-negative patients with insulin-dependent diabetes mellitus (IDDM) before and after either improvement, no change, or deterioration in glycemic control. Hypoglycemia was induced by an i.v. insulin infusion (30 mU/m2 · min for 1 h) after maintenance of euglycemia overnight with i.v. insulin.

In 13 patients with long duration of IDDM (9 ± 0.5 yr, mean ± SEM) and initially poor glycemic control (mean diurnal blood glucose, MBG 199 ± 8 mg/dl, ketoamine-HbA1 12.4 ± 0.2%; nondiabetic subjects 104 ± 4 mg/dl and 6.8 ± 0.09%, respectively), rates of plasma glucose recovery from hypoglycemia (0.30 ± 0.01 versus 0.60 ± 0.01 mg/dl · min in nondiabetic subjects, P < 0.001) and plasma glucagon (AUC 0.56 ± 0.09 versus 6.3 ± 0.50 ng/ml ·150 min in non-diabetic subjects, P < 0.01) and epinephrine (AUC 16.9 ± 0.2 versus 25.7 ± 0.2 ng/ml 150 min in nondiabetic subjects, P < 0.001) responses to hypoglycemia were impaired. Intensive therapy (three daily injections of insulin) instituted in 7 out of 13 IDDM patients for up to 9 mo improved MBG (124 ± 6 mg/dl, P < 0.01) and ketoamine-HbA1 (7.9 ± 0.02%, P < 0.01) but not rates of plasma glucose recovery (0.31 ± 0.01 mg/dl · min) and plasma glucagon (AUC 0.69 ± 0.07 ng/ml · 150 min) and epinephrine (AUC 14.9 ± 0.17 ng/ml ·150 min) responses. Similarly, no changes in glucose counterregulation were observed in the six IDDM patients in whom glycemic control was poorly maintained for 6–8 mo (MBG 201 ± 7 mg/dl, ketoamine-HbA112.3 ± 0.2%).

In five patients with short duration of IDDM (11 ± 0.5 mo) and optimized glycemic control (MBG 131 ± 8 mg/dl, ketoamine-HbA1 7.9 ± 0.2%), rates of plasma glucose recovery, as well as counterregulatory responses of plasma glucagon and epinephrine, were normal. Neither deterioration of glycemic control for 2 wk (MBG 229 ± 12 mg/dl, ketoamine-HbA1 9.1 ± 0.2%) nor its improvement over the 2 subsequent wk (MBG 120 ± 8 mg/dl, ketoamine-HbA1 8.5 ± 0.2%) resulted in any significant change in glucose counterregulation. It is concluded that near-normalization of glycemic control for up to 9 mo is insufficient to reverse the impairment in glucose counterregulation found in long-term IDDM. Short-term deterioration in glycemic control, when not accompanied by alterations in counterregulatory hormone secretion, does not impair glucose counterregulation.
Effect of Insulin Antibodies on Insulin Pharmacokinetics and Glucose Utilization in Insulin-dependent Diabetic Patients
Authors: Waldhäusl Werner K , Bratusch-Marrain Paul , Kruse Viggo , Jensen Ivan , Nowotny Peter , Vierhapper Heinrich
Published: September 2013

To determine the impact of insulin-binding antibodies on total (TIRI) and free insulin (FIRI) as well as on insulin sensitivity, 10 insulin-dependent diabetic patients (IDDM) with poststimulatory C-peptide <100 pmol/L and an insulin binding capacity (IBC) between <1 and 294 μg/L serum were studied during and after a 1-h nonprimed, constant-rate insulin infusion (study 1: 0.057 U/kg body wt, study 2: 0.286 U/kg body wt). Euglycemia was maintained by variable glucose infusion. Control studies were performed in 5 healthy subjects.

Basal TIRI (mU/L) was lowest in healthy subjects (16 ± 1 [SE]) and elevated in diabetic patients (IBC <25 μg/L: 72 ± 11, IBC >25 μg/L: 1772 ± 842), whereas serum concentrations of FIRI were considerably smaller but still two- to threefold greater (P < 0.01) in the patients than in healthy subjects (13 ± 1). After intravenous (i.v.) insulin administration, almost identical increments in serum TIRI were seen in healthy subjects and in diabetic patients with low IBC (<25 μg/L), whereas those with high IBC (<25 μg/L) had a heterogeneous response.

