bluuebunny/crossref_metadata_2025 huggingface dataset:
1,25-Dihydroxyvitamin D3 and 24,25-Dihydroxyvitamin D3 Production by Isolated Renal Slices is Modulated by Diabetes and Insulin in the Rat
Authors: Wongsurawat Nirandon , Armbrecht H James , Zenser Terry V , Davis Bernard B , Thomas Mary L , Forte Leonard R
Published: September 2013

The alterations of mineral homeostasis observed in the streptozotocin-induced diabetic rat have been attributed to low circulating levels of 1,25-dihydroxyvitamin D3 [1,25-(OH)2-D3], the biologically active metabolite of vitamin D3. However, the effect of diabetes and subsequent insulin repletion on production of 1,25-(OH)2-D3 and 24,25-(OH)2-D3 by the kidney has not been investigated. We studied the renal production of 1,25-(OH)2-D3 and 24,25-(OH)2-D3 in diabetic and insulin-treated diabetic rats using the renal slice technique. In rats made diabetic with streptozotocin, the renal production of 1,25-(OH)2-D3 was markedly reduced (0.195 ± 0.011 pg/min/mg) compared with controls (0.685 ± 0.107 pg/min/ mg, P < 0.05). The renal production of 24,25-(OH)2-D3 in diabetic rats was markedly increased (0.529 ± 0.052 pg/min/mg) compared with controls (0.233 ± 0.035 pg/min/mg, P < 0.05). Treatment of diabetic rats with insulin for 12 days resulted in a significant increase in 1,25-(OH)2-D3 production (0.331 ± 0.053 pg/min/mg) and decrease in 24,25-(OH)2-D3 production (0.329 ± 0.054 pg/min/mg). The possibility that diabetes decreased renal 1,25-(OH)2-D3 production by decreasing parathyroid hormone (PTH) secretion or depressing the action of PTH on the kidney was also studied. Diabetes caused no decrease in serum PTH levels relative to the control group and urinary cyclic AMP excretion in the diabetic group was not depressed. The cyclic AMP content of diabetic rat kidney slices in response to PTH was similar to controls (39.8 ± 3.3 versus 39.7 ±4.6 pmol/mg wet weight). Serum immunoreactive calcitonin (iCT) was significantly depressed in diabetic rats compared with controls. Insulin treatment of diabetic rats resulted in a marked increase in serum iCT. These data suggest the impairment of renal 1,25-(OH)2-D3 production was not related to impairment of the PTH activation of adenylate cyclase in the diabetic State.
Cysteamine Blocks Somatostatin Secretion Without Altering the Course of Insulin or Glucagon Release: A New Model for the Study of Islet Function
Authors: Sorenson Robert L , Grouse Lynette H , Elde Robert P
Published: September 2013

Cysteamine (300 mg/kg) administered subcutaneously depletes pancreatic somatostatin to 36% of control levels, but does not alter pancreatic insulin or glucagon content. Although perfusion of pancreata from normal animals with glucose (300 mg/dl) markedly stimulated somatostatin release, pancreata from cysteamine-treated animals failed to secrete somatostatin in response to glucose. Cysteamine treatment was without effect on insulin and glucagon release under the conditions tested. The isolated perfused pancreas from the cysteamine-treated rat provides a model for further investigations into regulation of islet hormone release in the absence of stimulated somatostatin release.
Examination of the Role of the Pituitary-Adrenocortical Axis, Counterregulatory Hormones, and Insulin Clearance in Variable Nocturnal Insulin Requirements in Insulin-dependent Diabetes
Authors: Skor Donald A , White Neil H , Thomas Lorraine , Shah Suresh D , Cryer Philip E , Santiago Julio V
Published: September 2013

In insulin-dependent diabetics, insulin requirements increase significantly after 0600 h, resulting in prebreakfast hyperglycemia with either conventional insulin therapy or constant insulin infusions with insulin infusion devices. In order to clarify the role of the pituitary-adrenocortical axis and further examine the mechanisms of the phenomenon of nocturnal variability in insulin requirements, we studied five IDDs using a closed-loop insulin infusion device (Biostator, GCIIS). The subjects were given saline (SAL) or dexamethasone (DEX) i.v. from 1800 to 0900 h on successive nights. From 2400–0300 to 0600–0900 h, mean insulin infusion rates required to maintain blood glucose values between 109 and 120 mg/dl increased by 0.21 ± 0.05 mU/kg/min during the SAL infusion, and 0.16 ± 0.04 mU/kg/min during the DEX infusion, when plasma cortisols were suppressed to ≤ 2 μg/dl. Mean free insulin concentrations did not increase and remained constant throughout both study nights in spite of the significantly higher 0600–0900-h insulin infusion rates. Growth hormone, glucagon, epinephrine, and norepinephrine concentrations showed normal nocturnal and early morning patterns during both study nights. We conclude that the nocturnal variability in insulin requirements persists despite suppression of the pituitary-adrenocortical axis, and that increased free insulin clearance or degradation may contribute to the “dawn phenomenon” of rising prebreakfast glucose despite constant insulin infusion.
The Effects of Streptozotocin Diabetes on the Activities of Rat Liver GlycosyItransferases
Authors: Tepperman Helen M , Dewitt Janet , Tepperman Jay
Published: September 2013

