bluuebunny/crossref_metadata_2025 huggingface dataset:
Chronic Hyperinsulinemia in the Fetal Rhesus Monkey: Effects on Fetal Growth and Composition
Authors: Susa John B , McCormick Kenneth L , Widness John A , Singer Don B , Oh William , Admsons Karlis , Schwartz Robert
Published: September 2013

The delivery of 19 U of insulin a day for 21 days to the rhesus monkey fetus, coupled with the permeability properties of the placenta, has made it possible to produce fetal hyperinsulinemia in the presence of euglycemia. Fetal plasma insulin concentrations of 3525 μU/ml were attained with no apparent effects on the mother. In fetal macrosomia, a 34% increase in body weight was observed above the expected weight for gestational age (466 vs. 348 g). Relative organomegaly, matched for gestational age of fetal weight, was seen in the hyperinsulinemic fetuses with enlarged livers, placentas, hearts, and spleens. Liver composition in the fetuses was only slightly affected by hyperinsulinemia. Glycogen concentration was elevated, but not sufficiently, to explain the relative hepatomegaly produced. The lipid, protein, DNA, and RNA concentrations were not affected by hyperinsulinemia. Based on the similar DNA concentrations and protein/DNA ratios observed in hyperinsulinemic and control groups, the hepatomegaly appears to be the result of insulin-stimulated hyperplasia and not of hypertrophy. In the presence of normal plasma concentrations of growth substrates, insulin in the subhuman fetus has been shown to be a growth-promoting hormone that has specific growth-stimulating effects.
Ionic Basis of Phenytoin Sodium Inhibition of Insulin Secretion in Pancreatic Islets
Authors: Pace Caroline S , Livingston Elizabeth
Published: September 2013

The possibility that phenytoin sodium has a dual role in blocking Ca and Na channels in the pancreatic β-cell membrane was studied to clarify the mechanism of action of the drug in inhibiting insulin release. Glucose and veratridine were used to activate the Ca and Na channels, respectively. The increase in insulin release induced by 16.7 mM glucose or 200 μM veratridine was inhibited 77% and 60%, respectively, by 100 μM phenytoin, whereas the increase in the rate of glycolysis was inhibited 74% and 100%, respectively. An increase in extracellular Ca from 2.5 to 5.0 mM attenuated the inhibitory effect of phenytoin oh both the metabolic and secretory responses to glucose and veratridine. This Ca-dependent reversal was greater for glucose than for veratridine. Furthermore, 1,0 mM ouabain partially reversed the inhibitory effect of phenytoin on the secretory responses to both stimulants. This may be attributed to the indirect effect of ouabain in increasing intracellular Ca. However, ouabain together with phenytoin almost completely blocked glycolytic flux measured in the presence of either glucose or veratridine. These results suggest that phenytoin may block the stimulation of passive Na and/or Ca transport in the β-cell membrane.
Renin-Angiotensin System in Phlorhizin Compared with Alloxan Diabetes in the Rat
Authors: Christlieb A Richard , Long Rosemary
Published: September 2013

In alloxan-treated diabetic rats, plasma renin activity (PRA) is decreased. One possible mechanism that may explain the decreased PRA is an increased delivery of sodium to the macula densa produced by the glucose osmotic diuresis, resulting in decreased renin release. To evaluate this possible mechanism, rats with phlorhizin diabetes, which produces a glucose osmotic diuresis without hyperglycemia, were studied and ompared with rats with alloxan-induced diabetes. Whereas phlorhizin-treated rats had low blood glucose and alloxan-treated rats had elevated glucose, the glucose osmotic diuresis was similar in the two groups. PRA and plasma renin concentration (PRC) were significantly increased in the phlorhizin group. In the alloxan group, PRA was decreased and angiotensin II sensitivity increased, both significantly. Plasma renin substrate (PRS) remained adequate in each group. These results suggest that the decreased PRA in alloxan-induced diabetes is due neither to factors associated with the glucose osmotic diuresis including changes in renal tubular sodium nor to decreased PRS.
Ultrasonography in the Detection of Residual Urine
Authors: Piters Kenneth , Lapin Stuart , Bessman Alice N
Published: September 2013

Eleven patients with distended urinary bladders (10 diabetics with autonomic neuropathy and one patient with prostatic hypertrophy) were examined by ultrasonography in an attempt to define the volumes detectable by this technique. Incremental volumes of saline were instilled through indwelling Foley catheters with ultrasonographic examination at each volume. The ultrasonograms obtained were presented to two radiologists trained in ultrasonography, who were asked to identify the presence of residual urine. There was 97% certainty of recognizing the presence of 100-cc residual volumes and 100% certainty of recognizing 150-cc residual volumes. It is concluded that residual urine volumes greater than 100 cc are clinically detectable by ultrasonography, a noninvasive alternative for diagnosing this common diabetic condition.
The Effect of Age on Insulin-degrading Activity in Rat Tissue
Authors: Runyan K , Duckworth W C , Kitabchi A E , Huff G
Published: September 2013

Insulin-degrading activity was measured in the 100,000g supernatant fraction of muscle, liver, and kidney from rats of varying ages. Young animals (four weeks old) had the highest activity in all three tissues. By seven weeks of age the activity in both muscle and liver had decreased significantly as compared with four-week-old animals. A slight but nonsignificant decrease occurred in kidney. In animals over one year of age the insulin-degrading activity in all three tissues was significantly less than the activities at either four or seven weeks. In contrast the effect of age on degradation of albumin and parathormone was much less marked.
Reduced Monoamine Oxidase Activity in Blood Platelets from Insulin-dependent Diabetic Subjects
Authors: Mosnaim Aron D , Wolf Marion E , Huprikar Shankar , Singh Sant P , Zeller E Albert
Published: September 2013

Insulin-dependent diabetics show a significant reduction in blood platelets' monoamine oxidase (MAO) activity when compared with age-matched and sex-matched controls. This does not appear to be the case for diabetics either on diet control or receiving oral hypoglycemic drugs.