The increments of free serum insulin induced by the infusion of 0.057 and 0.286 U of insulin/kg body wt were almost identical for healthy subjects and diabetic patients, whereas overall glucose utilization, as estimated by the glucose infusion rate necessary to maintain euglycemia, was mostly lower in the diabetic patients (low IBC: study 1: 3.2 ± 0.4 mg/kg min, study 2: 4.0 ± 0.3; high IBC: study 1: 1.2 ± 0.1, study 2: 3.8 ± 0.3) than in healthy subjects (study 1: 3.9 ± 0.6, study 2: 6.1 ± 0.5). Insulin clearance remained constantly within the normal range, but tended to be elevated in diabetic patients with high IBC, who also displayed a 50–100% rise in apparent insulin half-life.

We conclude that elevated insulin-binding capacity: (1) increases serum total insulin, but affects serum free insulin only to a minor extent, and (2) is associated with a rise in apparent free insulin half-life in diabetic patients with high IBC only. (3) Insulin sensitivity is significantly impaired in IDDM and is not improved by short-term normal glycemia; and (4) the relative contribution of insulin antibodies and nonimmunologic factors to impaired insulin action in IDDM cannot be separated by the euglycemic clamp technique.
Effects of Lysosomal Inhibitors on 125I-Insulin and 125I-Asialofetuin Degradation by the Isolated, Perfused Rat Liver and Isolated Rat Hepatocytes
Authors: Ward Walter F , Moss Anita L
Published: September 2013

To further evaluate the role of the lysosomal system in insulin degradation, we have compared the effects of inhibitors of lysosomal function on the degradation of 125I-insulin with 125I-asialofetuin, a lysosomally targeted molecule, by the intact, perfused rat liver and the isolated rat hepatocyte. The inhibitors employed were chloroquine (125 μJLM), NH4CI (10 mM), and leupeptin (50 μg/ml). In the intact, perfused liver the observed inhibition of 125I-asialofetuin degradation at 30 min was as follows: chloroquine, 38%; NH4CI, 32%; and leupeptin, 86%. Chloroquine also inhibited 125Nnsulin degradation in the intact, perfused liver (29%), but NH4CI and leupeptin had no effect. Using the isolated hepatocyte, the observed values for inhibition of 125I-asialofetuin at 60 min were: chloroquine, 85%; NH4CI, 76%; and leupeptin, 81%. Chloroquine produced a 28% inhibition of 125I-insulin degradation, while NH4CI and leupeptin had no effect. Chloroquine and NH4CI decreased cell-associated radioactivity when isolated hepatocytes were incubated with 125I-asialofetuin (leupeptin had no effect), whereas chloroquine caused a 107% increase in cellassociated radioactivity when 125I-insulin was added to the incubation media (NH4CI and leupeptin had no effect), These results indicate that the effects of chloroquine on insulin degradation are an extralysosomal action and that lysosomes appear not to be involved in the physiologic degradation of the insulin molecule.
Measurement of Circulating Human Proinsulin Concentrations Using a Proinsulin-specific Antiserum
Authors: Deacon C F , Conlon J M
Published: September 2013

Antibodies have been raised against biosynthetic human proinsulin that show < 1% cross-reactivity with human insulin and C-peptide. A sensitive (IC50 0.16 pmol/ml; minimum detectable concentration 0.004 pmol/ml) radioimmunoassay has been developed using this antiserum and 125I-proinsulin that will measure proinsulin-like immunoreactivity in human serum without the need for prior separation of insulin or C-peptide. In healthy, fasted subjects (N = 23), the serum proinsulin concentration was 0.015 ± 0.001 pmol/ml (mean ± SEM). In six healthy subjects, serum proinsulin rose to 250% of basal after 120 min in response to 100 g oral carbohydrate, but to only 130% after 60 min following 25 g oral carbohydrate. The proinsulin/total immunoreactive insulin ratio and the proinsulin/C-peptide ratio fell sharply after both high and low carbohydrate loads. Endogenous human serum proinsulin-like immunoreactivity released into the circulation after 100 g carbohydrate was eluted from a Mono Q high-performance, ion-exchange column with the same retention time as biosynthetic human proinsulin. Treatment of biosynthetic proinsulin with trypsin under mild conditions led to a decrease in proinsulin-like immunoreactivity concomitant with an increase in C-peptide and insulin-like immunoreactivity, indicating that the proinsulin-specific antiserum did not preferentially recognize intermediates of proinsulin cleavage.
Effects of NIDDM on Very-Low-Density Lipoprotein Triglyceride and Apolipoprotein B Metabolism: Studies Before and After Sulfonylurea Therapy
Authors: Taskinen M-R , Beltz W F , Harper I , Fields R M , Schonfeld G , Grundy S M , Howard B V
Published: September 2013