The effects of streptozotocin diabetes on the activities of rat liver glycosy Itransferase enzymes have been investigated. Liver microsomal fractions were prepared from rats that had been injected with streptozotocin (65 mg/kg, i.v.) 3 wk to 2 mo earlier. Preparations from diabetic rats had decreased activities of N-acetylglucosaminyl transferase compared with those of agematched controls (0.98 ± 0.11 nmol transferred per mg protein in 30 min versus 3.19 ± 0.34 for controls, P < 0.001). Galactosyltransferase activity was also lower in diabetic rat livers (1.48 ± 0.26 nmol transferred per mg protein in 30 min versus 3.32 ± 0.56 for controls, P < 0.025). Sialytransferase activities were not significantly different between diabetic and control rat livers. There were no significant differences between the diabetic and control rat liver microsomes in the activities of UDP N-acetylglucosamine pyrophosphatase, UDP galactose pyrophosphatase, or CMP sialic acid phosphatase. The glycosidases, N-acetylglucosaminidase and galactosidase, had similar activities in the livers of both groups of rats. Sialidase activity could not be detected in microsomal preparations from either diabetic or control rat livers. These results are discussed in relation to our previously reported alterations in glycosyltransferase activities, and plasma membrane glycoprotein composition in the livers of rats made insulin-resistant by a carbohydrate-free, high-fat diet and to the observation of Carter and his colleagues (FEBS Lett. 1979; 104:389–92.) that streptozotocin diabetes alters the glycoprotein composition of rat liver plasma membranes.
Physical Stability of Insulin Formulations
Authors: Lougheed W D , Albisser A M , Martindale H M , Chow J C , Clement J R
Published: September 2013

Insulin aggregation remains a fundamental obstacle to the long-term application of many insulin infusion systems. We here report the effects of physiologic and nonphysiologic compounds on the aggregation behavior of crystalline zinc insulin (CZI) solutions. Under conditions chosen to simulate the most severe that would be encountered in delivery systems (presence of air, continuous motion, and elevated temperature), both highly purified and regular CZI at 5 U/ml formed turbid gels in 5 days. At concentrations of 100 and 500 U/ml stability was increased with turbid gels forming at 12 and 15 days, respectively. Under identical conditions, 5 U/ml CZI formulations containing the physiologic surfactant lysophosphatidylcholine (0.02%) or the synthetic surfactants SDS (1%), Brij 35 (0.1%), Tween (0.01%), or Triton X (0.01%) retained a transmittance at 540 nm of >96% for 67–150 days. These nonionic and ionic surfactants containing the hydrophobic group, CH3(CH2)N, with N = 7–16, remarkably stabilized CZI formulations while those lacking such groups demonstrated little or no effect. The alcohols glycerol (30–50%) and isopropanol (10–50%) were moderately effective stabilizers. Silicone rubber drastically accelerated aggregation in all but one formulation (1% SDS). Emphasis in this study was placed on the properties of 5-U/ml formulations. Controls run at higher concentrations indicated a positive correlation between concentration and stability. It was concluded that the aggregation of insulin into high-molecular-weight polymers may be inhibited by reducing the effective polarity of the solvent. In this regard, anionic and nonionic surfactants containing appropriately long hydrophobic groups demonstrated the greatest degree of stabilization. Finally, of all the medical grade materials likely to be used in pumps, silicone rubber is the most active in promoting insulin aggregation.
Normoglycemia after Reflux of Islet-containing Pancreatic Fragments into the Splenic Vascular Bed in Dogs
Authors: Warnock G L , Rajotte R V , Procyshyn A W
Published: September 2013

Canine pancreata obtained at total pancreatectomy were cannulated via the ducts and perfused with collagenase to prepare a tissue suspension that was isografted into the spleen (preparation ⋍2 h, mean graft vol = 10 ± 1 ml containing 24% of the B-cell mass/pancreas). In 13 dogs the tissue was implanted by reflux into terminal splenic veins: two died postoperatively, and in two the intrasplenic vein wall was inadvertently punctured during cannulation. In the remaining nine, mean fasting blood glucose (BG) was ≤150 mg/dl initially; one was killed at 2 wk (distemper) and one at 6 wk (sepsis, diabetes), and one died at 9 wk (intestinal obstruction). Mean BG was 94 ± 4 mg/dl at 1 mo and remained in this range until the dogs were killed at 5 mo (91 ± 1 3 mg/dl). During glucose-tolerance testing 1 wk preimplantation and 1 mo and 2-3 mo postimplant, mean values were: K (decline in glucose concentration, %/min), 3.4 ± 0.2,1.4 ± 0.1, and 1.5 ± 0.1; peak insulin μU/ml), 50 ± 5,12 ± 1, and 11 ± 2; fasting serum glucagon (mg/ml), 33 ± 3, 59 ± 12, and 53 ± 9, with no change in the glucagon response. Histologically, the spleens contained prominent islets. In five other dogs, the tissue was injected into the splenic pulp: mean BG rose to ≥250 mg/dl at 2 wk (compared with initial series, P < 0.001) and remained elevated until death at 6 wk, when histologic examination of the spleens showed severe fibrosis and no islets. Apancreatic controls (N = 4) survived 10 ± 3 days; BG was 343 ± 11 mg/dl terminally. We conclude that this modified method for collagenase perfusion of a single large-mammal pancreas via the ducts provides sufficient viable islets to induce prolonged normoglycemia (5 mo) and preserve the response to glucose challenge. Reflux of pancreatic fragments into splenic veins appears more efficient than intrapulp implantation.
Quantitative Determination of Islet Cell Surface Antibodies Using 125I-Protein A
Authors: Huen A H -J , Haneda M , Freedman Z , Lernmark A , Rubenstein A H
Published: September 2013