The important role of this enzyme in the regulation of the circulating levels of a number of monoamines known to have an inhibitory effect on insulin secretion may indicate the possible inclusion of the MAO/monoamine system in the pathophysiology of diabetes. The possibility of using platelets' MAO activity as a biologic marker for a subgroup of insulin-dependent diabetics is also discussed.
Synthesis and Accumulation of Triglycerides in Liver of Diabetic Rats: Effects of Insulin Treatment
Authors: Murthy Veeraraghavan K , Shipp Joseph C
Published: September 2013

The relationship between the severity of diabetes and the hepatic accumulation of triglycerides in the streptozotocin-diabetic rat was studied. A consistent increase in hepatic triglyceride content was observed only in the ketotic diabetic state. Insulin treatment of the ketotic diabetic rat resulted in the reversal of increased plasma concentrations of glucose, free fatty acids, β-hydroxybutyric acid, and triglycerides to those observed in nondiabetic controls. Insulin treatment of the ketotic rats, however, did not completely eliminate the accumulation of the hepatic triglycerides.

To determine whether increased triglyceride synthesis was a contributory factor to the hepatic triglyceride accumulation, triglyceride synthesis was studied in vitro using nuclear-free homogenates of liver. Triglyceride synthesis increased in ketotic diabetes. Insulin treatment of the ketotic rats partially reversed the increased diglyceride synthesis observed in ketosis but had no effect on the accelerated triglyceride synthesis. The effects of ketotic diabetes and insulin treatment of the diabetic rat on the activities of two key enzymes of the triglyceride biosynthetic pathway were examined next. The maximal velocities (Vmax) of liver-soluble phosphatidate phosphohydrolase and of microsomal diglyceride acyltransferase increased in ketosis. Insulin treatment of the diabetic rats reverted the activity of these two enzymes to control values.

The results of our study demonstrate that hepatic triglyceride synthesis in ketotic rats accelerates at a time when hepatic triglyceride content increases, the increased synthesis of triglycerides was associated with increases in the activities of two key enzymes of triglyceride biosynthesis. There was a correlation between hepatic triglyceride content and triglyceride synthesis in control, ketotic diabetic, and insulin-treatedrats. These observations suggest a physiologic role for the observed changes in the accumulation of hepatic triglycerides in ketotic diabetes.
Variation in Thickness of the Capillary Basement Membrane in Single Muscles of Diabetic Subjects
Authors: Peterson Gregory E , Forsham Peter H
Published: September 2013

Thickness of skeletal muscle's capillary basement membrane was measured in gastrocnemius and quadriceps femoris muscles of diabetic subjects. At least two different sites of each muscle studied were biopsied. Comparison of these biopsy samples revealed a marked and significant variation in thickness of the capillary basement membrane at different sites within the same muscle. In one subject so studied, this variation was found regardless of the method of fixation or the method of measurement of capillary basement membrane thickness used. Variation was present in diabetics suffering from long-term complications of the disease as well as in diabetics free of such complications. A reevaluation of the biologic significance of basement membrane thickness measurements, as presently applied, is indicated in the light of the variability of base-line values.
High Concentration of Gamma-Aminobutyric Acid in Pancreatic Beta Cells
Authors: Taniguchi Hiroshi , Okada Yasuhiro , Seguchi Harumichi , Shimada Chicaco , Seki Michio , Tsutou Akimitsu , Baba Shigeaki
Published: September 2013

The γ-aminobutyric acid (GABA) concentration of pancreatic islets in rats treated with streptozotocin (STZ) and of human insulinoma tissue was studied. Seven hours after the administration of 65 mg/kg body weight of STZ, a distinct increase in serum insulin concentration and at the same time a decrease in blood glucose level were seen. Twenty-four hours after the injection of STZ, however, the level of serum insulin decreased much, whereas that of blood glucose increased considerably. On the other hand, the GABA concentration of the islet was reduced dramatically to about one-tenth the control level after both 7 and 24 h. The histologic investigations of the islets revealed the destruction of B cells but no changes in A and D cells 7 and 24 h after the treatment of STZ. Nerve fibers and nerve endings in the islets were preserved intact all through the study. The GABA and insulin contents of the two cases of human insulinoma were determined. One insulinoma, which was compactly occupied with B cells according to its histologic features, contained a high concentration of GABA. The other tumor, having a rather sparse distribution of B cells in it as compared with the former case, possessed a lower concentration of GABA, but it was still high compared with that of its surrounding tissues. The present observations indicate that a large amount of GABA is available in the B cells of the pancreaticislets.
Gastric and Pancreatic Release of Somatostatin-like Immunoreactivity During the Gastric Phase of a Meal: Effects of Truncal Vagotomy and Atropine in the Anesthetized Dog
Authors: Schusdziarra V , Rouiller D , Harris V , Unger R H
Published: September 2013