To evaluate mechanisms of diabetes-induced changes in very-low-density lipoprotein (VLDL), VLDL triglyceride (TG) and VLDL apolipoprotein B (apoB) metabolism were studied in 12 obese Pima Indian control subjects and in 15 Pima Indian obese non-insulin-dependent diabetics. Eleven of the diabetics were restudied after reduction of hyperglycemia with oral sulfonylurea therapy. In addition, adipose, muscle, and postheparin lipoprotein lipase and postheparin hepatic lipase activities were measured in all subjects. Obese diabetics as compared with obese controls showed a trend toward increased production of VLDL TG (46 ± 4 vs. 35 ± 6 g/day, P = .10) but not of VLDL apoB (1595 ± 106 vs. 1597 ± 164 mg/day, NS); production of VLDL TG declined to control levels (33 ± 4 g/day, P < .05) during therapy, whereas there was no change in production of VLDL apoB. Diabetics had a clearance defect for VLDL, indicated by significantly lower fractional catabolic rates for both VLDL TG (10.6 ± .9 vs. 13.1 ± .9 pools/ day, P < .05) and VLDL apoB (5.6 ± .4 vs. 7.5 ± 0.7, P < .05) as compared with controls; fractional catabolic rates increased after therapy (to 13.3 ± 1.5, P < .05, and 6.7 ± .4, P < .05, respectively). In the diabetics, this decrease in clearance was accompanied by a lower adipose lipoprotein lipase (.30 ± .09 vs. .92 ± .25 μmol · g−1 · h−1 P < .01), which increased during therapy (to .61 ± .17, P < .05). Hepatic lipase also decreased significantly after therapy (27.4 ± 3.6 to 26.4 ± 3.2, P < .01). Composition of VLDL in diabetics was also abnormal, indicated by a higher TG/apoB ratio (14.7 ± .6 vs. 11.7 ± .8, P < .01); this ratio fell during therapy (to 12.5 ± .8, P < .05). The data indicate there are multiple abnormalities in structure and metabolism of VLDL in non-insulin-dependent diabetics. Control of hyperglycemia with sulfonylureas has the capability of reversing some of these abnormalities.
Slow Component-a of Axonal Transport, Nerve myo-Inositol, and Aldose Reductase Inhibition in Streptozocin-Diabetic Rats
Authors: Tomlinson David R , Sidenius Per , Larsen Jørn Rolighed
Published: September 2013

This study measured sugars and polyols, weight/unit length, and slow component-a of axonal transport (SCa) in dorsal root afferents of the sciatic nerves of control rats and rats with streptozocin (STZ)-induced diabetes of 4-wk duration. The effects of two treatments—aldose reductase inhibition ‘Statil (“Static” is a trademark; the property of Imperial Chemical Industries PLC.) ICI 128436 at 25 mg/kg/day, p.o.’ and myo-inositol supplementation (650 mg/kg/day, p.o.)—were studied in control and diabetic groups. Inclusion of untreated controls and diabetics gave a total of six groups for the study. The treatments were begun on the day after injection of STZ and were maintained throughout the protocol.

The sciatic nerves of the diabetic (untreated) rats showed accumulation of sorbitol and fructose, depletion of myo-inositol, and an 8% increase in weight/unit length. All of these abnormalities were prevented by treatment with Statil. Treatment of diabetic rats with myo-inositol prevented its depletion in the sciatic nerve, but did not affect the accumulation of sorbitol and fructose nor the increase in weight/unit length. Neither treatment exerted any apparent effect on body weight, blood glucose, nerve weight, or nerve sugars and polyols in the control rats.