A quantitative method to measure islet cell surface antibodies in human patients has been developed using 125l-protein A. Isolated, dispersed, viable rat islet cells prepared by collagenase digestion were fixed in 4% paraformaldehyde to allow storage for up to 7 wk at 4°C. Human sera, heat inactivated and adsorbed with rat liver and kidney powder (100 mg/ml), were incubated with the fixed cells (50 × 103) for 60 min at 37° C. Thereafter the cells were washed and exposed to 5 × 105 cpm 125I-protein A, which binds to IgG attached to the cell surface. Assay precision (14%) and reproducibility (16%) were established by repeated analysis of pooled sera from healthy individuals and IDDM patients using pooled batches of islet cells. Using this method, islet cell surface antibodies were detected in 35% of insulin-dependent diabetic patients.
Impaired Counterregulation of Hypoglycemia in Insulin-dependent Diabetes Mellitus
Authors: Kleinbaum Jerry , Shamoon Harry
Published: September 2013

We evaluated the recovery of blood glucose after insulin-induced hypoglycemia in six insulin-dependent diabetics (insulin-infused and initially euglycemic) and six normal controls after comparable reductions in plasma glucose. In contrast to controls, the recovery of plasma glucose was delayed in diabetics (2-h plasma glucose 80 ± 5 mg/dl and 58 ± 5 mg/dl, respectively, P < 0.01). This delay was due to the absence of a rebound in hepatic glucose output in the diabetics, whereas glucose output rose two- to threefold above baseline in normals. The impaired rebound in glucose output in diabetics could not be attributed to hyperinsulinemia. Rather, hypoglycemia-induced secretion of epinephrine and glucagon was reduced in thediabetics by 60–80% as compared with normals (P < 0.001). The diabetics did not suffer from overt neuropathy and plasma cortisol, growth hormone, and norepinephrine increased normally following hypoglycemia. The data suggest that prolonged hypoglycemia may frequently occur in tightly controlled type I diabetics because of impaired rebound in hepatic glucose release which in turn may be a consequence of reduced secretion of epinephrine and glucagon.
Immunoglobulin from Insulin-dependent Diabetic Children Inhibits Glucose-induced Insulin Release
Authors: Kanatsuna Takahiro , Bæekkeskov Steinunn , Lernmark Åke , Ludvigsson Johnny
Published: September 2013

Immunogiobulin was separated from islet cell antibody positive plasma of six children with newly diagnosed insulin-dependent (type I) diabetes mellitus. The dynamics of insulin release in response to glucose and partially purified antibodies were determined in dispersed rat islet cells perifused on small columns of Biogel P-2 beads. After perifusion at 5.5 mmol/L D-glucose in the presence of healthy control immunoglobulin, the rate of insulin release increased in a biphasic manner after stimulation with 30 mmol/L D-glucose. In cells exposed to diabetic immunoglobulin, 30 mmol/L D-glucose had little, if any, effect on insulin release. These results suggest that islet cell antibodies in insulin-dependent diabetes may interfere with the insulin release mechanisms in the pancreatic B-cells.
Parallel Decreases in the Expression of Receptors for Insulin and Insulin-like Growth Factor I in a Mutant Human Fibroblast Line
Authors: Massague Joan , Freidenberg Gary F , Olefsky Jerrold M , Czech Michael P
Published: September 2013

The receptors for insulin and the insulin-like growth factor (IGF) I are two structurally homologous disulfide-linked multisubunit complexes of apparent Mr = 350,000. The similar subunit structures of these two types of receptors suggested that their genetic expression might be affected by common genetic defects. We have examined this possibility in an insulinresistant, diabetic patient who exhibits decreased insulin binding activity. The receptors for IGF-I and insulin in skin fibroblasts from this patient were affinity labeled with 125I-IGF-I and 125I-insulin, respectively, and visualized by electrophoresis and autoradiography in polyacrylamide gels. Control fibroblasts exhibited the usual affinity labeling of the disulfide-linked Mr = 350,000 insulin and IGF-I receptor structures.

The intensity of labeling of both receptor types in the patient's fibroblasts was less than in control fibroblasts. Binding data indicated that this decrease is due to a decreased receptor number with little or no decrease in affinity for the respective ligands. The highaffinity bIGF-II receptor in fibroblasts affinity labeled with 125I-IGF-II or 125I-IGF-I consists of a single polypeptide not disulfide linked to any other membrane component. The molecular size and intensity of labeling of the IGF-II receptor in the patient's fibroblasts were unaltered when compared with those of controls. These observations suggest that a common genetic defect alters the expression of the homologous receptor structures for insulin and IGF-I.
Muscle Capillary Basement Membrane Width and Its Relationship to Diabetes Mellitus in Monozygotic Twins
Authors: Ganda O P , Williamson J R , Soeldner J S , Gleason R E , Kilo C , Kaldany A , Miller J P , Garovoy M R , Carpenter C B
Published: September 2013

Quadriceps (Q) and gastrocnemius (G) muscle capillary basement membrane width (CBMW) were measured in 18 pairs of monozygotic (MZ) twins. Thirteen of these twin pairs were discordant for insulin-dependent diabetes (IDD) and five pairs were concordant for either IDD (two pairs) or for non-insulin-dependent diabetes (NIDD). In 12 of the 13 nondiabetic (ND) twin mates of IDD, 50 oral glucose tolerance tests performed in the years before or after determination of CBMW revealed mean blood glucose levels in the 36–52 percentile range, compared with normal controls.