The postprandial release of gastric and pancreatic somatostatin-like immunoreactivity (SLI) was examined in anesthetized dogs during the gastric phase of a meal, and the role of vagal and atropine-sensitive mechanisms in the responses was assessed. The intragastric instillation of liver extract at pH 7 elicited a significant rise in antral vein SLI (∼300 pg/ml) and gastrin concentration. After truncal vagotomy, both baseline and postprandial antral vein SLI and gastrin concentration increased significantly compared with the control group. The infusion of atropine (100 μg/kg/h) abolished the postprandial rise in antral vein SLI but not in gastrin. The liver meal at pH 2 elicited a sustained sixfold greater rise of antral SLI (∼2000 pg/ml) than that at pH 7, while gastrin concentrations did not rise significantly. The latter antral SLI response was not influenced by truncal vagotomy, but atropine infusion reduced it by about 50%. In response to the meal at pH 7, fundic vein SLI concentrations rose by about 300 pg/ml. The rise was augmented slightly by truncal vagotomy but was abolished completely by atropine infusion. In response to the meal at pH 2, fundic SLI decreased sharply below baseline levels. The response was not altered significantly by vagotomy, but was reversed completely by atropine infusion, during which fundic vein SLI concentrations rose significantly. Pancreatic vein SLI concentrations rose by about 350 pg/ml in response to the gastric meal at pH 7. That rise was not altered significantly by vagotomy but was abolished by atropine infusion. In response to the meal at pH 2, pancreatic SLI concentrations rose by about 1000 pg/ml above baseline, significantly greater than the response to the meal at pH 7. The pancreatic vein SLI response to the meal at pH 2 was not altered by vagotomy. It was reduced considerably by atropine infusion. It is concluded that SLI is released from the antrum, fundus, and pancreas during the gastric phase of a meal and that these responses are modified by acidification of the intragastric contents and by truncal vagotomy and atropine infusion. The greatly augmented release of antral SLI in response to the acidified meal raises the possibility of a role for somatostatin in acid-induced suppression of gastrin release.
Time Dependence of the Glycogenic Effect of Insulin in Cultured Fetal Hepatocytes
Authors: Plas C , Menuelle P , Moncany M L J , Fulchignoni-Lataud M C
Published: September 2013

Primary cultures of rat fetal hepatocytes were used to study the effect of insulin on glycogenesis during 24 h. Measurements of both glycogen content and 14C-glucose incorporation into glycogen gave similar results. A clear stimulatory insulin effect was observed within one hour, reaching a maximum after three hours. However, from 4 to 12 h, the effect declined strongly and then disappeared. Thereafter, it reappeared but to a lesser extent. These striking variations were not caused by a lack of active insulin in the medium or by any amplification of time variations in hepatocyte basal glycogenic capacity. They were induced by the hormone itself as a function of time of insulin presence at the hepatocyte level. Therefore, the effect of insulin is time dependent.

The transient cessation in the stimulatory effect of insulin on net glycogenesis limits the total amount of glycogen stored and permits a more progressive glycogen accumulation. It is not yet clear whether this cessation is a result of a decrease in the insulin-stimulated glycogen synthesis, or an increase of glycogen breakdown, or both; but we showed that it does not require prior maximal stimulation of glycogenesis. After four hours of incubation with varying doses of insulin, a dosedependent decrease in the insulin glycogenic effect of a second maximal dose of hormone was observed, whereas glucagon-induced glycogenolysis was unchanged. The characteristics of the time dependence of the insulin effect suggest that it may play a regulatory role in fetal hepatocytes.
Neural Release of Glucagon Is Inhibited by Hyperglycemia and Enhanced by Phentolamine
Authors: Miller Ralph E , Horton Edward S
Published: September 2013

Using an innervated, cross-perfused, canine pancreasstomach–duodenum preparation, direct neural effects on the immunoreactive glucagon secretion rate (GSR) were separated from bloodborne influences. Both splanchnic nerves were cut above the diaphragm and stimulated simultaneously for three separate 10-min-long periods, twice before and once during a pancreatic arterial phentolamine infusion. The extent of the decrease in GSR that occurred after nerve section was inversely correlated with the arterial plasma glucose concentration at the time of section. Splanchnic nerve stimulation caused a significant increase in GSR. Similar stimulation during a pancreatic arterial phentolamine infusion caused a significantly greater increase. Rapid infusion of glucose near the end of the experiment caused a significant decrease in GSR, demonstrating the responsiveness of the preparation. These data were collected in conjunction with a study of neural influences on insulin secretion rates.2 It can be concluded that the central nervous system can alter the secretion rate of glucagon by direct neural means. Alpha-adrenergic blockade, in the presence of splanchnic nerve stimulation, enhances GSR.
Insulin-degrading Activity of Plasma Membranes from Rat Skeletal Muscle: Its Isolation, Characterization, and Biologic Significance
Authors: Yokono Koichi , Imamura Yoshimichi , Sakai Hideyo , Baba Shigeaki
Published: September 2013

We isolated the plasma membrane from rat skeletal muscle without using drastic procedures such as extraction by salts or other agents. As a result of the purity of our preparations, evaluated by enzymatic markers, lipid composition, 125l-insulin specific binding, and morphologic examinations, we were able to use the plasma membrane to study insulin degradation.

Isolated plasma membranes were capable of degrading insulin, but 95% of total degrading activity was found in the cytosol fraction. The membranes proteolytically degraded the hormone with a high degree of specificity and a pH optimum of 7.0. The extent of degradation depended on time, temperature, and protein concentration. The apparent Km for insulin was 1.7 × 10−7 M. N-ethylmaleimide (NEM) and p-chloromercuribenzoate (PCMB) markedly inhibited insulin degradation by membranes, whereas glutathione (GSH) and dithíothreitol (DTT) were stimulatory. These characteristics of insulin-degrading activity in the plasma membrane (membrane-IDE) were similar to those of the partially purified insulin-degrading enzyme from the cytosol fraction (cytosol-IDE).