The diabetic rats showed a retardation of the wave of transported-labeled protein (shown as increased leftward skewness of the wave) and a reduction in mean transport velocity (calculated as the mean velocity for all segments contributing to the transport wave: 0.96 ± 0.09 mm/day in diabetics versus 1.15 ± 0.07mm/day in controls). These abnormalities were not affected significantly in the diabetic groups by treatment with either Statil or with myo-inositol.
Effects of Nephron Loss on Glomerular Hemodynamics and Morphology in Diabetic Rats
Authors: O'Donnell M P , Kasiske B L , Daniels F X , Keane W F
Published: September 2013

Loss of renal mass in rats with experimental diabetes mellitus leads to exaggerated hypertrophy of remaining nephrons and accelerated diabetic glomerulopathy. To examine factors responsible for glomerular injury in this setting, rats with preexisting diabetes were subjected to unilateral nephrectomy. Micropuncture studies and evaluation of glomerular morphology were performed 2–3 mo later. Nephrectomized diabetic rats demonstrated significant increases in kidney weight, superficial nephron glomerular filtration rate, and superficial nephron plasma flow compared with two-kidney diabetic rats and nephrectomized nondiabetic controls. Glomerular capillary hydraulic pressure was comparable in two-kidney and nephrectomized diabetic rats and was significantly reduced compared with nephrectomized nondiabetic controls. Nephrectomized diabetic rats demonstrated significant albuminuria, mesangial matrix expansion, and focal glomerulosclerosis, whereas two-kidney diabetic rats and nephrectomized nondiabetic controls showed only minimal alterations in glomerular morphology. It is concluded that diabetic rats can undergo glomerular functional compensation in response to nephron loss. Moreover, accelerated glomerular injury caused by nephron loss in diabetic rats could not be attributed to increased glomerular capillary pressure.
In Vivo Regulation of Non-Insulin-Mediated and Insulin-Mediated Glucose Uptake by Cortisol
Authors: Baron Alain D , Wallace Penny , Brechtel Ginger
Published: September 2013

In vivo glucose uptake (Rd) occurs via two mechanisms: insulin-mediated glucose uptake (IMGU), which occurs in insulin-sensitive tissues, and noninsulin-mediated glucose uptake (NIMGU), which occurs in both insulin-sensitive and non-insulinsensitive tissues. To determine whether these two pathways for in vivo glucose disposal are regulated independently, we studied the effect of stress levels of cortisol on IMGU and NIMGU in seven normal subjects after an overnight fast. To study NIMGU, somatostatin (SRIF, 600 μg/h) was infused to suppress endogenous insulin secretion and create severe insulinopenia, and glucose turnover was measured isotopically while serum glucose was clamped at ∼200 mg/dl for 240 min. Separate studies were performed during the overnight infusion of saline or hydrocortisone (HCT; 2.0 μg · kg−1 · min1). The final 120 min of each study were used for data analysis. Under these conditions, insulin action is absent, and Rd = NIMGU. NIMGU was 204 ± 11 mg/min and 208 ± 8 mg/dl during saline and HCT, respectively (P NS). Therefore, HCT did not modulate NIMGU. To measure the effect of cortisol on Rd, hyperglycemic (200 mg/dl)-hyperinsulinemic clamp studies (30 mU · m−2 · min1) were performed during the infusion of saline or HCT. The results demonstrate that during saline infusion, steady-state rates of Rd (10.4 ± 0.8 mg · kg−1 · min1) were achieved by 160 min; in contrast, during HCT infusion, Rd never reached steady state but increased from 4.5 ± 0.2 in the 2nd h to 7.6 ± 0.4 mg kg1 min1 in the 4th h, P < .01. In conclusion, 1) cortisol has no measurable modulatory effect on NIMGU, 2) IMGU and NIMGU are independently regulated and functionally distinct, and 3) cortisol causes insulin resistance by causing a decrease in the rate at which insulin activates the glucose uptake system.
Cytosolic Free-Calcium Concentrations in Normal Pancreatic Islet Cells: Effect of Secretagogues and Somatostatin
Authors: Sussman Karl E , Leitner J Wayne , Draznin Boris
Published: September 2013