The mean (±SD) age at the onset of IDD in discordant twins was 18.7 ± 10.1 (range 8–37) yr and the mean duration of discordance at the time of biopsy was 13.6 ± 8.3 (range 3–32) yr. CBMW data were compared within each twin (Q versus G) and between twin mates and age- and sex-matched controls. Overall, CBMW of IDD twins was greater than that of their ND twin mates. Differences between IDD and ND twins, however, were much more marked in gastrocnemius (1859 ± 643 versus 1222 ± 307 Å, P < 0.0003) than in quadriceps (1291 ±319 versus 1112 ± 302 Å; P < 0.04). CBMW in gastrocnemius was significantly thicker than that in the quadriceps of IDD twins (t = 4.55, P < 0.0008) but not in their ND twin mates (t = 1.15, P < 0.27). CMBW was significantly thicker in IDD than in their ND twin mates (in quadriceps and/or gastrocnemius) in 10 of the 12 twin pairs. Ten of 12 IDD twins (no tissue was available from 1 IDD twin) and 8 of their 13 ND twin mates had evidence of quad-riceps CBMT compared with controls. Repeat biopsies (taken 3–4 yr after the initial biopsy) of quadriceps muscle from five pairs of MZ twins (discordant for IDD) revealed only minor changes in mean CBMW in all but 1 subject.

The muscle CBMW of IDD twins is frequently, but not invariably, “thicker” than that of their ND twin mates. Although some ND twin mates of IDD may have “thick” CBM, this observation does not necessarily imply a greater propensity to development of diabetes.

Further studies are required to elucidate the interactions of genetic and local tissue-environmental factors in modulating the development of diabetic vascular disease.
Insulin-Glucagon Relationships in the Defense Against Hypoglycemia
Authors: Unger Roger H
Published: September 2013

The Berson Memorial Lecture was established to perpetuate the memory of a great scientist, Dr. Solomon Berson (Figure 1), who died tragically at age 53, just 5 years before Dr. Rosalyn Yalow received the Nobel Prize in Physiology or Medicine for their development of the radioimmunoassay. I am deeply honored to have been selected to be the first Berson Memorial Lecturer, but for me the award has a special significance that transcends the honor itself. This is because, in my eyes, Berson was more than the legendary figure whose brilliance as a scientist, clinician, mathematician, and classical musician won him the admiration and affection of his colleagues. To me, he was a special kind of friend, one who set the course of my research career.
Differences in the Ability of Anti-Insulin Antibody to Aggregate Monomeric Ferritin-Insulin Occupied Receptor Sites on Liver and Adipocyte Plasma Membranes
Authors: Lyen Kenneth R , Smith Robert M , Jarett Leonard
Published: September 2013

The effect of anti-insulin IgG (AlAb) on the ultrastructural organization of insulinreceptors on liver and adipocyte plasma membranes was studied using monomeric ferritin-insulin (Fm-I) as an electron microscopic marker for the insulin binding sites. Liver plasma membranes wereincubated at 0°C and 24°C with 250, 500, or 1000 μU monomeric Fm-I/ml, in the presence and absence of AlAb at a concentration that tripled 125I-insulin binding to livermembranes. Electron microscopy revealed that increasing concentrations of Fm-I increased the number of occupied receptors but had no effect on the distribution of occupied receptors into groups of various sizes on liver membranes. Over 60% of the receptors were observed as single molecules. At 24°C, AlAb produced a significant aggregation of occupied receptors, and only 30% of the receptors were found as single sites. At 0°C, AlAb hardly affected receptor distribution. On adipocyte membranes, however, about 60% of the occupied receptors normally occurred in groups of 2–6 molecules. Addition of AlAb to the adipocyte membranes, at concentrations just below that which inhibited 125I-insulin binding to these membranes, had no effect on the distribution pattern of Fm-I occupied receptors. Normal IgG had no effect on the Fm-I occupied receptors on either liver or adipocyte membranes. These data show that Fm-I occupied receptor sites on liver membranes at 24°C are mobile, and that insulin itself did not cause grouping of these receptors. AlAb-induced Fm-I aggregation probably occurred as the bivalent antibody cross-linked the occupied mobile receptors, since AlAb could not cause aggregation on liver membranes at 0°C, when the receptors were less mobile. The AlAb does not have a morphologically demonstrable effect on the already grouped receptor sites on adipocytes. Insulin receptors on liver and adipocyte membranes differ, not only in their organizational pattern, but also in their responses to AlAb, suggesting that unique differences exist between the membranes, the receptors in the membranes, or both.
Resting and Exercise Hyperemic Pulsatile Arterial Blood Flow in Insulin-dependent Diabetic Subjects
Authors: Cunningham L N , Labrie C , Soeldner J S , Gleason R E
Published: September 2013

Pulsatile arterial blood flow was studied in 20 normal (N), 20 short-term (STIDDM; mean: 5.17 yr), and 20 long-term insulin-dependent diabetic patients (LTIDDM; mean: 14.76 yr) between the ages of 18 and 30 yr with no clinically detectable peripheral vascular disease. Measurements were taken from waveforms obtained noninvasively using an electromagnetic flowmeter at rest and immediately after a 3-min isometric exercise challenge of the right leg. At rest, both groups of diabetics exhibited minute flow values similar to those in the normal group. This was achieved, however, by increased vasodilation in peripheral tissues as indicated by a difference in waveform configuration. Diabetic subjects showed a significantly smaller peak flow, a less steep ascending and descending slope, and a higher minute heart rate than normal controls. After 3 min of isometric exercise, the diabetic groups exhibited significantly less minute flow, flow/pulse, and a more vasodilated flow pattern similar to that recorded at rest. In addition, the LTIDDM groupshowed significantly less arterial elasticity than N or STIDDM groups as indicated by a shorter propagation time. These findings imply that apparent functional changes in pulsatile arterial blood flow occur early in the time course of diabetes and are independent of duration.
Characterization of the Receptor Binding Properties of Monoiodoinsulin Isomers and the Identification of Different Insulin Receptor Specificities in Hepatic and Extrahepatic Tissues
Authors: Podlecki D A , Frank B H , Kao M , Horikoshi H , Freidenberg G , Marshall S , Ciaraldi T , Olefsky J.M
Published: September 2013

Insulin was iodinated with 125I using a modification of the lactoperoxidase method. This iodinated insulin mixture was then separated by HPLC, allowing studies to be performed on the pure monoiodoinsulin isomers labeled at the tyrosyl residuesat A14, B16, or B26.