To clarify the biologic significance of IDE, we examined the cytosol- and membrane-IDE activities under various conditions in rats grouped as follows: fed, fasted for 1 day, fasted for 3 days, refed, refed + actinomycin D, diabetic, and hyperinsulinemic (insulinoma). A positive correlation with a high coefficient (r = 0.674, P < 0.001) was demonstrated between cytosol-IDE and the plasma insulin concentration but not between membrane-IDE and insulin levels. While actinomycin D had no effect on membrane-IDE levels, it abolished the insulin-mediated rise in cytosol-IDE activity, suggesting that the latter was dependent on RNA synthesis.
One Month of Sustained Release of Insulin from a Polymer Implant
Authors: Creque Halimena M , Langer Robert , Folkman Judah
Published: September 2013

Rats made diabetic with streptozotocin received a single subcutaneous implant of an insulin polymer pellet that released insulin continuously at approximately 2 U/day. Continuous normoglycemia was achieved for up to 1 mo. Mean glucose level for treated animals was 113 mg/dl as compared with 398 mg/dl for untreated diabetic controls. Diurnal blood glucose values for treated animals ranged from 71 to 116 mg/dl. Polyuria and glycosuria were corrected by the presence of the insulin + polymer. Treated animals gained weight normally and reached a mean weight of 350 g, whereas untreated control animals lost weight, to a mean of 150 g. When insulin + polymer preparations were periodically implanted and removed at 7-day intervals, normoglycemia was only associated with the presence of implants.
Perifusion and Culture of Human Fetal Pancreas
Authors: Maitland Janice E , Parry David G , Turtle John R
Published: September 2013

Human fetal pancreas has been obtained, after the therapeutic termination of pregnancy with prostaglandin F2α. Pancreatic explants were studied in a perifusion system and maintained in organ culture for up to 3 wk. Insulin biosynthesis was stimulated by glucose; however, the incorporation of 3H-L-leucine into proinsulin was surprisingly high (52% after 3 h) vs. insulin (48%), which suggests a possible block in the conversion of proinsulin to insulin. Insulin secretion was stimulated during perifusion with 19.3 mM glucose (1.6× prestimulation level), 1.5 μM glucagon (2.4), 5 mM leucine (2.4), 10 mM arginine (2.7), and 10 mM theophylline (10.0).

Insulin biosynthesis and secretion were maintained in organ culture. After 12 days there was a 3.4-fold increase in insulin secretion in the presence of 22 mM glucose compared with 5.5 mM glucose, and the explant insulin content rose in the presence of high glucose levels by 89%. The acute insulin secretory response to 10 mM theophylline was maintained after culture.

These studies suggest that human fetal pancreas should be considered as a potential source of donor tissue for pancreas transplantation in human diabetes.
Erythrocyte Insulin Receptors in Non-insulin-dependent Diabetes Mellitus
Authors: DePirro Roberto , Fusco Angelo , Lauro Renato , Testa Ivano , Ferreti Fabrizio , Demartinis Carlo
Published: September 2013

Insulin binding was studied on circulating erythrocytes isolated from 12 normal and 12 untreated, adult, nonobese, nonketotic, non-insulin-dependent diabetic subjects. Insulin binding in normal subjects was higher than in diabetics (P < 0.01); binding variation was caused mainly by a reduction in insulin receptor concentration. Insulin binding was inversely correlated with fasting serum insulin levels (R = 0.49; P < 0.01). The close agreement between the present data and previous data on other cell populations suggests that isolated erythrocytes may be a useful tool in clinical investigations on the human insulin receptor.
Decreased Insulin Binding and Degradation Associated with Depressed Intracellular ATP Content
Authors: Draznin Boris , Solomons Clive C , Emler Carol A , Schalch Don S , Sussman Karl E
Published: September 2013

The effect of metabolic inhibitors, 2,4-dinitrophenol (DNP) and NaF, on insulin binding and degradation has been studied in cultured Buffalo rat liver (BRL) cells. In control studies, 1.8 fmol of 125I-insulin binds to 1.2 × 106 cells, possessing approximately 40,000 receptor sites per cell with binding affinity of 5.52 × 10−8 M. When the cells were preincubated with increasing concentrations of either DNP or NaF, a dose- and time-dependent decrease in both insulin binding and degradation was observed. The total amount of 125I-insulin bound to BRL cells preincubated with metabolic inhibitors was reduced to 1.2 fmol per 1.2 × 106 cells. The point of ½ B max was achieved in the presence of 50 ng/ml of native insulin, 1.7 times that of the control level. The number of receptor sites was unaffected by either DNP or NaF, but an average affinity profile revealed a decrease in the affinity of the ATP-depleted cells for insulin (KD : 7.31 × 10 −8 M and 7.06 × 10−8 M in DNP- and NaF-treated cells, respectively). The decrease in insulin binding and degradation following the exposure of the BRL cells to the metabolic inhibitors was associated with a 20% reduction in intracellular ATP and adenylate energy charge. DNP and NaF did not affect the equilibrium constant for the myokinase catalyzed reaction and the intracellular concentration of hypoxanthine was stable, confirming the integrity of the cells during the experiments.