We have assessed the effect of somatostatin on glucose-, potassium-, forskolin-, and dibutyryl cAMP-induced changes in cytosolic free [Ca2+] in normal rat pancreatic islet cells with the new Ca2+ indicator fura 2. The cytosolic free [Ca2+] in islet cells incubated with nonstimulatory concentrations of glucose (30 mg/dl) ranged from 54 to 64 nM. In the presence of extracellular Ca2+ (1 mM), glucose (300 mg/dl) rapidly increased the cytosolic free [Ca2+] to a level of 90-110 nM. In the absence of extracellular Ca2+, glucose failed to increase the cytosolic free [Ca2+], which remained at a level of 55-60 nM. Somatostatin inhibited glucose-induced increases in cytosolic free [Ca2+] in a dose-dependent manner (maximal inhibition was 34%). Half-maximal inhibition was observed at 10−9 M somatostatin, which correlated well with somatostatin binding to islet cells (Kd = 2.6 × 1010 M). Potassium (50 mM) rapidly increased the cytosolic free [Ca2+] to 110-120 nM, and its effect was not influenced by the presence of somatostatin. Forskolin (20 μM) and dibutyryl cAMP (1 mM) rapidly increased cytosolic free Ca2+ both in the presence and absence of extracellular Ca2+. More than 80% of the overall increase in cytosolic free-Ca2+ levels could be accounted for by the mobilization of intracellular Ca2+ stores. Somatostatin effectively blocked the forskolin effect (32% inhibition) but not the dibutyryl cAMPinduced effect. Somatostatin appears to inhibit secretagogue-induced increases in cytosolic free [Ca2+] by interfering with cAMP production and probably with Ca2+ transport across the cell membrane.
Fibronectin Binding to Glomerular Basement Membrane is Altered in Diabetes
Authors: Cohen Margo Panush , Saini Romesh , Klepser Henry , Vasanthi L G
Published: September 2013

We examined the ability of fibronectin, an extracellular glycoprotein that interacts with cell surfaces and matrix components, to bind to glomerular basement membrane and the effect of diabetes on this binding. 125I-Iabeled fibronectin binding to rat glomerular basement membrane (GBM) was dose dependent, related to time and amount of basement membrane, and inhibited by unlabeled fibronectin but not by unrelated proteins. Binding was reduced ∼60% when GBM was pretreated with collagenase and ∼24% when pretreated with chondroitinase plus heparinase. Treatment with NaCI had little effect on binding, whereas reduction with p-mercaptoethanol removed ∼25% of the bound 125I-fibronectin. Binding to samples prepared from rats with streptozocin-induced diabetes was significantly increased compared with that observed with control preparations at all concentrations of fibronectin and of basement membrane tested. The findings provide direct evidence that fibronectin binds to GBM and that this binding, which represents a biologic function of the protein, is enhanced in diabetes.
Decreased Acylation of Phosphatidylcholine in Diabetic Rat Erythrocytes
Authors: Le Petit-Thévenin Josette , Nobili Odette , Boyer Jean
Published: September 2013

We investigated the effects of streptozocin-induced diabetes on composition and metabolism of rat erythrocyte lipids. Diabetes produced no change in contents of cholesterol, total phospholipids, and proportions of phosphatidylcholine and phosphatidylethanolamine in phospholipids. The acylation of total phospholipids with palmitic, oleic, or arachidonic acids was decreased (P < .01) in intact erythrocytes from diabetic versus control animals. This anomaly was underlaid by a decrease (P < .01) in acylation of phosphatidylcholine, whereas phosphatidylethanolamine was unaffected. The impaired acylation of phosphatidylcholine was unchanged in vitro by insulin or coenzyme A but was restored to control values by ATP and by insulin treatment of the diabetic rats. We conclude that diabetes specifically alters the acylation of at least phosphatidylcholine in rat erythrocyte, an effect that might modify the remodeling of erythrocyte phospholipids and thereby the membrane function.
Pancreastatin Inhibits Insulin Secretion and Stimulates Glucagon Secretion in Mice
Authors: Ahrén Bo , Lindskog Stefan , Tatemoto Kazuhiko , Efendić Suad
Published: September 2013