Binding studies in adipocytes revealed the relative affinity for these isomers to be 1.00:0.99:1.30 (A14, B16, and B26, respectively). The increase in binding affinity was shown to be due to a slower dissociation rate rather than an accelerated association rate. Receptor mediated internalization of the different iodinated isomers was assessed and these results showed that the isomers were internalized and degraded in proportion to their relative binding affinities. The biologic activity of the B26 isomer to stimulate adipocyte glucose oxidation was measured and showed that the B26 isomer had 30% greater biologic potency compared with native insulin, consistent with its 30% increase in binding potency. Thus, the B26 monoiodoinsulin isomer behaves as a superagonist.

Binding studies using extrahepatic tissue (i.e., adipocytes, cultured human fibroblasts, IM-9 lymphocytes, and human placental membranes) consistently revealed that the relative binding affinity for the B26 isomer was always greater than the A14 isomer (1.29, 2.11,1.22, and 1.35, respectively) while the binding affinity of the B16 isomer was generally comparable to the A14 isomer. In hepatic tissue (whole cell, liver plasma membranes, and partially purified, solubilized liver receptors) binding studies illustrated that the B26 isomer consistently had a lower relative affinity when compared to the A14 isomer (0.75 : 0.69 : 0.77).

In conclusion, (1) the B26 monoiodoinsulin isomer exhibits higher binding affinity and greater biologic activity in adipocytes than native insulin, and (2) the B26 monoiodoinsulin isomer has a higher binding affinity in extrahepatic tissues but exhibits a lower binding affinity in hepatic tissues when compared with the A14 monoiodoinsulin isomer, indicating a striking difference in functional specificity between the hepatic and the extrahepatic insulin receptor.
Regulation of Triacylglycerol Lipolysis in the Perfused Hearts of Normal and Diabetic Rats
Authors: Murthy Veeraraghavan K , Bauman Mitchel D , Shipp Joseph C
Published: September 2013

Effects of insulin, palmitate, and acetate on lipolysis of endogenous triacylglycerols were studied in the isolated perfused hearts of control and ketotic diabetic rats. In the absence of fatty acid, the rate and net amount of lipolysis were increased in diabetic hearts. After 120 min of perfusion, the residual triacylglycerol content was similar in both hearts, suggesting that the accumulation of triacylglycerols in the heart of ketotic diabetic rats was due to the expansion of the mobiliza-ble fraction. Insulin, in physiologic concentrations, inhibited triacylglycerol lipolysis in control hearts, provided glucose was present in the perfusate. In diabetic hearts perfused with glucose, insulin up to 500 μU/ml did not inhibit triacylglycerol lipolysis.

A physiologic concentration of palmitate (0.5 mM) completely inhibited triacylglycerol lipolysis in both control and diabetic hearts. Palmitate inhibition of lipolysis in both control and diabetic hearts could be observed in absence of perfusate glucose. Heart intra-cellular lipoprotein lipase activity was determined to check whether the inhibitory actions of insulin and palmitate could be explained by reduction in this lipase activity. Perfusion of control hearts with insulin and glucose had no effect on the lipase activity. Lipase activity of the control heart was unaffected and that of the diabetic heart was inhibited after perfusion with palmitate. The results of this study show (1) insulin in physiologic concentrations inhibits heart triacylglycerol lipolysis probably through increased glucose transport and glycolysis; (2) inclusion of palmitate in the perfusate results in inhibition of triacylglycerol lipolysis in both control and diabetic hearts and in the case of the diabetic heart, the palmitate effect is due to the inhibition of the intracellular lipoprotein lipase activity; and (3) an increased synthesis of triacylglycerol coupled with an inhibition of its lipolysis by free fatty acids and ketone bodies may explain the accumulation of triacylglycerols in the hearts of diabetic rats.
Normalization of Six Key Hepatic Enzymes After Fetal Pancreas Transplantation in Diabetic Rats
Authors: Makoff R K , Brown J , Mullen Y , Clark W R
Published: September 2013

In diabetic rats transplanted with fetal pancreata we measured the activities of six important enzymes to assess the return of liver metabolism to normal. Comparison was made among the responses of transplanted rats with and without renal-portal vein shunts and of those not transplanted and injected with insulin in varying doses. Insulin supply was not limited since three or four fetal pancreata were first grown in normal rats before transfer into the diabetic animals.

Transplantation normalized blood and urine glucose and the rate of disappearance of intravenous glucose. Glucokinase and pyruvate kinase activities in liver rose toward normal at 7 days after transplantation and reached normal levels at 30 and 90 days. The response of the other four enzymes, glucose-6-phosphate dehy-drogenase, citric lyase, fructose-1,6-bisphosphatase, and glucose-6-phosphatase, was more rapidly restored to normal at 7 days and remained normal at 30 and 90 days. No difference was observed in the enzyme activities of transplanted-shunted rats to nonshunted animals.

Glucokinase activity was restored to normal after 1 wk of daily injections of 1 U of PZI; pyruvate kinase restoration required 3 U/day. Glucose-6-phosphate dehydrogenase and citric lyase required 2 U/day to be restored to normal; 3 U daily resulted in temporary supernormal activities. The gluconeogenic enzymes, fructose-1,6-bisphosphatase and glucose-6-phospha-tase, were only partially suppressed toward normal by insulin even with 3 U daily for 3 wk.