It is suggested that ATP levels must remain intact to maintain normal insulin receptor affinity. Furthermore, the rate of insulin degradation by ATP-depleted cells is slower than that of intact cells. It is conceivable that the depression of insulin degradation by partially ATP-depleted cells results from either diminished binding or decreased endocytosis and lysosomal activity, all of which appear to be energy dependent.
HLA-DRw Antigens in Mexican-American and Black-American Diabetic Patients
Authors: Zeidler Adina , Loon Jimmy , Frasier S Douglas , Kumar Dinesh , Penny Robert , Terasaki Paul
Published: September 2013

HLA-A, -B, and -C antigens were studied in 67 Mexican-American and 38 black-American diabetic patients who had the onset of their disease before age 31 yr. Control populations consisted of 322 Mexican-American and 367 black-American subjects for HLA-A, -B, and -C antigens. In addition, HLA-DRw antigens were studied in 60 Mexican-American and 34 black-American diabetic patients. Control populations for HLA-DRw antigens consisted of 189 Mexican-American and 145 black-American subjects.

We found that juvenile-onset-diabetic patients of Mexican-American origin who had the onset of their disease before age 19 demonstrated a significant increase in HLA-DRw4. HLA-DRw4 was also significantly increased in black-American patients with juvenile-onset diabetes mellitus. HLA-DRw2 was not detected in any patient with juvenile-onset diabetes in either ethnic group. A significant association was found between HLA-B18 and HLA-DRw3 in Mexican-American juvenile-diabetic patients.

These findings, which are comparable to those in similar Caucasian patients, provide additional information to support the hypothesis that HLA-DRw antigens play a major role in determining the susceptibility to juvenile-onset diabetes mellitus.
Nonenzymatically Glycosylated Serum Protein: Spurious Elevation Due to Free Glucose in Serum
Authors: Kennedy Allan Laurence , Mehl Homas D , Merimee Thomas J
Published: September 2013

The presence of free glucose in serum was found to interfere with accurate measurement, by a colorimetric method, of nonenzymatically glycosylated serum proteins. A mean elevation to 241% of basal levels was observed in the serum of 11 nondiabetic subjects to which glucose, in a concentration of 300 mg/dl, had been added immediately before assay. After dialysis of serum samples to remove glucose, levels of nonenzymatically glycosylated serum proteins were 0.27 ± 0.11 and 0.79 ± 0.24 nmol 5-hydroxymethylfurfural/mg protein (mean ± SD), respectively, in 57 nondiabetic and 62 type I diabetic subjects. Levels observed before dialysis of serum were approximately two to three times higher. These studies indicate that removal of free sugar from serum is necessary for accurate measurement of glycosylated protein by thecolorimetric method, and this can be achieved by overnight dialysis of serum against normal saline.
Collagen Metabolism in the Myocardium from Streptozotocin-diabetic Rats
Authors: Modrak John
Published: September 2013

The effect of diabetes on myocardial collagen metabolism was investigated in rats. Diabetes was induced by an i.v. injection of streptozotocin (60 mg/kg). The animals were killed after 3, 6, 18, or 26 wk of diabetes. Urinary glucose levels were measured by Tes-Tape. The myocardium was quickly removed and homogenized. Collagen was estimated by hydroxyproline content. Collagen synthesis was measured by 14C-proline incorporation. Myocardial collagen concentration and collagen synthesis was not altered in the diabetic rats. Collagen concentration did appear to increase in the 18- and 26-wk rats, but this was probably an age-related phenomenon. These results suggest that diabetes does not alter collagen metabolism in rats.
Insulin Binding and Response in Erythroblastic Leukemic Cells
Authors: Galbraith Richard A , Wise Curtis , Buse Maria G
Published: September 2013

Mammalian erythrocytes have been shown to bind, but not to respond to, physiologic doses of insulin. Insulin binding was studied in normal rat erythrocytes and erythroblastic leukemie (EBL) cells by standard 125I-insulin competitive binding assays. EBL cells exhibited marked insulin degradation, which was time, temperature, and concentration dependent and was mediated by both cell-bound and soluble enzymes. Bacitracin or bovine serum albumin was used to inhibit such degradation in a dose-dependent fashion to allow meaningful data analysis. Insulin binding studies showed a greater than 10-fold increase of specific binding to EBL cells compared with erythrocytes. Scatchard analysis was consistent with increases predominantly in the number of receptors on EBL cells. Concordant with increased insulin binding, EBL cells demonstrated increased transport of α-aminoisobutyric acid and increased incorporation of uridine into ribonucleic acid in response to physiologic doses of insulin (100 μU/ml). It can be concluded that EBL cells may serve as useful models of erythroblasts to explore the relationships between insulin binding, response, and cell maturation.
Dissociation Between Plasma Triglyceride Concentration and Tissue Lipoprotein Lipase Deficiency in Insulin-deficient Rats
Authors: Chen Yii-Der I , Howard Julie , Huang Virginia , Kraemer Fredric B , Reaven Gerald M
Published: September 2013

Insulin deficiency was produced by streptozotocin in young (5–6 wk old) male rats, and measurements were made of plasma triglyceride and glucose concentrations and of lipoprotein lipase (LPL) activity of adipose tissue (epididymal) and muscle (gastrocne-mius and soleus). Rats with streptozotocin-induced diabetes underwent a significant reduction in adipose tissue LPL activity (both total and heparin releasable), but the fall in LPL activity in these rats bore little relationship to their rise in plasma triglyceride concentration. Furthermore, muscle LPL activity was essentially unchanged in diabetic rats. Qualitatively similar changes were observed when measurements were made at either 8 a.m. (after the normal evening access to food) or 2 p.m. (6 h after food withdrawal). It is concluded that the hypertriglyceridemia that occurs secondary to insulin deficiency is not a simple function of decreased tissue LPL activity.
Changes in Nerve Conduction Velocity After Six Weeks of Glucoregulation With Portable Insulin Infusion Pumps
Authors: Pietri Angel , Ehle Albert L , Raskin Philip
Published: September 2013