Recently a new peptide, pancreastatin, was isolated from porcine pancreatic extracts. It contains 49 amino acids and shows a structural similarity to chromogranin A, which occurs in secretory granules of the endocrine pancreas. Furthermore, pancreastatin has been found to inhibit glucose-induced insulin secretion in the perfused rat pancreas. However, its effects under in vivo conditions have never been studied. We have therefore investigated the effects of this peptide on insulin and glucagon secretion in vivo in the mouse. We found that an intravenous injection of pancreastatin (4.0 nmol/kg) lowered basal plasma insulin concentration at 6 min from 55 ± 8 μU/ml in control mice to 21 ± 7 microU/ml (P < .01). The peptide also inhibited the plasma insulin response to both glucose (P < .01) and the cholinergic agonist carbachol (P < .001). Furthermore, 2 min after injection of pancreastatin, plasma glucagon concentration had increased to 301 ± 19 pg/ml compared to 190 ± 12 pg/ml in control mice (P < .001). The peptide did not, however, affect the carbachol-induced plasma glucagon response. In addition, pancreastatin induced a transient hyperglycemia. Combined adrenergic blockade by means of a pretreatment of phentolamine and propranolol did not prevent pancreastatin from exerting its effects on plasma insulin levels, whereas the increase in plasma glucagon levels was abolished. Thus, in the mouse, the newly discovered intrapancreatic peptide pancreastatin 1) lowers baseline plasma insulin levels, 2) inhibits glucose- and cholinergically induced insulin secretion, 3) stimulates baseline glucagon secretion, and 4) induces hyperglycemia. The effects seen were modest in potency, and the effects on plasma insulin levels were seen also after combined α- and β-adrenoreceptor antagonism. We suggest that pancreastatin could be a regulator of islet function.
Effect of Intensive Diabetes Treatment on Low-Density Lipoprotein Apolipoprotein B Kinetics in Type I Diabetes
Authors: Rosenstock Julio , Vega Gloria Lena , Raskin Philip
Published: September 2013

The metabolism of low-density lipoprotein (LDL) was studied in six insulin-dependent (type I) diabetic patients during a 7-wk period of conventional and intensive therapy with insulin. Plasma glucose and HbA1c were normalized, demonstrating the effectiveness of our intensive treatment program. Plasma lipoprotein profiles and LDL apolipoprotein B kinetic parameters were estimated during conventional and then during intensive therapy for each patient. Intensive therapy resulted in a significant reduction of plasma and LDL cholesterol and an increase in high-density lipoprotein (HDL) cholesterol. The lower LDL levels resulted from a decreased production of lipoprotein rather than an increased fractional catabolic rate. These results are consistent with our previous observations of very-low-density lipoprotein (VLDL) metabolism during intensive therapy. VLDL production is significantly reduced; thus, a decreased production of LDL supports the contention that intensive therapy with insulin in normolipemic type I diabetic patients reduces the production of lipoproteins containing apolipoprotein B rather than increasing the clearance, and therapy also increases HDL cholesterol. Both of these effects may be beneficial in reducing the risk for coronary heart disease in type I diabetes.
Effect of Statil (ICI 128436) on Erythrocyte Viscosity In Vitro
Authors: Rillaerts Eddy G , Vertommen Jan J , De Leeuw Ivo H
Published: September 2013

The hypothesis that sorbitol accumulation could contribute to a reduced erythrocyte deformability in diabetes was investigated. Erythrocyte sorbitol and erythrocyte viscosity at high and low shear rates were studied in 20 insulin-dependent diabetic (IDDM) and 20 matched control subjects. An increased erythrocyte sorbitol and an increased low-shear erythrocyte viscosity were found in the IDDM patients, but there was no significant correlation (r = .11, NS) between the parameters. Incubation (3 h, 37 degrees C) in a Krebs buffer containing 33.3 mM glucose resulted in a significant sorbitol accumulation, but erythrocyte viscosity was not affected. Despite this fact, addition of 1 mM statil (ICI 128436) in the 5.5- and 33.3-mM glucose media not only prevented erythrocyte sorbitol accumulation but also improved erythrocyte viscosity in diabetic and control subjects. The effect was more pronounced at the low (∼16%) than at the high (∼2%) shear rate. The effect on erythrocyte viscosity disappeared by washing the erythrocytes after incubation, although erythrocyte sorbitol remained different. Our results suggest that sorbitol accumulation does not contribute to an increased erythrocyte viscosity in diabetes, and statil shows a positive effect on erythrocyte viscosity independent of its aldose reductase–inhibiting property.