These findings indicate that pancreas transplantation is a more effective regulator of liver metabolism in diabetes than insulin injections.
Defective Oxidative Metabolism of Heart Mitochondria from Genetically Diabetic Mice
Authors: Kuo Tuan H , Moore Kathleen H , Giacomelli Filiberto , Wiener Joseph
Published: September 2013

Long chain saturated β-hydroxy fatty acid content and oxidative metabolism were studied in hearts of diabetic mice (C57BL/KsJ db/db) with a progressive cardiomy-opathy at intervals of 7, 10, 16, and 26 wk of age. Total β-hydroxy fatty acid (BHFA) content increases progressively with age in diabetic hearts with a mean value of 143.5 nmoi/g dry wt as compared with a mean of 59.6 nmol/g dry wt in control hearts. There was also a redistribution of BHFA in myocardium of diabetic mice when compared with controls, with a relative decrease in β-hydroxymyristate and an increase of β-hy-droxypalmitate. Oxidative phosphorylation studies using isolated mitochondria from diabetic mice demonstrated depressed state 3 oxidation rates with both palmityl carnitine and pyruvate as substrates. While mitochondrial NADH-oxidase activity was not statistically different from that of controls, there was a significant decrease in mitochondrial total NAD + NADH content in diabetic hearts. In addition, treatment of my-ocardfal tissue with lanthanum demonstrated an abnormal permeability of sarcolemmal, intercalated disc as well as mitochondrial membranes in myocytes of diabetic mice. The data indicate that deficiencies in total NAD + NADH content can account for the depressed state 3 oxidation of palmitylcarnitine and pyruvate in diabetic mice that in turn may explain the abnormal accumulation of BFHA. The latter could play a role in altering the permeability of cardiac cell membranes.
Biphasic Modulation of K+ Permeability in Pancreatic Islets During Acute Stimulation with Glucose
Authors: Sehlin Janove , Freinkel Norbert
Published: September 2013

The regulation of 86Rb+ efflux (marker of K+ permeability) during acute secretagogic stimulation with glucose was studied with cultured as well as freshly isolated pancreatic islets from rats and freshly isolated islets from mice. A perifusion system with minimal dead-space and “flow-through” characteristics conducive to abrupt, steep increases in ambient glucose was combined with multiple samplings of effluent to achieve high temporal resolution. Under these conditions, acute increases in perifusate glucose concentration from 4 to 16.7 mM or from 1 to 27.8 mM effected a biphasic change of the 86Rb+ fractional efflux rate. A rapid reduction of 86Rb+ efflux was interrupted by an evanescent increase in 86Rb+ outflow, which appeared to be temporally coincident with the initiation of the first phase of stimulated insulin release. It is suggested that the glucose-induced biphasic oscillations in K+ permeability may contribute to the well-known initial biphasic changes in β-cell membrane potential and insulin release during the inception of β-cell stimulus secretion coupling.
The Effect of Growth Hormone on Splanchnic Glucose and Substrate Metabolism Following Oral Glucose Loading in Healthy Man
Authors: Bratusch-Marrain Paul R , Gasić Slobodan , Waldhäusl Werner K , Nowotny Peter
Published: September 2013

Elevated plasma concentrations of growth hormone impair glucose tolerance and insulin sensitivity of peripheral tissues. To study the effect of short-term exposure to growth hormone concentrations elevated into the upper physiologic range (7–10 ng/ml) on splanchnic carbohydrate metabolism, both splanchnic glucose output (SGO) and substrate exchange after ingestion of a 75-g glucose load were determined by means of the liver vein catheter technique in six healthy volunteers after growth hormone administration. Growth hormone was infused at a rate of 2 μg/ kg · h starting 120 min before and continuing for 150 min following the glucose load. Control studies without growth hormone administration were performed in seven subjects. SGO was 104 ± 10 (SEM) mg/min in the postabsorptive state and increased to 43.4 ± 2.2 g during the 150-min period following glucose ingestion. Growth hormone infusion did not alter basal SGO (130 ± 14 mg/min), nor the splanchnic exchange of lactate, pyruvate, and free fatty acids, whereas basal production of β-OH-butyrate was increased twofold; following glucose ingestion a higher proportion of the given glucose load escaped the splanchnic bed after growth hormone exposure (66.9 ± 6.8 g/150 min; P < 0.005). The insulin production rate (basal 14 ± 2 mU/min; following oral glucose 7.0 ± 0.8 U/150 min) as calculated from C-peptide release from the splanchnic area was unaltered by growth hormone exposure in the basal state (14 ± 3 mU/min), but augmented after glucose ingestion (14.8 ± 1.5 U/150 min). The higher rate of glucose release by the splanchnic bed after glucose ingestion despite a more marked hyperglycemia and increased insulin secretion suggests that acute elevations of plasma growth hormone within the physiologic range, besides causing insulin resistance of peripheral tissues, affect glucose disposal of splanchnic tissues as well.
Insulin in the Rat Fetus: A New Form of Circulating Insulin
Authors: Watts Colin , Gain Kevin R
Published: September 2013