Near normal glucoregulation was maintained in 10 patients with insulin-dependent (type I) diabetes mellitus for 6 wk with preprogrammed continuous subcutaneous insulin infusion using a portable battery-powered infusion pump (CSII). This form of therapy resulted in a statistically significant increase in motor nerve conduction velocity in the median and peroneal nerves compared with baseline values. There was no significant change in the motor nerve conduction velocity in the ulnar nerve or in the sensory nerve conduction studies. No changes occurred in five additional patients studied in similar fashion while on a conventional insulin regimen. These results suggest that the prevention of sustained hyperglycemia with CSII could theoretically result in the prevention of diabetic neuropathy. However, only long-term studies of CSII will provide the information necessary to determine the clinical relevance of the findings.
The Effect of Insulin and of Anti-insulin Serum on Handling of Sodium by the Isolated, Perfused Kidney of the Streptozotocin-diabetic Rat
Authors: Rostand Stephen G , Watkins Joyce B , Clements Rex S
Published: September 2013

We studied the effects of insulin and of anti-insulin serum (AIS) on sodium excretion in isolated diabetic and normal, fed, rat kidneys perfused at a constant pressure of 100 mm Hg with Krebs-Ringer bicarbonate buffer containing 7.5% bovine serum albumin and 5 m M glucose and gassed with 95% O2 and 5% CO2. The addition of insulin produced antinatriuresis in kidneys from diabetic rats but not in those from normal rats. Moreover, before the addition of insulin, the baseline rate of sodium excretion was greater in the diabetic rat kidney than in the normal rat kidney, while no differences in glomerular filtration rate (GFR) were observed. The addition of AIS (enough to bind 1.4 mU of insulin per milliliter of perfusate) produced a brisk natriuresis in kidneys from normal rats, and this natriuresis could be reversed by the addition of saturating amounts of insulin. By contrast, the addition of normal guinea pig serum to the perfusate of normal kidneys or AIS to the perfusate of diabetic kidneys was unassociated with natriuresis. We conclude that (a) insulin promoted antinatriuresis in diabetic but not in normal rat kidneys, (b) AIS promoted natriuresis in normal kidneys, and (c) this natriuresis was a specific effect of AIS and was reversed by the addition of insulin. Our data support the view that insulin-induced antinatriuresis depends on endogenous levels of circulating insulin and on the availability of insulin-binding sites in the kidney.
Effects of Glucose and Amino Acids on the Biosynthesis of DNA and Insulin in Fetal Rat Islets Maintained in Tissue Culture
Authors: Swenne Ingemar , Bone Adrian J , Howell Simon L , Hellerström Claes
Published: September 2013

Islet growth and insulin biosynthesis in fetal rat islets have been studied in a recently developed in vitro culture system. Collagenase-digested pancreases of 22-day-old rat fetuses were maintained for 5 days in culture medium RPMI 1640 (11.1 mM glucose) to allow degeneration of acinar tissue. Intact pure islets, composed of more than 90% B-cells, were then collected and subsequently treated for 3 days in culture with different concentrations of glucose and amino acids. Islets were then labeled for 24 h with 3H-thymidine (DNA synthesis) or for 2 h with 3H-phenylalanine (insulin/protein biosynthesis). The specificity of the incorporation of 3H-thymidine into B-cell DNA has been investigated and characterized by both Chromatographie and autoradiographic studies. Moreover, using a colchicine metaphase arrest technique, a relationship was demonstrated between incorporation of labeled thymidine and mitotic incidence in the B-cells.

Incorporation of 3H-thymidine was significantly higher in the fetal than in the adult islets following treatment in all the different concentrations of glucose and amino acids studied. In the presence of 10% fetal calf serum, however, there was no increase in DNA synthesis in the fetal islets in response to high concentrations of glucose or amino acids. In media containing 2.5% fetal calf serum, there was a significant increase in the rates of DNA synthesis after addition of high concentrations of either glucose or amino acids.

Fetal islets showed a marked insulin biosynthetic response to an acute glucose challenge. Both the basal and stimulated rates of insulin biosynthesis were increased after treatment in high concentrations of glucose but not after treatment with amino acids. The results suggest an important role of nutrients in the regulation of B-cell growth and insulin biosynthesis and also a dissociation between the regulatory mechanisms of these two processes.
Temporal Relationship of Tissue Somatostatin-like Immunoreactivity to Metabolic Changes in Genetically Obese and Diabetic Mice
Authors: Berelowitz Michael , Coleman Douglas L , Frohman Lawrence A
Published: September 2013