The apparent hyperinsulinemia seen in the fetal rat in late gestation (day 20) is shown to be due to an immunoreactive insulin (IRI) species peculiar to the fetal rat, and not monomeric insulin. This species disappears within 12 h after birth and only insulin as found in the adult can then be detected. The fetal serum IRI has a molecular weight of approximately 11,000 and differs from monomeric insulin and proinsulin in its behavior on gel filtration and anion-exchange chromatography. The activity of fetal serum in an insulin bioassay based on the incorporation of glucose into lipid was half that expected on the basis of the serum IRI concentration. The addition of fetal serum to purified insulin or proinsulin caused an increase in the apparent molecular weight of both hormones by 2000. In the case of insulin, the generated species of mol wt 8000 was clearly differentiated from the endogenous species (11,000) on Sephadex G50. The fetal pancreas at this age was found to contain predominantly monomeric insulin that had full biologic activity in the insulin bioassay. It is postulated that the fetal pancreas secretes monomeric insulin, which is then modified in the circulation to a complex with reduced biologic activity. This process appears to have two steps in that in vitro fetal serum increases the apparent molecular weight of added insulin to 8000, whereas in vivo an additional component of approximately 3000 mol wt is involved to produce the endogenous insulin complex.
Degradation of Insulin by Isolated Mouse Pancreatic Acini: Evidence for Cell Surface Protease Activity
Authors: Goldfine Ira D , Williams John A , Bailey Anne C , Wong K Y , Iwamoto Yasuhiko , Yokono Koichi , Baba Shigeaki , Roth Richard A
Published: September 2013

In the present study, we have used isolated mouse pancreatic acini to investigate the relationship between 125I-insulin binding and its degradation in order to probe the nature and cellular localization of the degradative process. In these cells, the proteolysis of 125I-insulin was dependent on time and cell concentration, and was saturated by unlabeled insulin with a Km of 290 nM. Since this value was much higher than the Kd for insulin binding to its receptor (1.1 nM), the data indicated that 125I-insulin degradation by acini occurred primarily via nonreceptor mechanisms.

Several lines of evidence suggested that insulin was being degraded by the neutral thiol protease, insulin degrading enzyme (IDE). First, insulin degradation was inhibited by thiolreacting agents such as N-ethylmaleimide and p-chloromercuribenzoate. Second, the Km for degradation in acini was similar to the reported Km for IDE in other tissues. Third, the enzyme activity had a relative mol wt of approximately 130,000 by gel filtration, a value similar to that reported for purified IDE. Fourth, the degrading activity was removed with a specific antibody to IDE.

Other lines of evidence suggested that enzymes located on the cell surface played a role in insulin degradation by acini. First, the nonpenetrating sulfhydryl reacting agent 5,5′dithiobis-2-nitrobenzoic acid blocked 125I-insulin degradation. Second, a specific antibody to IDE identified the presence of the enzyme on the cell surface. Third, chloroquine, leupeptin and antipain, agents that inhibit lysosomal function, did not influence 125I-insulin degradation. Fourth, highly purified pancreatic plasma membranes degraded 125I-insulin.
Immunohistochemically Detectable Vitamin D-Dependent Calcium-Binding Protein Is Reduced in Cerebellum of Diabetic Subjects
Authors: Bruhlmann-Papazyan M , Garcia-Segura L M , Pizzolato G P , Norman A W , Orci L
Published: September 2013

The immunohistochemical distribution of the vitamin D-dependent calcium-binding protein (CaBP) was studied in the cerebellar vermis of 14 adult type (NIDDM) diabetic and 13 nondiabetic human subjects. In both groups, CaBP-immunoreactive material was found in Purkinje cells and in axons of the cortical white matter, but, when compared with controls, the positive structures had a significantly (P < 0.001) reduced volume density in 12 of the 14 diabetic subjects. The reduction of the volume density of CaBP-positive structures in diabetic subjects was not due to an overall reduced volume of the molecular layer or of the cortical white matter. These results complement previous reports of a reduction of CaBP levels in duodenum and kidney of diabetic subjects.
Possible Role of Endogenous Arachidonic Acid Metabolites in Stimulated Release of Insulin and Glucagon from the Isolated, Perfused Rat Pancreas
Authors: Walsh Mary F , Pek Sumer Belbez
Published: September 2013

Enhancement of arachidonic acid metabolism results in increased insulin secretion. To determine which pathways of arachidonic acid metabolism were involved in this stimulation, we studied the effects of various inhibitors of arachidonate metabolism on arginine-induced insulin and glucagon secretion in the isolated, perfused rat pancreas. The release of PGE2 from the pancreas was monitored to document the efficacy of the inhibitory drugs.

p-Bromophenacyl bromide, a phospholipase A2 inhibitor, diminished PGE2 release and significantly inhibited both the early and late phases of insulin and glucagon release in response to arginine. Flurbiprofen, a specific cyclooxygenase inhibitor, decreased the early phase of insulin release and inhibited both phases of arginine-stimulated glucagon secretion; these decreases were concurrent with a large inhibition of PGE2 release. Nordihydroguaiaretic acid, a li-poxygenase inhibitor, at a dose of 10-5 M did not affect PGE2 release, inhibited the early phase of insulin release, and did not modify glucagon secretion. The combination of flurbiprofen and nordihydroguaiaretic acid, although the most potent in inhibiting PGE2, lowered only the early phase of insulin and had no effect on glucagon secretion.

We conclude that: (1) endogenous cyclooxygenase-derived metabolites of arachidonic acid promote insulin and glucagon release, (2) endogenous lipoxygenase products preferentially stimulate insulin release, and (3) phospholipase A2 activity has an intrinsic modulatory effect on insulin and glucagon secretion.
Nicotinamide Does Not Protect Islet B-Cell Metabolism Against Alloxan Toxicity
Authors: Sandler S , Welsh M , Andersson A
Published: September 2013

Nicotinamide, a poly(ADP-ribose)synthetase inhibitor, protected NMRI mice against alloxan-induced hyperglycemie when given 10 min before, but not 10 min after, the injection of the drug. Pretreatment in vivo with nicotinamide induced hyperglycemia at the time of alloxan injection, and this could account for the protective action of nicotinamide against alloxan diabetes. Exposure of islets to alloxan (2 mM) in vitro caused a marked inhibition of both glucose-stimulated proinsulin biosynthesis and insulin release, and this was not affected by the action of nicotinamide. Alloxan-impaired islet glucose oxidation was partly restored by nicotinamide. The decreased islet content of NADH plus NAD, which was observed after alloxan treatment, could be prevented by nicotinamide. Glucose-stimulated islet oxygen uptake was abolished after treatment with alloxan, and nicotinamide had no protective effect in this process. Leucine (10 mM) plus glutamine (10 mM), however, were still able to evoke an islet respiratory response after alloxan exposure. Alloxan caused an immediate increase in the islet efflux of radiolabeled nucleotides, which was followed after about 5 min by a further increase. This latter increase of the radio efflux was inhibited by the addition of nicotinamide.

The inability of nicotinamide to prevent the alloxan-induced impairment of proinsulin biosynthesis, insulin release, and oxygen uptake, together with the failure of nicotinamide to prevent the development of diabetes when given after alloxan, does not support a current hypothesis that the major cytotoxic effect of alloxan is primarily due to DNA damage. The present data suggest that organelles other than the nuclei, e.g., the mitochondria or the plasma membrane, are the primary sites of B-cell injury by alloxan. It is also concluded that the B-cell-toxic action of alloxan is different from that Of StreptOZOCin.
The Onset of Liver Glycogen Synthesis in Fasted-Refed Rats: Effects of Streptozocin Diabetes and of Peripheral Insulin Replacement
Authors: Werve G Van De , Sestoft L , Folke M , Kristensen L Ø
Published: September 2013

The mechanism of liver glycogen synthesis after refeeding has been investigated in diabetic rats, diabetic insulin-treated rats, and in control rats fasted for 48 h. The accumulation of liver glycogen was the same in diabetic rats and in control rats after 2 h of feeding, but did not proceed any further in the diabetic group during the next 2 h. Insulin-treated diabetic rats synthesized five times more hepatic glycogen than the control rats after 1 h of refeeding, but the amount accumulated at the end of the refeeding period was the same. Feeding resulted in a transient activation of glycogen synthase in untreated as well as in treated diabetic rats. In control rats, however, glycogen synthase was already partially in the active form before access to food, and the onset of glycogen synthesis occurred without further activation of the enzyme. A transient inactivation of phosphorylase was observed in all groups during the meal, but was very slight in the untreated diabetic rats in which phosphorylase a values were already reduced before the access to food.

Peripheral glycemia was markedly increased upon refeeding in treated and untreated diabetic rats, but remained normal in control rats. Peripheral insulinemia was increased by feeding in the control rats and remained low in the diabetic rats and high in the insulin-treated diabetic rats.

The results indicate that, in normal controls in contrast to diabetic rats, synthase activation is not a prerequisite for the initiation of glycogen synthesis after a meal; phosphorylase inactivation may be of major importance in normal controls, but also appears to play a role in the diabetic animals. The changes are triggered by glucose as well as insulin.
Glucagon Binding to Liver Membranes of Mt-T-W15 Tumor-bearing and Hypophysectomized Rats: A Possible Role for Insulin and Growth Hormone
Authors: Walsh Mary F , Dunbar Joseph C
Published: September 2013

To assess the effect of the endocrine environment and, more specifically, of growth hormone and insulin on glucagon receptors, we studied 125I-glucagon binding to liver membranes in five groups of rats: (1) controls, (2) streptozocin (STZ)-treated, (3) tumor-bearing (growth hormone-producing, Mt-T-W15), (4) STZ-treated tumor-bearing, and (5) hypophysectomized rats. Glucagon binding was decreased in tumor-bearing, hypophysectomized, and STZ-treated rats. Basal and glucagon-stimulated cAMP levels were determined in isolated hepatocytes. The basal cAMP levels were increased in STZ-treated, tumor-bearing, STZ-treated tumor-bearing, and hypophysectomized animals. All groups responded and produced more cAMP in response to glucagon stimulation, although the maximal response was greater in STZ-treated, tumor-bearing STZ-treated, and hypophysectomized groups than in the controls. Our data confirm that hyperglucagonemia downregulates the hepatic glucagon receptors and suggest that insulin and growth hormone may play a stimulatory role in their regulation. The results also suggest that the downregulation of glucagon receptors is not directly correlated to the basal or glucagon-stimulated CAMP levels.
Effects of Monensin on Conversion of Proinsulin to Insulin and Secretion of Newly Synthesized Insulin in Isolated Rat Islets
Authors: Gold Gerald , Pou José , Nowlain Robert M , Grodsky Gerold M
Published: September 2013

When isolated rat islets were incubated with 10−10–10−6 M monensin, a sodium and proton ionophore, glucose-stimulated insulin release was inhibited in a concentration- and time-dependent manner. After removal of monensin, inhibition of insulin secretion persisted during stimulation with a variety of secretagogues, including 5 mM glucose plus 15 mM arginine, 20 mM glucose, and 20 mM glucose plus 1 mM 3-isobutyl-1-methylxanthine. Within the same low range of monensin concentrations, proteolytic conversion of newly synthesized proinsulin to insulin was also blocked. At each concentration, prohormone-to-hormone conversion was inhibited to almost the same extent as inhibition of insulin secretion. Therefore, both processes may have equal or common dependency on a subcellular ionic gradient. Although monensin decreased total insulin secretion, the glucose-regulated marking process was unaffected. Regardless of the monensin concentration or the overall rate of insulin secretion, the percentage of secreted newly synthesized versus older insulin remained the same, and the threefold differences in the fractional secretory rates of newly synthesized versus total insulin also remained the same. Thus, rather than specifically blocking protein traffic through the Golgi apparatus of the beta cell, monensin probably first inhibited insulin secretion by disrupting proton gradients in secretory vesicles and, thereby, also inhibited other processes occurring within this organelle.