Somatostatin-like immunoreactivity (SRIF-LI) content in 2 N acetic acid extracts of hypothalamus, gastric antrum, and pancreas was measured in genetically obese (C57BL/6J ob/ob and db/db) and diabetic (C57BL/KsJ db/db and ob/ob) mice and normal litter-mate controls from 5 to 24 wk to determine the relationship of previously reported changes to the development of metabolic abnormalities. Hypothalamic SRIF-LI concentration was similar in control, diabetic, and obese mice at all ages and increased progressively with age in all groups. Gastric antrum SRIF-LI was similar in all groups of mice at all ages. Obese mice gained weight progressively and showed moderate hyperglycemia and marked hyperinsulinemia from 5 wk of age. Pancreatic SRIF-LI content in obese (C57BL/6J) animals was similar to that in lean litter-mate controls, but pancreatic SRIF-LI concentration (expressed by weight or protein content) was decreased until 18 (6J ob/ob) and 10 (6J db/db) wk. Diabetic (C57BL/KsJ) mice showed a similar metabolic pattern until 10 wk with no change in pancreatic SRIF-LI content or concentration. Thereafter a progressive fall in serum insulin and a marked rise in serum glucose was associated with increasing pancreatic SRIF-LI content and concentration. These studies suggest that the genetically hyperphagic syndromes are unassociated with any change in hypothalamic or gastric SRIF-LI; that pancreatic SRIF-LI increases occur in response to, rather than as the cause of, relative hypoinsulinemia; and that the genetic background of the mice (KsJ or 6J) rather than the mutant gene (db or ob) determines the defect in carbohydrate metabolism and the pancreatic SRIF-LI response.
Reversal of the Enhanced Somatostatin Release from the Isolated, Perfused Diabetic Rat Pancreas after the Amelioration of Diabetes by Whole Pancreas Transplantation
Authors: Kadowaki Seizo , Taminato Tomohiko , Chiba Tsutomu , Goto Yasuo , Nozawa Masumi , Seino Yutaka , Matsukura Shigeru , Fujita Takuo
Published: September 2013

To investigate the pancreatic endocrine function in streptozotocin-diabetic rats before and after the whole pancreas was transplanted, pancreatic insulin, gluca-gon, and somatostatin content and release were measured.

Highly inbred Lewis male rats were divided into the following three groups: normal control rats; streptozo-tocin-induced-diabetic rats; and streptozotocin-diabetic rats transplanted with whole pancreas. Studies in vivo revealed normalization of elevated blood glucose and marked improvement of the impaired arginine-induced plasma insulin release by the transplantation of a healthy pancreas into the diabetic rats. Neither basal nor arginine-induced plasma glucagon levels in the diabetic rats were significantly different from the normal group, but significantly higher plasma glucagon levels were found in the transplanted rats. Studies in vitro using the isolated perfused rat pancreas revealed a significant increase of somatostatin release from the diabetic pancreas, with a marked reduction of insulin release and almost normal glucagon release. In the transplanted rats, on the other hand, arginine-induced somatostatin release from the host pancreas was reduced to normal without a significant change in insulin or glucagon release. In addition, the endocrine function of the graft remained normal in the transplanted rats.

Pancreatic somatostatin release, thus, appears to be affected by changes in circulating insulin, since pancreatic transplantation effectively corrects the circulating insulin level.
Body Weight, Skeletal Muscle Morphology, and Enzyme Activities in Relation to Fasting Serum Insulin Concentration and Glucose Tolerance in 48-year-old Men
Authors: Lithell H , Lindgärde F , Hellsing K , Lundqvist G , Nygaard E , Vessby B , Saltin B
Published: September 2013

Tissue samples were taken from the gastrocnemius muscle of 26 randomly selected, glucose-tolerant, 48-yr-old men. Hexokinase, phosphorylase, lactate dehy-drogenase (LDH), succinate dehydrogenase, and lipo-protein lipase activity (LPLA), as well as the area per fiber type and capillary density, were determined.

Mean fiber area correlated positively with relative body weight (r = 0.53, P < 0.01), but capillary density did not. The result is that, in cases of high body weight, each capillary supplies a larger muscle fiber area.

Serum insulin concentration in the fasting state correlated positively with body weight (r = 0.77, P < 0.001) and with mean fiber area per capillary (r = 0.87; P < 0.001). Only during the latter part of an oral glucose tolerance test (OGTT) did blood glucose concentrations correlate with relative body weight and mean fiber area per capillary (r = 0.42, r = 0.51, P < 0.05). A stepwise multiple regression analysis showed that the different muscle morphology measurements could account for % of the variation in the fasting serum Insulin concentration, the fasting insulin/glucose ratio, and the blood glucose concentration at 120 min in the OGTT. Of the intracellular enzymes, only LDH (r = -0.71, P < 0.001) correlated with the mean fiber area per capillary.

LPLA correlated with capillary density (r = 0.66, P < 0.001), and, along with the muscle morphology measurements, could account for ¾ of the variation in serum triglyceride concentrations.

The results show that a large mean muscle fiber area/capillary ratio indicates a morphologic imbalance, which is related to both glucose tolerance and various degrees of insulin sensitivity.
Metabolic Characteristics of Renal Insulin Uptake
Authors: Rabkin Ralph , Gottheiner Toby I , Tsao Tan-Sheng
Published: September 2013

The kidney removes insulin from the circulation by glomerular and peritubular clearance, thus exposing both the luminal and the contraluminal tubular surfaces to the hormone. In the proximal tubule, filtered insulin is internalized by means of pinocytosis and eventually localizes in the lysosomes. The fate of insulin removed from the peritubular circulation is less well understood. We previously reported that tubular uptake of insulin is a temperature-sensitive process consisting of two different systems. The system associated with the luminal aspect of the cell appears to be dependent on oxidative metabolism, whereas the system associated with the contraluminal aspect of the cell appears to be independent thereof. To further characterize renal tubular insulin uptake, a study was performed with (OCI) was 1336 ± 51 μl/min (SEM), glomerular filtration rate (GFR) was 1019 ± 54 fillmin, peritubular clearance of insulin was 317 ± 56 μl/min, fractional excretion of sodium (FEna) was 5.1 ± 0.7%, and fractional excretion of insulin (FEi) was 1.3 ± 0.6%. Addition of inhibitors of glycolysis (iodoacetate), Na-K-ATPase (ouabain), and lysosomal degradation (chloroquine) was associated with a significant increase of Fei (4.7 ± 0.4%, 6.4 ± 1.4%, and 13.9 ± 2.4%, respectively) and of FEna (28%, 43%, and 12%, respectively). By contrast acetazolamide with furosemide increased FEna to 39%, without altering FEi. Analysis of all data revealed no correlation between FEna and FEi, indicating that tubular insulin absorption is at least partly independent of sodium reabsorption. None of the inhibitors altered OCI, GFR, or peritubular clearance significantly. The above observations, together with our previously reported data, indicate that luminal insulin uptake is a temperature and chloroquine sensitive process that is dependent on metabolic energy. The source of this energy appears to be ATP, which is derived from oxidative metabolism and glycolysis. These characteristics, consistent with a process involving pinocytosis and lysosomal degradation, were not features of the contraluminal insulin uptake process.
Triphenyltin Fluoride (TPTF) as a Diabetogenic Agent: TPTF Induces Diabetic Lipemia by Inhibiting Insulin Secretion from Morphologically Intact Rabbit B-Cell
Authors: Manabe Shigeo , Wada Osamu
Published: September 2013

Recent work in our laboratory has shown that oral administration of triphenyltin fluoride (TPTF) evokes hypertriglyceridemia in rabbits. The present experiments were conducted to elucidate the mechanism of TPTFinduced hypertriglyceridemia in rabbits by a combined biochemical and ultrastructural approach. After a single TPTF administration, fasting blood glucose and plasma triglyceride levels increased significantly (P < 0.02) for about 10 days. On the other hand, both plasma and adipose tissue lipoprotein lipase (LPL) activity was markedly decreased (P < 0.001) during this period, and triglyceride production rates on day 2 after TPTF administration were significantly decreased (P < 0.01). Density-gradient ultracentrifugation showed a remarkable accumulation of chylomicron and VLDL in the composition of plasma lipoproteins. Insulin injection to the hypertriglyceridemic rabbits induced a significant recovery of the decreased plasma LPL activity with a concomitant decrease of plasma triglyceride levels, while abeyance of insulin injection resulted in a decrease of LPL activity again. A significant inhibition of insulin release in response to the loading of glucose, glucagon, or arginine was observed in the TPTF rabbits (P < 0.02). Inhibition of glucagon release was also observed in the arginine-loading test (P < 0.01). Electron microscopic studies showed small abnormalities in the pancreatic islets of TPTF-treated rabbits. These findings suggest that TPTF inhibits insulin release from rabbit islets, subsequently inducing diabetic lipemia due to the insulin deficiency. Furthermore, it is possible to provide a new animal model for diabetes and diabetic lipemia by administration of TPTF to rabbits.
Hormones and Factors That Stimulate Growth of a Rat Islet Tumor Cell Line in Serum-Free Medium
Authors: Fong Henry K W , Chick William L , Sato Gordon H
Published: September 2013

A number of hormones and factors were found to stimulate growth of cultured rat islet tumor cells, the RIN-r cell line. A serum-free supplemented medium for RIN-r cells was formulated. It consisted of a 1:1 mixture of Ham's F12 and DME media with the addition of insulin, transferrin, triiodothyronine, prolactin, growth hormone, and an extract of proteose peptone (medium IM). The growth rate of RIN-r cells in this medium is as great as it is in 10% serum-supplemented medium. Up to 10 population doublings occurred over a period of 20 days. Insulin is a very effective mitogen for RIN-r cells and has an effect on concentrations as low as 30 ng/ml. In addition, the insulin-like somatomedin, multiplication stimulating activity (MSA), is a growth factor at 50 ng/ml. It was found that RIN-r cells proliferate and continue to produce immunoreactive insulin in a hormonally and nutritionally defined medium. This medium is derived from medium IM, in which insulin is replaced with MSA and proteose peptone is omitted. Variations of this medium may prove useful in studies on the growth and function of normal islets in long-term primary culture.
Increased Platelet Arachidonic Acid Metabolism in Diabetes Mellitus
Authors: Halushka P V , Mayfield R , Wohltmann H J , Rogers R C , Goldberg A K , McCoy S A , Loadholt C B , Colwell J A
Published: September 2013

Platelets obtained from some diabetic patients show enhanced in vitro platelet aggregation. This study sought to determine if platelets obtained from insulindependent diabetic subjects synthesize increased quantities of the labile aggregating substance, thromboxane A2 (TXA2), and if it may play a role in the enhanced platelet aggregation. Arachidonic acid (1 mM)-stimulated TXA2 synthesis, as determined via radioimmunoassay of its stable metabolite TXB2, was significantly greater (P < 0.01, N = 12) in platelet-rich plasma obtained from diabetics compared with matched controls. Arachidonic acid-stimulated TXB2 synthesis in the diabetic platelet-rich plasma was positively correlated with the ambient fasting plasma glucose (r = 0.61, P < 0.02, N = 15). Platelet aggregation induced by arachidonic acid (0.4–0.8 mM) was inhibited significantly less by 13-azaprostanoic acid (P < 0.04, N = 14), a competitive antagonist of the actions of prostaglandin H2 or TXA2 on platelets, compared with matched controls. The results support the notion that platelets obtained from some insulin-dependent diabetic subjects manifest increased synthesis of TXA2, which may contribute to the enhanced platelet aggregation.